theevolutionofdevelopmentinstreptomyces ......which development is initiated, or to changes in...
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Theevolutionofdevelopment inStreptomyces analysedbygenomecomparisonsKeith F. Chater & Govind Chandra
John Innes Centre, Norwich Research Park, Colney, Norwich, UK
Correspondence: Keith F. Chater,
Department of Molecular Microbiology, John
Innes Centre, Norwich Research Park, Colney,
Norwich NR4 7UH, UK. Tel.: 144 1603
450297; fax: 144 1603 450778; e-mail:
Received 21 February 2006; revised 10 April
2006; accepted 12 April 2006.
First published online 22 June 2006.
DOI:10.1111/j.1574-6976.2006.00033.x
Editor: Simon Cutting
Keywords
comparative genomics; evolution of
development; sporulation; actinobacteria;
Streptomyces .
Abstract
There is considerable information about the genetic control of the processes by
which mycelial Streptomyces bacteria form spore-bearing aerial hyphae. The recent
acquisition of genome sequences for 16 species of actinobacteria, including two
streptomycetes, makes it possible to try to reconstruct the evolution of Strepto-
myces differentiation by a comparative genomic approach, and to place the results
in the context of current views on the evolution of bacteria. Most of the
developmental genes evaluated are found only in actinobacteria that form
sporulating aerial hyphae, with several being peculiar to streptomycetes. Only four
(whiA, whiB, whiD, crgA) are generally present in nondifferentiating actinobacter-
ia, and only two (whiA, whiG) are found in other bacteria, where they are
widespread. Thus, the evolution of Streptomyces development has probably
involved the stepwise acquisition of laterally transferred DNA, each successive
acquisition giving rise either to regulatory changes that affect the conditions under
which development is initiated, or to changes in cellular structure or morphology.
Introduction: morphology, ecology,phylogeny and evolutionary history ofstreptomycetes
Streptomycetes are among the most complex of bacteria
(Chater & Losick, 1997). They grow, usually in soil, as
branching thread-like hyphae to form a vegetative or
substrate mycelium. This growth habit, coupled with the
activities of an abundance of extracellular hydrolytic en-
zymes, helps them to gain access to the nutrients locked up
in the insoluble polymers of soil. As the substrate mycelium
becomes nutrient-limited, it goes on to manifest two
properties for which streptomycetes are particularly famous:
the production of antibiotics and other secondary metabo-
lites; and the formation of reproductive aerial branches,
which are at least partially parasitic on the mycelium from
which they emerge. The aerial hyphae form aseptate tip
compartments up to c. 100 mm long, often containing many
tens of copies of the genome. When such a compartment
stops growing, it undergoes synchronous multiple septation
to give a string of unigenomic prespore compartments. The
cell walls of prespore compartments undergo thickening and
rounding up, and usually accumulate spore pigment, as they
turn into chains of spores. Unlike the highly resistant
endospores of bacilli, Streptomyces spores do not show
conspicuous resistance to heat, but they do survive well in
including dry conditions, and they presumably facilitate
dispersal by wind and water, and via animals. Although
most streptomycetes do not seem to sporulate in submerged
culture, a few can (including some strains of Streptomyces
griseus and Streptomyces venezuelae). Presumably, differ-
ences in the detailed developmental regulatory circuitry are
involved in this departure from the norm, but at the time of
writing, there is insufficient published information for us to
consider this interesting question here.
Phylogenetically, streptomycetes are part of the Actino-
bacteria, the class of gram-positive and morphologically
diverse bacteria that have DNA comparatively rich in G1C.
Among free-living actinobacteria, G1C content ranges from
54% in some corynebacteria to more than 70% in strepto-
mycetes, but in the obligate pathogen Tropheryma whipplei it
is less than 50%. Figure 1 is a phylogenetic tree of sequenced
actinobacteria based on 16S rRNA gene sequence compar-
isons. It appears that the last common ancestor of the
actinobacteria lived about 2–1.5 Ga, when evidence suggests
that the atmosphere first began to be oxygenated (Embley &
Stackebrandt, 1994). Streptomycetes are believed to have
originated about 440 Ma, apparently immediately following
the invasion of the land by green plants, and coinciding with
a rapid rise of oxygen in the atmosphere towards present
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
levels (Heckman et al., 2001). Reflecting this timing, strep-
tomycetes are particularly well adapted to the utilization of
refractory plant remains, are aerobic, and have a great
number of genes and systems associated with oxidative
stress (Lee et al., 2005). Earthworms and arthropods such
as springtails that consume decaying plant remains, includ-
ing the associated microorganisms, appeared soon after the
first land plants, so it is quite possible that they exerted other
major selective pressures on Streptomyces evolution. For
example, the frequent passage of most soil through earth-
worms (Darwin, 1881) must have selected for resistance of
Streptomyces spores to the chemically and enzymically harsh
conditions in worm guts. The frequent consumption of
small soil animals by vertebrates (such as reptiles, small
mammals and birds) in more recent evolutionary time,
while it may have contributed to the dispersal and diversi-
fication of streptomycetes, could not have been a funda-
mental element in their early evolution.
Comparative genomics of actinobacteria:the bigger picture
In the last few years, genome sequences have been obtained
for diverse actinobacteria (19 complete and annotated
genomes, 16 species, 10 genera at the time of writing), as
indicated in Table 1. The analysis in this review is largely
confined to these sequenced genomes, about which more
comprehensive statements can be made. Of the two se-
quences from streptomycetes, one is from the academic
model organism Streptomyces coelicolor A3(2) (note that this
strain is improperly named, being more similar to the
Streptomyces violaceoruber clade; but the name is too deeply
enshrined in the literature to justify making the correction),
and the other is from Streptomyces avermitilis, an important
antibiotic producer (Bentley et al., 2002; Ikeda et al., 2003).
These two species are phylogenetically fairly well separated
from each other and from Streptomyces griseus, another
species that has been important in the study of development,
and whose genome sequence is currently being determined
(S. Horinouchi, pers. commun.). The genome sequence of
another species, the plant pathogen Streptomyces scabies,
is also available, though currently not annotated (ftp://
ftp.sanger.ac.uk/pub/pathogens/ssc/Ssc.dbs). This makes it
possible to use comparative genomics to attempt to trace the
evolution of Streptomyces development, and in doing this,
perhaps to deepen our understanding of developmental
mechanisms.
For such an approach, criteria must first be adopted for
whether related genes can be considered to be orthologues
(i.e. are the direct, functionally equivalent derivatives of a
gene present in the last common ancestor of the species
being compared) or are merely paralogues (related through
ancestral gene duplication). A first criterion is sequence
relatedness between gene products. One would expect most
(though not all) orthologous genes to show a fairly consis-
tent degree of amino-acid identity in comparisons between
particular organisms. This ‘orthology value’ could be deter-
mined by initially making assumptions about obvious
candidates for orthology, such as genes for RNA polymerase
core enzyme subunits, or genes for the biosynthesis of
amino acids etc. (as long as such candidate genes are
represented only once in the genomes). We compared seven
such gene products among S. coelicolor, S. avermitilis,
Thermobifida fusca, Corynebacterium glutamicum and My-
cobacterium tuberculosis (Table 2). Within this set of
orthologues, comparisons between the two streptomycetes
indicated a range of 88–97% amino-acid identity, while
identities between S. coelicolor and the other organisms
ranged from 69–82% (T. fusca) to 52–71% (C. glutamicum).
On the other hand, when sets of multiple paralogues
are encoded within one organism, members of each
set are generally well under 50% identical. We therefore
used 50% as a threshold value when considering
orthology in the developmental genes discussed in this
article.
The second criterion for orthology takes advantage of
similarity of gene arrangement along the chromosome
(‘synteny’), which is usually extensive between members of
the same genus. Across different genera of actinobacteria
there are many patches of local synteny, and there is a
0.10
100 Mtb
MboMav
MleNfa
CdiCefCgxCgl
Pac
TfuSco
Sav
BloSth
Bsu
100100
100
100
1009970
97
75100
100
7987
98
LxyTwaTwx
Fig. 1. Phylogeny of 16S rRNA gene of sequenced actinobacterial
genomes. The analysis used the Phylip package. Abbreviations are as
follows: Mtb, Mycobacterium tuberculosis; Mbo, Mycobacterium bovis;
Mav, Mycobacterium avium; Mle, Mycobacterium leprae; Nfa, Nocardia
farcinica; Cdi, Corynebacterium diphtheriae; Cef, Corynebacterium effi-
ciens; Cgx, Cgl, Corynebacterium glutamicum; Tfu, Thermobifida fusca;
Pac, Propionibacterium acnes; Sco, Streptomyces coelicolor; Sav, Strep-
tomyces avermitilis; Lxy, Leifsonia xyli; Twa, Twx, Tropheryma whipplei;
Blo, Bifidobacterium longum; Sth, Symbiobacterium thermophilum; Bsu,
Bacillus subtilis (outgroup).
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
652 K.F. Chater & G. Chandra
Table 1. Sequenced actinobacterial genomes consulted in this article
Organism
Morphology,
special features Accession number Database Genome attributes
Streptomyces
coelicolor
Mycelial, spore
chains on aerial
hyphae
AL645882 http://streptomyces.org.uk 8.66 Mb, 72.1% GC,
7825 genes,
146 (2%) with TTA
Streptomyces
avermitilis
Mycelial, spore
chains on aerial
hyphae
BA000030 http://www.bio.nite.go.jp/dogan/
MicroTop?GENOME_ID=sav_G1
9.02 Mb, 70.7% GC,
7575 genes,
260 (3%) with TTA
Thermobifida
fusca
Mycelial, spores
clumped on aerial
hyphae
CP000088 http://genome.jgi-psf.org/
draft_microbes/thefu/
thefu.home.html
3.64 Mb, 67.95% GC,
3419 genes,
449 (13%) with TTA
Nocardia farcinica Mycelial, substrate
and aerial hyphae
form fragments
AP006618 http://nocardia.nih.go.jp/ 6.02 Mb, 70.8% GC,
5683 genes,
242 (4%) with TTA
Mycobacterium
avium
Irregular rods AE016958 http://www.cbc.umn.edu/
ResearchProjects/AGAC/Mptb/
Mptbhome.html
4.83 Mb, 69.2% GC,
4350 genes
589 (14%) with TTA
Mycobacterium
bovis
Irregular rods BX248333 http://www.sanger.ac.uk/Projects/
M_bovis/
4.35 Mb, 65.6% GC,
3953 genes
1442 (36%) with TTA
Mycobacterium
tuberculosis
Irregular rods AL123456
AE000516
http://www.sanger.ac.uk/Projects/
M_tuberculosis/
http://www.tigr.org/tigr-scripts/
CMR2/
GenomePage3.spl?database=gmt
4.41 Mb, 65.6% GC,
3999/4189 genes
1462/1489 (36%) with TTA
Leifsonia xyli Rods AE016822 http://leifsonia.lbi.ic.unicamp.br 2.58 Mb, 67.6% GC,
2030 genes
109 (5%) with TTA
Bifidobacterium
longum
Irregular rods,
sometimes
branched
AE014295 2.26 Mb, 60.1% GC,
1727 genes
450 (26%) with TTA
Corynebacterium
efficiens
Rods BA000035 http://www.bio.nite.go.jp/dogan/
MicroTop?GENOME_ID=ce__G1
3.15 Mb, 63.1% GC,
2942 genes
717 (24%) with TTA
Corynebacterium
glutamicum
Rods BA000036
BX927147
http://genome.ls.kitasato-u.ac.jp/
esequenced.html
http://www.uni-bielefeld.de/
3.31/3.28 Mb, 53.8% GC,
3099/3058 genes
1942/1885 (62%) with TTA
Corynebacterium
diphtheriae
Rods BX248353 http://www.sanger.ac.uk/Projects/
C_diphtheriae/
2.49 Mb, 53.4% GC,
2320 genes
1535 (66%) with TTA
Propionibacterium
acnes
Pleomorphic rods AE017283 http://www.g2l.bio.uni-
goettingen.de/projects/
f_projects.html
2.56 Mb, 60.0% GC, 2297
genes
1249 (53%) with TTA
Symbiobacterium
thermophilum
Obligate symbiont;
possibly not
actinobacterial
AP006840 http://genome.ls.kitasato-u.ac.jp/
esequenced.html
3.57 Mb, 68.6% GC,
3337 genes
272 (8%) with TTA
Tropheryma
whipplei
Obligate pathogen BX072543
AE014184
http://www.sanger.ac.uk/Projects/
T_whipplei/
http://www.genoscope.cns.fr/
externe/English/Projets/
projets.html
0.93 Mb, 46.3% GC,
784/808 genes
687/718 (88%) with TTA
Mycobacterium
leprae
Obligate pathogen AL450380 http://www.sanger.ac.uk/Projects/
M_leprae/
3.27 Mb, 57.7% GC,
2720 genes
1972 (73%) with TTA
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
653Evolution of Streptomyces development
recognizable tendency for a substantial number of genes to
retain a similar relative distance from the replication origin,
even though the overall organization of the chromosomes
has usually undergone many rearrangements (Fig. 2). The
rearrangements mostly involve deletions, insertion of newly
acquired DNA, and more or less symmetrical exchanges
from opposite sides in relation to the replication origin
(Bentley et al., 2002).
Table 2. Percentage amino-acid identity of representative gene products from four actinobacteria to their orthologues in Streptomyces coelicolor
Gene or enzyme
Streptomyces avermitilis Mycobacterium tuberculosis Corynebacterium glutamicum Thermobifida fusca
(70.7% GC) (65.6% GC) (53.8% GC) (70% GC)
Amino-acid biosynthesis
hisA 96.2 68.2 59.9 74.3
hisB 99.0 57.4 52.5 75.8
leuB 93.1 67.8 67.8 69
Intermediary metabolism
Aconitase 92.7 70.4 68.3 78.5
Malate dehydrogenase 94.8 70.8 64.6 82
Transcription
rpoB 95.1 75.3 71.2 80.6
rpoC 96.8 71.6 63.6 78.2
Mean (‘orthology value’) 95.4 68.8 64.0 77.0
S. coelicolor
S. coelicolor
S. coelicolor
S. coelicolor vs S. avermitilis S. coelicolor vs M. tuberculosis
`S. a
verm
itilis
T. fu
sca
S. a
verm
itilis
T. fu
sca
T. fu
sca
M. t
uber
culo
sis
M. tuberculosis
8000
7000
6000
5000
4000
3000
2000
1000
0
3500
3000
2500
2000
1500
1000
500
0
3500
3000
2500
2000
1500
1000
500
0
3500
4000
3000
2500
2000
1500
1000
500
00 1000 2000 3000 4000 5000 6000 7000 8000
0 1000 2000 3000 4000 5000 6000 7000 8000 0 500 1000 1500 2000 2500 3000 3500 4000
0 1000 2000 3000 4000 5000 6000 7000 8000
S. coelicolor
S. coelicolor
S. coelicolor vs T. fusca M. tuberculosis vs T. fusca
M. tuberculosis
S. coelicolor
Fig. 2. Plots of synteny between some actinobacterial chromosomes. The points represent positions of genes whose products reciprocally show closest
relatedness in Blast comparisons between the chromosomes. A simple diagonal line would indicate fully conserved gene arrangement in plots
comparing chromosomes aligned from a comparable point (i.e. the end of linear chromosomes, which have an roughly centrally located replication
origin, in the case of Streptomyces avermitilis against Streptomyces coelicolor; and the origin of replication of circular chromosomes, in the case of
Mycobacterium tuberculosis against Thermobifida fusca). The extent to which the diagonal becomes a cross reflects the occurrence of symmetrical
exchanges on either side of the replication origin. When circular chromosomes are compared with linear ones, synteny is expected to give inflected
plots, because the coordinates of the chromosomes start from different positions. Symmetrical exchanges on either side of the replication origin are
then expected to give a diamond-shaped plot.
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
654 K.F. Chater & G. Chandra
Most of the genes that are conserved between streptomy-
cetes and other genera are located in a central core region of
the linear Streptomyces chromosomes. Thus, of the order of
1–2 Mb at each end consists mostly of laterally transferred,
comparatively recently acquired, genes. The core regions of
the S. coelicolor and S. avermitilis chromosomes show
remarkable synteny, involving some 4000 genes. Only a very
small number of exchanges between chromosome arms have
taken place, all more or less symmetrically in relation to the
origin of replication (Ikeda et al., 2003). The synteny is
punctuated by about 2000 nonhomologous genes. This
indicates that the two genomes have been repeatedly ex-
posed to foreign DNA over evolutionary time, with the cores
acquiring new genes or losing old ones on average several
times in every million years in the course of the estimated
220 Myr since the last ancestor common to the two species
(A. Ward, pers. commun.). Nevertheless, despite the numer-
ous insertions and deletions that separate the species, the
conserved gene set has almost entirely retained its organiza-
tion. Some powerful selection must exist to maintain this, so
it is interesting that there are marked differences in genome
organization between different genera. Possibly, new genera
arise initially in response to dramatic or cataclysmic novel
selective circumstances (such as were briefly discussed in the
previous section), when some major rearrangement
of the genome confers an adaptive value that would not be
favourable in gradual adaptation to slowly changing envir-
onmental circumstances. The extensive preservation of
genomic arrangement within streptomycetes suggests that
their subsequent evolution has largely involved microadap-
tation to environments that have changed little until the
present.
Although the 1–2 Mb end regions of Streptomyces
chromosomes are evidently a major primary depot for
receiving horizontally transferred segments of DNA,
their well-documented instability is not well-suited to
long-term retention of particular genes; so those genes
whose selective benefits are repeatedly acted upon may
well find their way into the more stable central region.
There are experimental accounts of unequal exchange
between chromosome ends, and between the ends of
chromosomes and linear plasmids (Chen et al., 2002;
Wenner et al., 2003), such as might permit the effective
migration of valuable DNA towards the central region.
Once a gene has migrated inwards far enough to be
closer to the core than some other selectively advantageous
gene, it will become less susceptible to loss involving
chromosome ends. This mechanism does not preclude
other routes for chromosomal gene acquisition, particularly
those involving transposition or site-specific integration
of specialized elements such as prophages (see final section
for a discussion of the possible roles of phages in gene
acquisition).
The genetic basis of developmentalcomplexity in streptomycetes
Over several decades, some progress has been made in the
genetic dissection of the complex multicellular development
briefly outlined above, particularly in S. coelicolor and S.
griseus (Chater, 2001; Chater & Horinouchi, 2003; Flardh &
van Wezel, 2004). Several tens of genes have been identified
whose products play regulatory, catalytic, organizational or
structural roles in development, and there is a moderate
body of information about the integrated action of these
genes and gene products. Much of this information ema-
nates from the isolation of S. coelicolor mutants that fail to
complete normal development, most being defective either
in aerial growth (‘bald’, or bld, mutants) (Hopwood, 1967;
Merrick, 1976) or in the formation of mature grey spores on
the fluffy aerial mycelium (‘white’, or whi, mutants) (Hop-
wood et al., 1970; Chater, 1972). Most of these mutants are
affected in genes with regulatory character. The details of
these genes and their interactions are explored in several
recent reviews (Chater & Losick, 1997; Chater, 1998, 2001;
Chater & Horinouchi, 2003; Chater, 2006), and are only
briefly summarized here.
When grown near to each other on certain rich media,
most of an early collection of bld mutants exhibit directional
interactions that lead to aerial growth (Willey et al., 1993;
Kelemen & Buttner, 1998). These interactions have been
interpreted as a signalling cascade that is transmitted in the
sequence bldJ/bldK,L/bldAH/bldG/bldC/bldD,M/ram (Fig.
3). Although some of the more recently isolated bld mutants
do not fit in this cascade, its features have helped in the
formulation of developmental models (Chater, 2001). It has
also been shown that some mutants that fall outside the
cascade can be phenotypically corrected by pH buffering,
indicating that their morphological defects are a secondary
SapB
Chaplins, rodlins
Signal 1(oligopeptide)
Signal5
Signal2 Signal
3
Signal4
bldC(regulatory)
bldG(regulatory)
bldH(regulatory)
bldA (tRNA forrare codon
UUA)
bldK(Oligopeptidetransporter)
bldD, M(regulatory)
bldJ(unknown)
bldL(unknown)
ramStartHere!
Fig. 3. Extracellular signalling dependent on bld genes leading to the
production of surface proteins involved in aerial growth in Streptomyces
coelicolor. The figure has been updated from that of Chater (1998).
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
655Evolution of Streptomyces development
consequence of the secretion of organic acids (Viollier et al.,
2001). In other species, different extracellular interactions
leading to development have been described. One example,
the restoration of sporulation to a bald mutant of S.
alboniger by the addition of an antibiotic, pamamycin, has
recently been interpreted in terms of calcium signalling
(Danilenko et al., 2005); but the most extensively studied is
the dependence of development in some S. griseus strains on
A-factor, a member of the species-specific g-butyrolactone
signalling molecules that are often involved in switching on
antibiotic biosynthesis in streptomycetes (Horinouchi,
2002). The variety of extracellular signals used during
Streptomyces development may reflect some selective benefit
of preventing competitors from responding to each other’s
signals, and thus be a cause or reflection of speciation
(Chater & Horinouchi, 2003).
Seven of the whi loci (whiA,B,D,E,G,H,I) identified in
early studies of S. coelicolor sporulation (Chater, 1972) have
been studied further [two ‘loci’ have not: whiF99 proved to
be a whiG allele; and whiC mutants did not survive
prolonged storage (C.J. Bruton and K.F. Chater, unpub-
lished)]. Figure 4 indicates the present understanding of the
regulatory interactions of some of these genes. The most
clear-cut gene-to-gene interactions are the requirements of
the whiH and whiI promoters for an RNA polymerase
holoenzyme that contains the s factor specified by whiG.
The only known gene-to-gene interaction between the bld
and whi gene cascades is the repression of whiG by BldD
(Elliot et al., 2001).
In this article we mainly focus on the genes shown in Figs
3 and 4, because it has been possible to fit them into
interaction networks. We also include several other devel-
opmentally and morphogenetically important genes, such as
the ssgA-like gene family, mutations in some of which give a
white colony phenotype. (Keijser et al., 2003; Traag et al.,
2004; Noens et al., 2005). This anthology of genes was used
to search the genomes of actinobacteria for likely ortholo-
gues.
Genes implicated in the regulation offormation of aerial hyphae
bldA -- the determinant of the tRNA for a rarecodon
In streptomycetes, the very high GC content of the DNA
makes it inevitable that GC-rich codons are preferentially
used. As a consequence, the third position in codons is a G
or a C in about nine out of every 10 codons, providing the
basis for one of the main tools used to delineate probable
coding sequences in Streptomyces DNA sequences (Bibb
et al., 1984; Ishikawa & Hotta, 1999). In the standard genetic
code, one of six codons for leucine (UUA) has no G or C
residues. Not surprisingly, this codon is rarely used in
streptomycetes – in fact, it is by far the rarest codon in S.
coelicolor, being used in only 2% (145/7825) of chromoso-
mal genes (260, i.e. 3%, of the 7575 chromosomal genes of S.
avermitilis contain TTA codons) (Fig. 5). These observations
are of more than anecdotal interest, because the only tRNA
capable of reading UUA codons efficiently is the product of a
gene that can be completely deleted without diminishing
growth (Lawlor et al., 1987; Leskiw et al., 1991). However, as
Growth stops(glycogendeposition)
Growth of longapical compartment
Multiple septation(FtsZ, ParAB, SALPs,DNA condensation,glycogen degraded)
Rounding ofpresporecompartments,wall thickening,spore pigmentdeposition
σ WhiG
FtsZ
ParAB
GlgBI etc DNA condensation
WhiB (I) WhiB (II)
WhiA (I) WhiA (II)
WhiI (I) WhiI (II)
WhiH (I) WhiH (II)
σ F
WhiD
MaturationFig. 4. Regulation of aerial hyphal differen-
tiation into spore chains in Streptomyces
coelicolor. The regulatory scheme uses infor-
mation from Kelemen et al. (1996, 1998),
Ryding et al. (1998), Tan et al. (1998), Flardh
et al. (1999), Ainsa et al. (1999), Noens et al.
(2005) and Jakimowicz et al. (2006).
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
656 K.F. Chater & G. Chandra
indicated by this gene’s designation (bldA), such mutations
do have phenotypic effects – most obviously, they give rise
to very easily seen bald colonies that are devoid of pigmen-
ted secondary metabolites on most media (Merrick, 1976).
The complexity of this phenotype suggests that bldA may
have a regulatory role in aspects of developmental and
stationary phase biology. The bldA tRNA, with its 30-AAU-
50 anticodon, should also be able to read the alternative
leucine codon UUG via ‘wobble’ interaction, whereas a
tRNA with the 30-AAC-50 anticodon cognate with the UUG
codon is not expected to be able to translate UUA codons
(Crick, 1966). Both tRNAs are present in streptomycetes,
perhaps in part because of the need to translate UUG codons
under conditions in which translation of UUA codons
would be disadvantageous.
A bldA-like tRNA locus is present in all actinobacteria. In
the current absence of any attempts to inactivate it in any
nonstreptomycete, it is not experimentally ruled out that
UUA codons are developmentally significant beyond strep-
tomycetes. However, TTA codons are more frequent in the
genomes of other actinobacteria, though those with high GC
content generally have the lowest frequency (Table 1).
Nocardia farcinica, with 70.8% GC, has TTA codons in only
4% of genes, but these include at least one important gene of
primary metabolism (hisA), so we doubt whether the
developmental role of bldA in streptomycetes has a close
parallel in N. farcinica. Moreover, the distribution of TTA
codons within annotated N. farcinica genes shows less
positional bias than in S. coelicolor and S. avermitilis (Fig.
5), suggesting that they do not fulfil a similar role to that of
TTA codons in streptomycetes: in S. coelicolor, 22% of TTA
codons occur in the first 10 codons, compared to 9% in N.
farcinica (Fuglsang, 2005). Since bldA mutants defective in
aerial mycelium formation have been obtained in species as
diverged as S. coelicolor (Merrick, 1976), S. griseus (Kwak
et al., 1996)and S. clavuligerus (Trepanier et al., 2002), bldA
was probably adopted as an element of the developmental
biology of streptomycetes very early in their evolution.
Although the focus of this article is on morphological
development, we note in passing that the frequently ob-
served effects of bldA mutations on antibiotic production
are mostly exerted directly via UUA codons in the mRNAs of
pathway-specific regulatory genes (Chater & Horinouchi,
2003; Chater, 2006).
In a comparison between the genomes of S. coelicolor and
S. avermitilis, it turns out that only 11 orthologous genes
contain TTA codons in both species (while a further 10 TTA-
containing S. coelicolor genes have TTA-free orthologues in
S. avermitilis. Seven of those 11 have some kind of functional
annotation. One, SCO2792 (bldH), is dealt with in the next
section. The others, and their predicted functions, are:
SCO5495 (cyclic di-GMP phosphodiesterase); SCO1980
and 3423 (both abaA orfA-like, see section on whiJ below);
SCO1434 (cbxX-, cbqX-like); SCO1242 (DNA-binding pro-
tein, WhiJ-like; see below); and SCO4114 (a sporulation-
associated protein; Babcock & Kendrick, 1990). The func-
tion-unknown genes are SCO4395, 5040, 6623 and 7251.
Thus, although a small number of the TTA-containing genes
found in modern streptomycetes were already present early
in the genus, the great majority (83% in S. coelicolor) have
been acquired by lateral transfer in the comparatively recent
evolutionary past.
bldH , SCO2792 (= SAV5261), is the main targetthrough which bldA affects differentiation
One of the 11 TTA-containing genes common to S. coelicolor
and S. avermitilis is adpA, or bldH (Nguyen et al., 2003;
0.35
0.30
0.25
0.20
0.15
0.10
0.05
0.000.1 0.2 0.3 0.4 0.5 0.6
Relative position of the first TTA codon in the genes
Per
cent
of T
TA
con
tain
ing
gene
s
0.7 0.8 0.9 1.0
ScoSavNfar
Fig. 5. The relative positions of TTA codons in
TTA-containing genes of Streptomyces coelico-
lor (black), Streptomyces avermitilis (dark grey)
and Nocardia farcinica (pale grey). Positions of
TTA codons are plotted as a fraction of total
number of codons. Note that in the Strepto-
myces genes the TTA codons are much more
likely to be close to the start of the gene.
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
657Evolution of Streptomyces development
Takano et al., 2003). This gene was first studied in S. griseus
(Ohnishi et al., 1999), in which it also contains a TTA codon,
in the same position within the gene as in the other two
orthologues. In S. coelicolor, this TTA codon is the major
route through which bldA influences development, and
AdpA is apparently not involved in regulating antibiotic
production; while in S. griseus the influence of bldA on
antibiotic production is partially mediated via adpA. This
suggests that the way in which adpA is integrated into the
whole physiology of its particular host has some species-
specificity, a notion reinforced by the regulation of adpA
itself. Thus, in S. griseus, adpA expression is directly
repressed by a protein called ArpA (‘A-factor receptor
protein’), repression being lifted in response to an extra-
cellular accumulation of the g-butyrolactone A-factor, a
small membrane-diffusible signalling molecule, which dif-
fuses back into cells and interacts with ArpA. In S. coelicolor,
a comparable g-butyrolactone signalling system seems to
have no marked effects on morphological differentiation or
adpA expression (Horinouchi, 2002; Kato et al., 2002;
Takano et al., 2003; Takano et al., 2005). The differences in
regulation of adpA are possibly reflected in divergences in
upstream DNA sequences; an overall similarity between
these regions of S. coelicolor and S. griseus is interrupted at
the region known to bind ArpA and impart A-factor-
dependence in S. griseus (Takano et al., 2003; Kato et al.,
2005).
From these observations, it seems that the cascade from
bldA via adpA to differentiation is a more ancient aspect of
Streptomyces evolution than the integration of g-butyrolac-
tones into their biology. g-Butyrolactones are themselves
mainly confined to streptomycetes, among which they are
widespread. They exhibit considerable diversity, and may
well have coevolved with antibiotic biosynthetic gene sets
that have been laterally transferred among streptomycetes
(Takano et al., 2005).
In S. griseus, adpA is autoregulatory, contributing to its
own transcriptional repression (Kato et al., 2005). Four out
of six of the AdpA-binding sites of the adpA promoter of
S. griseus are conserved in the adpA promoter regions of
S. coelicolor and S. avermitilis, so autoregulation may be a
general and ancient function of AdpA (Kato et al., 2005).
Although AdpA orthologues are known only in strepto-
mycetes, AdpA belongs to a subfamily within the family of
activator/repressor proteins whose best-known member is
AraC of E. coli (Yamazaki et al., 2004; Kato et al., 2005).
AraC-like proteins are characterized by the features of a dual
helix-turn-helix DNA-binding domain typically located far
from the N-terminus. In the case of AdpA, this domain is
from aminoacids 240–332. Residues 57–195 conform to a
Pfam domain DJ-1 PfpI that is involved in dimerization
(Yamazaki et al., 2004). The rather large size of AdpA
(around 400 aa) suggests that it may respond to some other
molecule, but there are no experimental data addressing this
possibility.
Extensive studies in the laboratory of S. Horinouchi have
revealed a substantial regulon of target genes for AdpA in S.
griseus, and a consensus AdpA binding site has been defined.
Among the target genes are several for proteases and
protease inhibitors (Ohnishi et al., 2005; Kim et al., 2005).
It seems likely that one of the main ways in which AdpA
influences development is via the extracellular interactions
of proteases and protease inhibitors (Chater, 2006). Many
streptomycetes produce extracellular proteases related to
trypsin, the activity of which is held in check by binding to
small proteinaceous inhibitors, such as leupeptin (Taguchi
et al., 1993; Kim & Lee, 1995). In some cases it has been
found that specific proteolysis of the inhibitor by a second
protease is used to activate the trypsin-like protease, which
is then involved in activating development (Kim & Lee,
1995). Since AdpA-dependent extracellular protease: inhibi-
tor interactions are found in phylogenetically diverse spe-
cies, it is likely that they are an ancient feature of
streptomycetes (but among other sequenced actinobacterial
genomes, only that of T. fusca apparently encodes such a
protease inhibitor). The interactions may be important in
the reuse of the biomass of the vegetative mycelium that
provides nutrients for growth of the aerial mycelium (Man-
teca et al., 2006). It will be interesting to find out whether
these interactions have any connection with the extracellular
signalling cascade discussed earlier for S. coelicolor (Fig. 3),
and whether species-specificity is involved, as is often the
case for extracellular signalling.
Several other bld genes appear to be confinedto actinomycetes with sporulating aerialmycelium
In the first two steps in the cascade shown in Fig. 3, bldJ,
which is still uncharacterized, is required for the production
of signal 1, an extracellular oligopeptide that has been
partially characterized (Nodwell & Losick, 1998). Signal 1 is
believed to be imported by a multicomponent ATP-depen-
dent transport system specified by the genes of the bldK
locus (SCO5112–5116; Nodwell et al., 1996). The bldK locus
is absent from other actinobacterial genomes. However, the
next three bld genes ‘downsteam’ of the bldA/adpA stage in
the extracellular signalling cascade are found not only in
streptomycetes, but also in T. fusca, the only other actino-
mycete among those with known genome sequences that has
a well-developed sporulating aerial mycelium (Fig. 6). This
is surprising, in view of the rather wide apparent phyloge-
netic distance between these two genera based on 16S rRNA
gene (Fig. 1), though the comparatively high orthology
value for S. coelicolor and T. fusca (Table 2) suggests that
the two genera may be more closely related than Fig. 1
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
658 K.F. Chater & G. Chandra
indicates. There is currently no information bearing on
whether the T. fusca bld gene orthologues play a role in
aerial growth.
bldG , SCO3549 ( = SAV4614)
This gene encodes a member of a protein family found
exclusively in gram-positive bacteria, and with multiple
representatives in many of them (there are 13 in S. coelicolor)
(Bignell et al., 2000). In the characterized cases, these
proteins are known as antagonists of anti-s factors. The
interaction between the two proteins depends on the
phosphorylation state of the anti-anti-s factor. When
phosphorylated, it does not bind to the anti-s factor
efficiently, so the latter protein is free to bind to, and hold
inactive, its target s factor, whereas the unphosphorylated
anti-anti-s sequesters the anti-s, thus releasing the s factor
to interact with RNA polymerase and activate a specific set
of promoters. s factors of a particular subclass are regulated
in this way, the best characterized being sB of B. subtilis
(Chen et al., 2003).
The phosphorylation state of the anti-anti-s is deter-
mined both by the kinase activity of the anti-s factor and by
dephosphorylation by a sensor phosphatase; and a phos-
phorylated form of BldG has been demonstrated (Hesketh
et al., 2002; Bignell et al., 2003). In sporulating bacilli and
clostridia, some of these systems are involved in forespore
development (Errington, 2003) and others, notably sB, in
stress responses (Chen et al., 2003). In those cases, the bldG-
like gene is clustered with the target anti-s factor gene and
the cognate s factor determinant; there are six examples of
such clusters in the S. coelicolor genome. However, although
bldG is adjacent to a gene (SCO3548, = SAV4615) for a
probable anti-s factor, the two genes are not located close
to a recognizable s factor determinant. It remains comple-
tely unknown whether BldG is involved in regulating
activity of any of the nine s B-like proteins encoded
in the S. coelicolor genome. In view of the evidence of
interplay between stress responses and differentiation in
S. coelicolor (Chater, 2001; Kelemen et al., 2001; Viollier
et al., 2003; Lee et al., 2005), it may well be that particular sfactors are subject to regulation by more than one anti-ssystem.
The T. fusca orthologues of bldG (Tfus0984, 77% iden-
tity) and its partner gene are at one end of a region
containing 12 syntenously arranged genes. Several of those
adjacent genes (but not the bldG pair) show widespread
synteny among actinobacteria.
bldC , SCO4091 ( = SAV4130)
This gene encodes a small protein, identical in S. coelicolor
and S. avermitilis, consisting of little more than a MerR
family DNA-binding domain (Hunt et al., 2005). The bldC
noncoding upstream region is conserved for more than
400 bp between S. coelicolor and S. avermitilis (4 90%
identity, interrupted by one nonrelated segment about
310 bp upstream of the coding region in each case). The
400 bp includes (at least in S. coelicolor) 145 bp of tran-
scribed leader sequence. The length of this conserved region
is quite surprising in view of the apparently constitutive
expression of bldC (Hunt et al., 2005). Although there are
other genes encoding single-domain MerR-like proteins in
these and other genomes, none of them is closely related to
bldC except the bldC orthologue (Tfus1491, 75% aminoacid
identity) in T. fusca (Hunt et al., 2005). Tfus1491 is located
at one end of a stretch of six genes that show synteny
with a region of the S. coelicolor genome (with two
additional genes in the streptomycete). Three of these
adjacent genes are present and syntenous in most actino-
bacteria. Figure 7 shows the extent of synteny in this
genomic region, as a typical example of the relative genome
strucures of S. coelicolor, S. avermitilis, T. fusca and other
actinobacteria.
bldD , SCO1489 ( = SAV6861)
BldD is a regulatory protein that possesses a putative DNA-
binding domain similar to those of Xre-like regulators, but
bldJ BldK1
2bldA
AdpA
AdpA*
3
BldG-P
BldG:anti-σ
releasesunknown σ
4 BldC
5
BldD
BldD*
ramRRamRRamR*RamCSAB
SapB
Aerialgrowth
Alsopresent inT. fusca
Peculiar toStreptomyces
BldC*σBldN
BldM
BldM*
bldN
bldMchpA-H
chaplins
Fig. 6. Distribution among sequenced actinobacteria of gene functions
involved in the extracellular cascade for aerial mycelium formation.
Except for blbD (see text) none of the genes for the proteins in this
diagram has orthologues in any characterized genomes apart from those
of streptomycetes and Thermobifida fusca (the latter are indicated on
yellow backgrounds). Gene products remaining inside the cell are inside
coloured areas. The numbered dotted arrows correspond to the extra-
cellular signals in Fig. 3, and unbroken lines or arrows represent likely
sequential events based on the authors’ interpretation of the literature
(see text). Asterisks indicate hypothetical activated forms of proteins.
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
659Evolution of Streptomyces development
is not otherwise closely similar to any known class of
regulators (Elliot et al., 1998). It represses its own expression
and that of several other developmental genes including
bldN and whiG (Elliot & Leskiw, 1999; Elliot et al., 2001). It
has been suggested that it may play a role similar to that of
the transition-state regulators SinR and AbrB of B. subtilis
(Elliot et al., 2001). The T. fusca orthologue (Tfus1058; 77%
identity) is located at the end of an 11-gene segment
showing synteny with the bldD region of streptomycetes,
much of which (but not including bldD) is widespread
among actinobacteria. Surprisingly, an apparent orthologue
is present in Kineococcus (Deinococcus) radiotolerans, an
outlying member of the actinobacteria (Accession no.
NZ_AAEF01000025). The bldD orthologues in S. coelicolor,
S. avermitilis and S. griseus show considerable conservation
in the promoter region , including complete identity in a
sequence near the � 10 promoter element shown to bind
BldD in vitro in S. coelicolor and S. griseus (Elliot et al., 2001;
Ueda et al., 2005). There is no such similarity upstream of
the T. fusca bldD orthologue, perhaps indicating that the
gene is regulated differently.
bld genes at the downstream end of theextracellular signalling cascade are peculiar tostreptomycetes
The bldG, bldC and bldD genes, which are also found in T.
fusca (see above), may have roles in the stress responses of
mycelial organisms. Such a role was previously suggested for
Streptomyces bld genes, in a model in which the extracellular
signal cascade was considered as a succession of indications
that all possible alternative responses to hard times had been
tried before commitment to reproductive aerial growth,
which is the ‘last resort’ (Chater, 2001). However, like the
genes involved in the start of the extracellular cascade (bldK,
bldA, bldH/adpA), those at its finish (bldM, bldN, ramC-
SABR) appear to be confined to streptomycetes (Fig. 6).
Conceivably, then, they fulfil roles that are specifically
appropriate to the particular features of Streptomyces aerial
growth.
bldM , SCO4768 ( = SAV4998), and bldN , SCO3323( = SAV4735)
BldM is an aberrant orphan response regulator (Molle &
Buttner, 2000), and BldN an ECF s factor (Bibb et al., 2000).
It appears that sBldN is directly required for the transcrip-
tion of one of two promoters of bldM (Bibb et al., 2000). An
important consequence of the expression of bldM is the
activation of the chp genes encoding the morphogenetic
chaplin proteins (Elliot et al., 2003; see below). Interestingly,
some bldM and bldN alleles give rise to a white aerial
mycelium phenotype (Ryding et al., 1999), suggesting that
they are both needed at more than one stage of develop-
ment. Both of the S. avermitilis orthologues show consider-
able conservation in their upstream noncoding regions with
their S. coelicolor equivalents: in the case of bldM this
extends for about 350 bp, showing 88% identity with the
exception of one short mismatched segment (20 bp in S.
avermitilis, 10 bp in S. coelicolor); while the homology
upstream of bldN extends for about 160 bp of 94% identity.
The same segment is about 84% identical in the S. griseus
orthologue of bldN, called adsA, which is a direct target of
AdpA ( = BldH) (Yamazaki et al., 2000); and bldN transcrip-
tion is also bldH-dependent in vivo in S. coelicolor (Bibb
et al., 2000). Curiously, much of the divergence is in the
segment shown in vitro to be protected by AdpA in S. griseus,
which, unusually, is within the transcribed region upstream
of adsA (Yamazaki et al., 2000). In S. coelicolor, bldN loses its
temporal control in a bldD mutant, being strongly expressed
even early in vegetative growth; and the equivalent segment
of the bldN upstream region of S. coelicolor binds to BldD
protein in vitro (Elliot et al., 2001). However, it is not clear if
AdpA and BldD both interact with the bldN upstream region
of either organism. A second BldD-binding site for bldN
corresponds to the segment of the promoter region expected
to interact with RNA polymerase (Elliot et al., 2001), which
is extensively conserved among the three species. We sup-
pose that the temporal expression of bldN reflects both
repression by BldD and activation by AdpA in all three
species, and that the regulation of bldN by these two
proteins is likely to be an ancient trait of streptomycetes.
However, divergence in the shared BldD-/AdpA-binding
DNA segment may cause the fine details of any competition
or interaction between the two proteins to differ in S. griseus
from those in S. coelicolor and S. avermitilis.
Most actinobacteria
bldC
S. avermitilis
T. fusca
S. coelicolor
Fig. 7. Synteny in the bldC region of actinobacterial genomes, com-
pared with Streptomyces coelicolor. Connecting bridges indicate the
absence of a gene in an otherwise syntenous series of genes; gaps
indicate that no likely orthologue is present in the genome; and genes in
brackets represent apparent orthologues located elsewhere in the
genome.
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
660 K.F. Chater & G. Chandra
The ramCSABR genes, SCO6681--6685(= SAV7499--7503)
A second important morphogenetic endpoint of the cas-
cade, that seems to bypass bldN and bldM and the produc-
tion of chaplins, is the five-gene ramCSABR cluster
( = amfTSBAR in S. griseus) (Chater & Horinouchi, 2003;
Ueda et al., 2005; Willey et al., 2006). [There is a paradox in
the literature here, since it was reported that a bldM mutant
behaved the same as a bldD mutant in the extracellular
complementation cascade (Molle & Buttner, 2000); but
these experiments may be complicated by the fact that some
of the mutants used to define the cascade have different
genetic backgrounds.] The first step in ram cluster expres-
sion is the transcription of ramR, which encodes a response
regulator that contains most of the residues conserved in
phosphorylation pockets that are the targets of histidine
protein kinases in bacteria. However, ramR is not located
close to a sensor kinase gene, and evidence for RamR
phosphorylation has proved elusive (Keijser et al., 2002;
Nguyen et al., 2002; O’Connor & Nodwell, 2005). Never-
theless, it seems that a change in the state of RamR results in
full activation of the ramCSAB operon. The membrane-
bound RamC, previously thought to be a serine-threonine
protein kinase, has now been found to have another role, in
processing RamS peptide (Kodani et al., 2004). RamA/AmfB
and RamB/AmfA are subunits of an ABC transporter believed
to transport processed RamS/AmfS out of the cell (Ueda
et al., 2002; Willey et al., 2006). The primary RamS product
(38 amino acids) undergoes both cleavages and amino-acid
modifications to end up as a 17-mer amphipathic oligopep-
tide, called SapB in S. coelicolor, that contains lanthionine
cross-bridges, and hence resembles lantibiotics, though it has
not yet been found to have antibiotic properties (Kodani
et al., 2004). The name SapB originates from its first discovery
as one of several ‘spore associated proteins’ (Guijarro et al.,
1988). Remarkably, the extracellular addition of SapB can
bring about aerial growth of any of the bld mutants that can
participate in the extracellular cascade, except bldN (Willey
et al., 1993; Bibb et al., 2000).
Interestingly, the ram/amf cluster shows lower sequence
conservation (just over 50% identity for every gene product)
between Streptomyces species than do any of the other
components of the bld gene cascade. This leads to the
hypothesis that SapB may have a species-specific aspect – in
other words, that there may have been positive selection for
changes in SapB-like oligopeptides in the course of evolu-
tion. Arguing against this, the simple morphogenetic activ-
ity of SapB lacks specificity, since aerial growth of bld
mutants can be stimulated equally by certain other amphi-
pathic proteins (Kodani et al., 2004, 2005). Future work with
the equivalent systems of a wider variety of species may help
to resolve this paradox.
The differentiation of aerial hyphae intospores
A section of the sporulation regulatory networkis found only in streptomycetes
Four of the regulatory genes shown in Fig. 4 that are needed
for aerial hyphae to form mature grey spores are absent from
other actinobacteria (though one of them, whiG, is found in
many nonactinobacterial bacteria, including E. coli and B.
subtilis). Three of the four genes (whiG, H, I) form a
particular regulatory subsection of the early whi gene net-
work, and the fourth (sigF) encodes a s factor for late gene
expression.
whiG , SCO5621 ( = SAV2630)
When aerial hyphae begin to grow, it appears that they are
developmentally indeterminate. They acquire a specific
developmental fate when the sporulation-specific WhiG sfactor becomes active (Mendez & Chater, 1987; Chater et al.,
1989) (perhaps partially as a result of the release of whiG
from repression by BldD; see above). WhiG is an orthologue
of the s factors that, in many bacteria, direct transcription
of some of the genes associated with motility and chemo-
taxis (Tan et al., 1998). Although the S. coelicolor chromo-
some contains a prophage-like insertion upstream of whiG
that is absent from S. avermitilis, the genes downstream are
conserved and syntenous between the species. The whiG
genes of several other streptomycetes have been shown to
play a similar role in committing aerial hyphae to sporula-
tion, and, where sequenced, all are highly conserved in their
coding regions and in their promoters at least from � 1 to
� 45, the region containing one of the two characterized
BldD-binding sites in the whiG upstream region of S.
coelicolor (Soliveri et al., 1993; Kormanec et al., 1994; Elliot
et al., 2001; Catakli et al., 2005).
whiH , SCO5819 ( = SAV2445) and whiI , SCO6029( = SAV2230)
Two sporulation regulatory genes, whiH and whiI, are well-
established targets for sWhiG RNA polymerase holoenzyme
(Ryding et al., 1998; Ainsa et al., 1999; Kormanec et al.,
1999). There are significant matches between S. coelicolor
and S. avermitilis in the upstream regions for each of these
genes. Each gene encodes a member of a well known and
very widespread family of regulatory DNA-binding proteins.
WhiH is in the family containing GntR, many of whose
members respond to small organic acids, prompting spec-
ulation that WhiH responds to a change in concentration of
such a metabolite during aerial mycelium growth (Ryding
et al., 1998). WhiH appears to induce the strong
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
661Evolution of Streptomyces development
developmentally controlled ftsZp2 promoter to bring about
sporulation septation (Flardh et al., 2000).
WhiI resembles the response regulators associated with
bacterial two-component systems (but has important differ-
ences in the normally conserved phosphorylation pocket,
which are likewise changed in the S. avermitilis orthologue)
(Ainsa et al., 1999). There is circumstantial evidence that
different genes are induced or repressed by different forms of
WhiI, and it is postulated that WhiI (and perhaps other
orphan response regulators, including BldN and RamR; see
above) responds to a phosphorylated intermediary metabo-
lite rather than being phosphorylated by a histidine protein
kinase (Y. Tian, K. Fowler, K.C. Findlay and K.F. Chater, in
preparation). WhiI may induce the chromosome condensa-
tion that accompanies sporulation (Ainsa et al., 1999).
sigF, SCO4035 ( = SAV4185)
The s factor encoded by sigF is required for the late stages of
sporulation (Potuckova et al., 1995; Kormanec et al., 1996;
Kelemen et al., 1996). Thus, a sigF mutant undergoes
sporulation septation, but the resulting spore chains are
irregular, thin-walled and more or less unpigmented, and
contain uncondensed DNA. There is no detectable tran-
scription of sigF in whiA, B, G, H, I or J mutants, but the
reason for this is unknown (Kelemen et al., 1996). The S.
aureofaciens orthologue is also dependent on at least whiG
(Kormanec et al., 1996), indicating that sigF fulfils the same
role in diverse species, and therefore probably acquired this
role early in Streptomyces evolution. Interestingly, sF is a
member of the same subgroup of gram-positive-specific sfactors as the two forespore ss of B. subtilis and the B. subtilis
stess-responsive sB, two of which are controlled by anti-s/
anti-anti-s cascades (see bldG above). However, no genes
for such a cascade are located in the vicinity of sigF or its
orthologue in S. avermitilis. No direct targets for sF have yet
been identified, but one of the whiE promoters (for orfVIII)
is inactive in a sigF mutant (Kelemen et al., 1998; Kelemen &
Buttner, 1998).
A section of the sporulation regulatory networkinvolves genes that are found inmorphologically simple actinobacteria
whiA , SCO1950 ( = SAV6294), whiB , SCO3034( = SAV5042) and whiD , SCO4767 ( = SAV3216)
whiA and whiB mutants both have unusually long and curly
aerial hyphae that completely lack sporulation septa (Flardh
et al., 1999). A whiB mutant of S. aureofaciens had a similar
phenotype (Kormanec et al., 1998), and the whiB ortholo-
gue of Streptomyces griseocarneum complemented a whiB
mutant of S. coelicolor (Soliveri et al., 1993). Thus, WhiA
and WhiB are thought to be required to stop the continued
extension of aerial hyphae. Probably, sporulation septation
can take place only in nongrowing hyphae. It is likely that
the two gene products are closely intertwined in their
actions, either by an intimate regulatory interplay (they
certainly influence each other’s expression) or by virtue of
some protein–protein interaction (Jakimowicz et al., 2006).
The developmentally controlled expression of whiA and
whiB in S. coelicolor (Soliveri et al., 1992; Ainsa et al., 2000)
and at least of whiB in S. aureofaciens (Soliveri et al., 1992;
Kormanec et al., 1998) appears to be largely independent of
whiG.
whiA is located in a gene cluster that shows significant
conservation across all gram-positive bacteria. For example,
homologues of whiA and the two upstream genes are
similarly clustered in B. subtilis (Ainsa et al., 2000). A whiA-
free version of the cluster is found in many gram-negative
bacteria. There appear to be no reports of the physiological
role or molecular structure and function of WhiA-like
proteins in any organisms except streptomycetes. Perhaps
in other bacteria too it is required to bring growth to an
orderly halt, but the elimination of this function might not
give rise to an obvious phenotype in unicellular organisms.
whiA is expressed at a low level throughout growth by
readthrough transcription extending from the upstream
transcription unit, but is strongly upregulated during aerial
hyphal development, from an additional promoter of its
own within the immediately upstream coding sequence
(Ainsa et al., 2000). Presumably whiA acquired its sporula-
tion-specific function by mutations affecting its regulation
that occurred very early in the Streptomyces lineage. The
distribution of whiB-like (‘wbl’) genes is quite different from
that of whiA homologues. WhiB orthologues are omnipre-
sent in free-living actinobacteria, but absent from all other
organisms. They are members of a substantial family of
generally small (80–120 aa) proteins whose major conserved
features are a set of four cysteine residues and a short
glycine- and tryptophan-rich segment (Soliveri et al.,
2000). Several of these proteins, like WhiB itself, have
orthologues widespread among actinobacteria, but others
show species-specificity, implying that members of the
family have often been acquired laterally. This is consistent
with the frequent occurrence of wbl genes on phages and
plasmids of actinomycetes – for example, the large linear
plasmid SCP1 present in S. coelicolor A3(2) carries three wbl
genes (Bentley et al., 2004). Several wbl genes and their
products have been studied experimentally in streptomy-
cetes and mycobacteria. One of these, whiD, is a late
sporulation gene in S. coelicolor (Chater, 1972; Molle et al.,
2000), while its orthologue (whmB, or whiB3) in mycobac-
teria is involved in a physiologically ill-defined but impor-
tant role in the pathogenicity of M. tuberculosis and M. bovis
(Steyn et al., 2002). Studies of the whiD/whmB/whiB3
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
662 K.F. Chater & G. Chandra
orthologues have provided insights into the function and
structure of WhiB-like proteins. Evidence that WhiB3 inter-
acts directly with the principle s factor of M. smegmatis
(Steyn et al., 2002) may have broad relevance to the action of
Wbl proteins, particularly since another wbl gene (the wblP
gene of plasmid SCP1) encodes a fusion of a Wbl domain
with an ECF s factor domain (Bentley et al., 2004); and four
other wbl genes of S. coelicolor and its plasmid SCP1 are
located very close to s factor determinants. The physiologi-
cal stimulus for the activity of Wbl proteins has been
postulated to be some kind of internally generated redox
change associated with the switch from growth to non-
growth, based on the four conserved cysteine residues
(Soliveri et al., 2000); and the idea of redox sensing has
received support from evidence that WhiD contains an
oxygen-sensitive 4Fe,4S cluster (Jakimowicz et al., 2005).
Other developmental genes peculiar to complexactinobacteria
The ssg gene family
The ssgA gene (SCO3926, = SAV4267) was identified first in
S. griseus as a high copy-number suppressor of a hyperspor-
ulating mutant (Kawamoto & Ensign, 1995; Kawamoto
et al., 1997), and was shown to be required for correct
sporulation septation of S. griseus (Jiang & Kendrick, 2000),
and of S. coelicolor on most media (the exception being
minimal medium with mannitol as carbon source) (Van
Wezel et al., 2000). It has been suggested that the different
abilities of S. griseus and S. coelicolor to sporulate in
submerged culture may reflect differences in ssgA regulation
in the two organisms, since it is a target of AdpA (and hence
is A-factor-dependent) in S. griseus, but is AdpA-indepen-
dent in S. coelicolor, in which it is regulated solely by the
adjacent ssgR gene (SCO3925, = SAV4268, encoding an
IclR-like protein) (Traag et al., 2004). There also seem to be
functional differences between SsgR and its S. griseus
orthologue, SsfR, since ssfR did not complement an ssgR
mutant.
The whi genes described above have little obvious influ-
ence on ssgA expression, but, on the other hand, whiH
appears to be overexpressed in ssgA or ssgR mutants. This is
an interesting observation in view of the implied involve-
ment of ssgA in septation, since whiH has been tentatively
implicated in the sporulation-specific regulation of the key
cell-division gene ftsZ (see above).
Streptomyces coelicolor has six other ssgA paralogues, and
S. avermitilis five (hence the term ‘SALP’ for SsgA-like
proteins). In addition, at least two ssgA-like genes are
present in T. fusca, and one in K. radiodurans, the gene
family being otherwise absent from the database (Noens
et al., 2005). One of the paralogues in S. coelicolor, ssgB
(SCO1541), is also involved in sporulation: a knockout
mutant was white and completely defective in sporulation
septation, and, interestingly, had a striking large-colony
phenotype (Keijser et al., 2003). It appears that ssgB is not
needed for ssgA expression. In a comprehensive gene knock-
out analysis of the ssgA-like genes of S. coelicolor, it has
emerged that all of the SALPs have roles in the precise
determination of the pattern of peptidoglycan biosynthesis
and hydrolysis that is needed for proper sporulation septa-
tion and spore morphogenesis (Noens et al., 2005).
samR , SCO2935 ( = SAV5141)
The Beijing laboratory of H. Tan has studied several white
colony mutants of Streptomyces ansochromogenes. Most of
these have proven to be defective in orthologues of known
whi genes of S. coelicolor, providing confirmation (together
with similar work on S. aureofaciens from the laboratory of J.
Kormanec in Bratislava) that these genes (whiG, B, A)
control development in a similar manner in diverse species.
In the course of this work, Tan et al. (1998) also discovered a
previously unknown developmental gene, samR, mutations
in which had a bald colony phenotype; but when they went
on to disrupt the orthologous gene of S. coelicolor, the
mutant colonies produced fluffy, nonsporulating aerial
mycelium similar to that of a whiG mutant. The product of
samR is a member of the IclR family of transcriptional
regulators, and has no orthologues outside of streptomy-
cetes.
whiJ , SCO4543 (SAV3425), and its relatives andtheir satellite gene families, including bldB
The literature on whiJ is complicated in several ways. The
term whiJ was first applied when Ryding (1995) succeeded in
complementing the whi-77 mutant analysed by Chater and
Merrick (1976) and several other previously little-studied
whi mutants from the collection first isolated by Hopwood
et al. (1970), from which the whiA, B, D, E, G, H and I loci
had been discovered (Chater, 1972). Complementation was
attributable to one gene, SCO4543, which was therefore
defined as whiJ in the EMBL database entry (AF106004) (J.
Ainsa, N.J. Ryding and K.F. Chater, unpublished). The
product of this gene has a lambda repressor-like DNA-
binding domain at its N-terminus. Subsequently, a transpo-
son-induced whi mutant was found to harbour a transposon
insertion in SCO4543, and it was pointed out that this gene
and its two neighbours (SCO4542 and 4544) are all mem-
bers of apparently Streptomyces-specific extensive gene fa-
milies (Gehring et al., 2000). These are found in various
combinations, often in combination with members of
further gene families. One of these families includes the bldB
gene cloned by Piret & Chater (1985). All of these three gene
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
663Evolution of Streptomyces development
families are peculiar to streptomycetes and, it now emerges,
T. fusca. One of the clusters containing members of these
families, termed abaA, was discovered in a search for genes
that, at high copy number, increased antibiotic production
(Fernandez-Moreno et al., 1992). The extent of the confu-
sion surrounding these genes is illustrated by the fact that
whiJ has inadvertently been used to refer to SCO4542
instead of the correct SCO4543, and abaA to refer to just
one of the five genes (ORFs1–5) in the abaA locus
(SCO0700–0704), in a comparison of BldB with some of its
paralogues (Eccleston et al., 2002).
The phenotypes of mutations in the genes flanking whiJ
are not yet known. Interestingly, a TTA-containing para-
logue of one of the genes flanking whiJ (SCO4544) , and of
abaA orfA (SCO0702), is present in both long terminal
inverted repeats of the linear plasmid SCP1 of S. coelicolor
(SCP1.58c, SCP1.295), where it is associated with the genes
for three spore-associated proteins (SapC, D and E) (Bentley
et al., 2004). It will be rewarding to subject these gene
families to more extensive analysis.
Genes directly involved in morphogenesis andspore structure
We have already discussed the SapB morphogenetic oligo-
peptide in some detail, because of its special place as an
endpoint of the bld gene cascade; but several other surface
components also play special – perhaps unique – roles in
morphogenesis (Elliot & Talbot, 2004).
chpA-H [in order, SCO2716, 7257, 1674, 2717,1800, 2705, 2699, 1675; there are Streptomycesavermitilis orthologues only of chpC (SAV6636),chpE (SAV6478), chpH (SAV6635)]
The growth of aerial hyphae into the air has turned out to
require a partially functionally redundant family of highly
surface-active proteins, called ‘chaplins’, encoded by the chp
genes (Claessen et al., 2003; Elliot et al., 2003). Some of the
chaplins are thought to be covalently linked by sortase
enzymes to the aerial hyphal peptidoglycan, and others
probably attach noncovalently to the anchored chaplins.
The only known occurrence of chaplins outside streptomy-
cetes is in the aerial mycelium-forming T. fusca, which has a
single chp gene. It is considered that chaplins enable aerial
hyphae to break through the surface tension barrier of a
water layer that separates the substrate mycelium from
direct contact with the air (Claessen et al., 2003; Elliot
et al., 2003; Elliot & Talbot, 2004; Gebbink et al., 2005); but
we think it is also possible that the chaplin layer acts to
capture a column of water between the aerial hyphal
membrane and the chaplin-containing layer, to provide an
aqueous environment within which cellular growth of aerial
hyphae can take place. This alternative theory might also
apply to the analogous hydrophobins of mycelial fungi. The
production of chaplins depends on the integrity of the
extracellular cascade involving bld genes (see above).
rdlA and rdlB (SCO2718, 2719)
The surface of aerial hyphae of S. coelicolor also contains
another class of amphipathic proteins, the rodlins, which
possibly interact with chaplins, and are encoded by two
adjacent rdl genes (Claessen et al., 2002; Claessen et al.,
2004). Although they are made only by aerial hyphae,
rodlins are not essential for aerial growth, and indeed they
are absent from S. avermitilis, apparently because of a
deletion of a genomic segment including the rdl genes (they
are present in many other streptomycetes, but are unknown
in other organisms) (Claessen et al., 2004). Interestingly,
mutants lacking rodlins (Claessen et al., 2002) or chaplins
(Gebbink et al., 2005) attach less well to hydrophobic
surfaces, loosely supporting the idea that aerial hyphae may
serve as exploratory hyphae as well as sporophores (Yeo &
Chater, 2005).
whiE (SCO5314–5321)
This is a cluster of eight genes, most of which encode
recognizable enzymes of polyketide biosynthesis (Davis &
Chater, 1990). Mutations in whiE genes cause a loss of spore
pigment or a change in spore colour, and there is consider-
able information about the roles of these enzymes in the
synthesis of a 24-carbon chain-length polyketide (Shen
et al., 1999; Moore & Piel, 2000). However, the mature
WhiE polyketide has not been characterized because it seems
to be covalently attached to the spore wall. The whiE genes
form two diverging transcription units, both of which are
switched on only during sporulation. Both promoters are
dependent on all the early whi genes, but only the one
reading into orfVIII is also dependent on the late sporula-
tion-specific SigF s factor (Kelemen et al., 1998).
The S. avermitilis cluster (SAV2837–2844) and the par-
tially sequenced cluster of S. halstedii are colinear with that
of S. coelicolor. Although the clusters are flanked on both
sides by quite different genes in S. avermitilis and S.
coelicolor, their overall positions in the genome are con-
served. The conservation of genomic position suggests that
the flanking genes have been subjected to extensive flux
(insertion and deletion differences), while the whiE genes
themselves have remained anchored. This interpretation
may be over-simple, in view of the results of a large-scale
survey of whiE ORFII genes in diverse streptomycetes
(Metsa-Ketela et al., 2002), which showed that the phylo-
geny of these strains obtained by ribosomal DNA sequen-
cing differed substantially from that obtained by analysis of
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
664 K.F. Chater & G. Chandra
the whiE genes. This surprising finding could mean that
whiE genes have intermittently been transferred laterally
between genomes, raising the question – yet to be addressed
by further genomic comparisons – of whether other seg-
ments of the core genome might also have been exchanged
between streptomycetes over evolutionary time.
Remarkably, the genome of S. griseus contains no whiE
gene cluster (Y. Ohnishi and S. Horinouchi, pers. com-
mun.). Instead, the pigmentation of both spores and myce-
lium is attributable to a form of melanin that results from
the action of a cytochrome P-450 (P-450mel) on 1,3,6,8-
tetrahydroxynaphthalene, itself made by the condensation
of five molecules of malonyl-CoA by a type III polyketide
synthase (RppA) (Funa et al., 2005). P-450mel and rppA
genes are also present in S. coelicolor (albeit somewhat
diverged fron their S. griseus counterparts), but they do not
appear to contribute to pigmentation (Izumikawa et al.,
2003).
crgA , SCO3854 (SAV4331)
Like samR, crgA (Del Sol et al., 2003) affects differentiation
differently in different species. It encodes a small, mem-
brane-bound protein that inhibits premature sporulation
septation in S. coelicolor. A crgA mutant showed premature
aerial mycelium growth and sporulation, as well as early
production of the blue antibiotic actinorhodin. On the other
hand, a crgA mutant of S. avermitilis had white, coiled,
mostly nonsporulating aerial mycelium. Multiple copies of
crgA in both species gave white colonies in which the aerial
hyphae did not undergo sporulation septation. The gene is
present (and shows local synteny) in all actinomycete
genomes, and it would be interesting to know whether it
always affects septation.
Other genes important for development
The morphological changes associated with development
involve several proteins that determine the location and
components of events associated with sporulation septation.
These include FtsZ (cell-division) and ParAB (chromosome
segregation), both of which have complex promoter regions,
with separate segments being responsible for expression
during vegetative growth or differentiation (Flardh et al.,
2000; Jakimowicz et al., 2006). On the other hand, a
different solution to the need for expression at more than
one developmental stage has been found for at least one set
of enzymes active at different developmental stages: glyco-
gen deposition at the end of vegetative growth or at the end
of aerial growth involves differently regulated but almost
identical operons for isoforms of four enzymes that include
the glycogen branching enzyme (Schneider et al., 2000; Yeo
& Chater, 2005).
Conclusion: an overview of the evolutionof Streptomyces development
The pattern of occurrence of the genes discussed in this
article in different genomes can be used in an attempt to
reconstruct the evolution of Streptomyces development (Fig.
8). It seems that the aerial growth-stimulating system
involving both SapB/AmfS-like lanthionine-containing oli-
gopeptides and chaplins was established very early in the
evolutionary emergence of streptomycetes, presumably pro-
viding a means by which a mycelium could proceed to aerial
Low GC Gram-positives
Other actinobacteria?
Gram-negatives
whiG, whiA, in ancient bacterial progenitor
Loss of whiA
Loss of whiG; acquisition of crgA;acquisition and diversification of whiB -like genes
mycobacteria corynebacteria
Thermobifida
Streptomyces
speciation
ram; bldA/adpA;whiG,H,I,E; samR; ssgA, other ssg genes
bldC,D,G; bldB/whiJ-like; chp; protease inhibitor; ssgB
rodlins, antibioticsγ-butyrolactones
First oxygenation of atmosphere
Second oxygenation of atmosphere
Aerial growth
Fig. 8. Proposed pathway for the evolution of
Streptomyces development. The scheme was
drawn to minimize the number of gene acquisi-
tion and gene loss events.
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
665Evolution of Streptomyces development
growth in a coordinated manner. The production of these
molecules in S. coelicolor is itself dependent on other
extracellular signals, most of which are still uncharacterized;
but at least one of them (signal 1) is an incompletely defined
oligopeptide (Nodwell & Losick, 1998). These signals within
the colony indicate activity of the BldH, G, C and D
regulatory proteins, as well as of the uncharacterized bldJ
gene product that is needed to generate signal 1. It has been
proposed that these gene products are involved in responses
to different stresses that would otherwise limit continued
growth (Chater & Horinouchi, 2003): thus, only when such
responses have all been activated would the ‘last resort’ of
terminal differentiation (aerial growth at the expense of the
accumulated substrate mycelial biomass) be undertaken. At
least in this sense, the signal cascade ending in SapB
accumulation might be seen to be equivalent to the intra-
cellular sporulation phosphorelay in Bacillus spp. and their
relatives, which culminates in the accumulation of phos-
phorylated Spo0A regulatory protein, the trigger for com-
mitment to endospore formation (Errington, 2003; Fujita &
Losick, 2005).
It seems likely that AdpA (BldH) played a key role in
development early in Streptomyces evolution (from some-
where between 440 and 220 Ma), and that even then the
UUA codon of adpA mRNA was being used as a route
through which bldA influenced development. However,
certain other details of adpA regulation show species-
specificity, such as its A-factor-sensitive repressibility by
ArpA in S. griseus. The nature and range of signal inputs
affecting AdpA levels and/or activity have probably devel-
oped as part of the adaptation of different species to
different ecological niches, or their adoption of different
strategies for responding to a shared niche. For example,
colonies of one species might sporulate comprehensively
and more or less synchronously at a hint of imminent hard
times, while those of another could have evolved to delay
sporulation as long as possible in case an environmental
change restores conditions favourable for growth, making it
possible to achieve greater biomass before commitment to
sporulation. Since there are orthologues of bldG, bldC and
bldD in T. fusca, but apparently not in other completely
sequenced actinobacteria, it is likely that any stress responses
in which these genes take part predated the emergence of
streptomycetes. T. fusca and streptomycetes are alone among
sequenced actinobacteria in being obligately mycelial and
reproducing principally via a sporulating aerial mycelium,
so these stresses might conceivably be associated with this
life-style. The absence of the amf system and of multiple
SALPs, chaplins and rodlins from all organisms except
streptomycetes suggests that the acquisition of these features
was associated with the development of the Streptomyces
aerial mycelium, and that T. fusca may have a different
means of solving the problems associated with reproduction
via aerial growth, albeit probably using its single chaplin and
two SALPs. It has been suggested that Streptomyces aerial
hyphae may serve not only as sporophores, but alternatively
as the means by which a mycelial organism can extend across
the air space of tiny cavities in the soil habitat to reach and
colonize other nearby surfaces (Yeo & Chater, 2005). This
suggests, in turn, that exploratory aerial growth could have
preceded the acquisition by aerial hyphae of their reproduc-
tive function, and provided the evolutionary opportunity
for it. The capture of whiG, whiH and whiI, followed by their
adaptation to become increasingly precise regulators of the
fragmentation process that became sporulation, presumably
happened around the time of the birth of the first ancestor
peculiar to streptomycetes. In this light, it is curious that
whiH and whiI are located about 200 and 400 genes away
from whiG, in both S. coelicolor and S. avermitilis, and that
the intervening regions, although not showing extensive
synteny with other actinobacterial genomes, include a
number of genes for which there are orthologues in most
actinobacteria. If whiG and its targets were acquired in a
single event, for example as a regulon on a plasmid, there
must have been several changes in chromosomal organiza-
tion in the whiG,H,I region quite soon after that initial
acquisition event.
If many of the genes for development have been horizon-
tally acquired, what is their source? Most of them are absent
from other known bacteria, but a huge pool of novel genes is
present in bacteriophages. Presumably, many of these genes
have evolved under extreme selective pressures in the face of
the constant evolution of host organisms to resist death by
phage infection, as well as under occasional competition
between phages infecting the same host (Hendrix, 2005).
This means that phages are potentially a rich source both of
novel regulatory genes and of genes for surface-acting and
other structural proteins. Both of these gene classes are
important for the evolution of developmental and morpho-
logical complexity. Moreover, phages provide the means by
which new genes can be efficiently introduced into bacterial
hosts (Hendrix, 2005). At least in the case of whiB-like genes,
there are examples in phage genomes. One can push the idea
of phages as a source of developmental possibilities further,
in considering how the use of the bldA-specified tRNA for a
rare codon might have acquired specific developmental
significance. Thus, a drift towards high GC content could
be associated with selection against phages through codon
bias, and could eventually permit a loss of the UUA-reading
tRNA, and an associated complete inability to translate UUA
codons that would confer resistance to phages that employ
this codon. Some phages would counter this by acquiring a
copy of the tRNA (tRNA genes are often present in phage
genomes). By this means, the tRNA might be reintroduced
into a proto-streptomycete as part of a prophage genome.
Because of continued selection for phage resistance, the
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
666 K.F. Chater & G. Chandra
newly acquired tRNA might not be expressed abundantly
during vegetative growth (the main phage-sensitive stage of
bacteria), but it could be expressed without disadvantage in
stationary phase, and come to be exploited by genes that are
specifically useful in stationary phase, as seems to be the case
for TTA-containing genes of streptomycetes (Chater, 2006).
Spores (as we know them) presumably evolved from
mycelial fragments, under selective pressures that may have
included the need to survive inside the guts of worms and
other invertebrates that consumed the plant remains on
which the early streptomycetes were growing. The ability of
spores to survive in such a hostile environment would have
been enhanced by the cross-linking of the aromatic spore
pigment to the spore wall in a lignification-like process; it is
not surprising, therefore, that whiE genes appear to have
been acquired early in the Streptomyces lineage (notwith-
standing their absence from S. griseus: possibly the melanin
pigment responsible for spore colour in that organism also
‘lignifies’ spore walls). Most antibiotics, including many
other aromatic polyketides, are much more strain-specific,
and have presumably been acquired more recently. Two
sporulation-specific gene sets, for SapB and spore pigment,
have some relatedness to genes for antibiotic production
(lantibiotics and aromatic polyketides, respectively). Since
their acquisition by Streptomyces appears to have preceded
the acquisition of antibiotic production genes, it is an
intriguing possibility that they may have been important in
the origin of such genes.
The whiAB part of the sporulation regulatory cascade
seems to have had a different evolution, involving vertically
inherited (rather than laterally acquired) genes whose initi-
ally nondevelopmental regulation and regulatory roles came
to be integrated into sporulation. The presence of whiA
orthologues in all gram-positive bacteria indicates a parti-
cularly ancient lineage, preceding the divergence of the low
and high GC gram-positive bacteria some 2 Ga. No role for
WhiA-like proteins has been found in any nonstreptomy-
cetes, but the evidence connecting WhiA with the orderly
shut-down of growth may provide a clue for studies of
orthologues in other bacteria. On the other hand, whiB
orthologues and their paralogues (generally termed wbl
genes) are confined to actinobacteria. In view of evidence
that the products of such genes may be redox-sensitive
regulators, it is interesting that actinobacteria share another
special feature of their redox regulation: they use mycothiol
as a thiol-disulphide buffer, a role played by glutathione in
many other bacteria (Fahey, 2001). Possibly, there is a
Wbl–mycothiol association, which could have been fixed at
the time when the actinobacteria emerged (about 1.5–2 Ga),
coinciding with the first appearance of oxygen at significant
levels in the atmosphere. Five wbl genes (whiB, whiD, wblA,
wblC and wblE) are present in most actinobacteria, so wbl
genes must have diversified very early in actinobacterial
evolution. One of them, wblC, is involved in the expression
of multiple antibiotic resistance in both mycobacteria and
streptomycetes (Morris et al., 2005), so, by extrapolation, it
is likely that each of the five had acquired a specific
physiological role at this early stage of evolution. We there-
fore suppose that whiB and whiD were subsequently (and
probably sequentially) sequestered for sporulation in strep-
tomycetes, rather than evolving by duplication and diver-
gence from a common sporulation-specific progenitor
during a gradual increase in the complexity of sporulation
control. Thus, the evolution of the later stages of sporula-
tion, which might be equated with an increase in the
effectiveness of mycelial fragments as survival and dispersal
agents, was probably associated with the acquisition of
successively later-acting regulons; hence the adoption of sigF
and whiD, each a member of a paralogous gene family, as
Streptomyces sporulation-specific controlling elements. This
process may have interesting differences from the evolution
of developmental complexity that has emerged from other
systems. For example, in B. subtilis endospore formation,
each of the two initial compartment-specific s factors, sE
and sF, is replaced in the later stages of sporulation by a
closely related paralogous s factor (sK and sG, respectively)
(Errington, 2003), strongly suggesting that the evolution of
the late stages of sporulation, during which the spores
acquire their resistance properties, involved the duplication
of the sporulation-specific progenitors of the sE and sF
genes, and their subsequent divergence. This theme of
developmental evolution by the duplication and divergence
of developmentally specific master control genes is also
found in the homeobox genes of vertebrates and the
MADS-box genes that determine floral organ identity in
plants (Cohen, 1999). We note here that the multiple SALPs
of streptomycetes, which play important roles in consecutive
stages of cell wall reorganization during sporulation, seem to
have evolved by duplication and divergence as the process of
sporulation of aerial hyphae evolved (Noens et al., 2005).
Unlike most other whi genes, both whiA and whiB have
two promoters, one of which is low-level and constitutive,
while the other is associated with development (Soliveri
et al., 1992; Flardh et al., 2000). The developmental promo-
ters appear to be part of an autoactivating circuit in which
both gene products affect the expression of both genes
(though it is not known if this regulation is direct) (Jakimo-
wicz et al., 2006). Although the constitutive promoters may
simply serve to provide a priming element in the autoregu-
latory developmental circuit, it is also possible that WhiA
and WhiB serve some other physiological purpose in
vegetative hyphae (perhaps similar to whatever their role is
in nondifferentiating actinobacteria). Likewise, the ftsZ and
parAB genes, both of which have somewhat different roles in
vegetative growth and sporulation, also have dual promo-
ters, one for vegetative functioning and one for sporulation
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
667Evolution of Streptomyces development
(Flardh et al., 2000; Jakimowicz et al., 2006). Thus, it is quite
common to find that genes have acquired additional RNA
polymerase interaction sites as developmental complexity
has evolved. In the cases of ftsZ and parAB, it seems that
WhiH and WhiA/WhiB are, respectively, closely involved in
this developmental activation (Flardh et al., 2000; Jakimo-
wicz et al., 2006). No doubt many other examples of the
evolutionary plasticity of promoter regions will emerge in
future analysis of Streptomyces development.
Note added in press
The genome sequence of a representative of the mycelial
nitrogen-fixing symbiont Frankia has very recently become
available (http://genome.jgi-psf.org/draft_microbes/fra_c/
fra_c.home.html). Although members of this genus are not
reported to form aerial mycelium, orthologues of several bld
genes are present (bldC, D, G, N), as well as an ssgB-like
gene. This profile is interestingly similar to that of T. fusca.
Acknowledgements
This work was supported by grant IGF12432 from the
BBSRC, a Competitive Strategic Grant to the John Innes
Centre from the BBSRC, and the John Innes Foundation. We
thank Alan Ward for discussions on Streptomyces phylogeny,
Sueharu Horinouchi and Yasuo Ohnishi for communicating
unpublished results, and David Hopwood for helpful com-
ments on the manuscript.
References
Ainsa JA, Parry HD & Chater KF (1999) A response regulator-like
protein that functions at an intermediate stage of sporulation
in Streptomyces coelicolor A3(2). Mol Microbiol 34: 607–619.
Ainsa JA, Ryding NJ, Hartley N, Findlay KC, Bruton CJ & Chater
KF (2000) WhiA, a protein of unknown function conserved
among gram-positive bacteria, is essential for sporulation in
Streptomyces coelicolor A3(2). J Bacteriol 182: 5470–5478.
Babcock M & Kendrick K (1990) Transcriptional and
translational features of a sporulation gene of Streptomyces
griseus. Gene 95: 57–63.
Bentley SD, Chater KF, Cerdeno-Tarraga AM, et al. (2002)
Complete genome sequence of the model actinomycete
Streptomyces coelicolor A3(2). Nature 417: 141–147.
Bentley SD, Brown S, Murphy LD, et al. (2004) SCP1, a 356
023 bp linear plasmid adapted to the ecology and
developmental biology of its host, Streptomyces coelicolor
A3(2). Mol Microbiol 51: 1615–1628.
Bibb MJ, Findlay PR & Johnson MW (1984) The relationship
between base composition and codon usage in bacterial genes
and its use for the simple and reliable identification of protein-
coding sequences. Gene 30: 157–166.
Bibb MJ, Molle V & Buttner MJ (2000) sigma(BldN), an
extracytoplasmic function RNA polymerase sigma factor
required for aerial mycelium formation in Streptomyces
coelicolor A3(2). J Bacteriol 182: 4606–4616.
Bignell DRD, Warawa JL, Strap JL, Chater KF & Leskiw BK (2000)
Study of the bldG locus suggests that an anti-anti-sigma factor
and an anti-sigma factor may be involved in Streptomyces
coelicolor antibiotic production and sporulation. Microbiology
146: 2161–2173.
Bignell DRD, Lau LH, Colvin KR & Leskiw BK (2003) The
putative anti-anti-sigma factor BldG is post-translationally
modified by phosphorylation in Streptomyces coelicolor. FEMS
Microbiol Lett 225: 93–99.
Catakli S, Andrieux A, Decaris B & Dary A (2005) Sigma factor
WhiG and its regulation constitute a target of a mutational
phenomenon occurring during aerial mycelium growth in
Streptomyces ambofaciens ATCC23877. Res Microbiol 156:
328–40.
Chater KF (1972) A morphological and genetic mapping study of
white colony mutants of Streptomyces coelicolor. J Gen
Microbiol 72: 9–28.
Chater KF (1998) Taking a genetic scalpel to the Streptomyces
colony. Microbiology 144: 1465–1478.
Chater KF (2001) Regulation of sporulation in Streptomyces
coelicolor A3(2): a checkpoint multiplex? Curr Opin Microbiol
4: 667–673.
Chater KF (2006) Streptomyces inside out: a new perspective on
the bacteria that provide us with antibiotics. Philos Trans Roy
Soc B 361: 761–768.
Chater KF & Horinouchi S (2003) Signalling early developmental
events in two highly diverged Streptomyces species. Mol
Microbiol 48: 9–15.
Chater KF & Losick R (1997) Mycelial life style of Streptomyces
coelicolor A3(2) and its relatives. Bacteria as Multicellular
Organisms (Shapiro JA & Dworkin M, eds), pp. 149–182.
Oxford University Press, New York.
Chater KF & Merrick MJ (1976) Approaches to the study of
differentiation in Streptomyces coelicolor A3(2). Second
International Symposium on the Genetics of Industrial
Microorganisms (MacDonald KD, ed.), pp. 583–593. Academic
Press, London.
Chater KF, Bruton CJ, Plaskitt KA, Buttner MJ, Mendez C &
Helmann JD (1989) The developmental fate of S. coelicolor
hyphae depends upon a gene-product homologous with the
motility sigma-factor of B subtilis. Cell 59: 133–143.
Chen CW, Huang CH, Lee HH, Tsai HH & Kirby R (2002) Once
the circle has been broken: dynamics and evolution of
Streptomyces chromosomes. Trends Genet 18: 522–529.
Chen C-C, Lewis RJ, Harris R, Yudkin MD & Delumeau O (2003)
A supramolecular complex in the environmental stress
signalling pathway of Bacillus subtilis. Mol Microbiol 49:
1657–1669.
Claessen D, Wosten HAB, vanKeulen G, Faber OG, Alves AMCR,
Meijer WG & Dijkhuizen L (2002) Two novel homologous
proteins of Streptomyces coelicolor and Streptomyces lividans
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
668 K.F. Chater & G. Chandra
are involved in the formation of the rodlet layer and mediate
attachment to a hydrophobic surface. Mol Microbiol 44:
1483–1492.
Claessen D, Rink R, de Jong W, Siebring J, de Vreugd P, Boersma
FGH, Dijkhuizen L & Wosten HAB (2003) A novel class of
secreted hydrophobic proteins is involved in aerial hyphae
formation in Streptomyces coelicolor by forming amyloid-like
fibrils. Genes Dev 17: 1714–1726.
Claessen D, Stokroos I, Deelstra HJ, Penninga NA, Bormann C,
Salas JA, Dijkhuizen L & Wosten HAB (2004) The formation of
the rodlet layer of streptomycetes is the result of the interplay
between rodlins and chaplins. Mol Microbiol 53: 433–443.
Cohen E (1999) The Art of Genes. Oxford University Press,
Oxford.
Crick F (1966) Codon–anticodon pairing: the Wobble
hypothesis. J Mol Biol 19: 548–555.
Danilenko V, Mironov VA & Elizarov SM (2005) Calcium as a
regulator of intracellular processes in actinomycetes: a review.
Appl Biochem Microbiol 41: 319–329.
Darwin C (1881) The Formation of Vegetable Mould, Through the
Action of Worms. John Murray, London.
Davis NK & Chater KF (1990) Spore colour in Streptomyces
coelicolor A3(2) involves the developmentally regulated
synthesis of a compound biosynthetically related to polyketide
antibiotics. Mol Microbiol 4: 1679–1691.
Del Sol R, Pitman A, Herron P & Dyson P (2003) The product of
a developmental gene, crgA, that coordinates reproductive
growth in Streptomyces, belongs to a novel family of
actinomycete-specific proteins. J Bacteriol 185: 6678–6685.
Eccleston M, Ali R, Seyler R, Westpheling J & Nodwell J (2002)
Structural and genetic analysis of the BldB protein of
Streptomyces coelicolor. J Bacteriol 184: 4270–4276.
Elliot M & Talbot NJ (2004) Building filaments in the air: aerial
morphogenesis in bacteria and fungi. Curr Opin Microbiol 7:
594–601.
Elliot MA & Leskiw BK (1999) The BldD protein from
Streptomyces coelicolor is a DNA-binding protein. J Bacteriol
181: 832–6835.
Elliot M, Damji F, Passantino R, Chater K & Leskiw B (1998) The
bldD gene of Streptomyces coelicolor A3(2): a regulatory gene
involved in morphogenesis and antibiotic production.
J Bacteriol 180: 1549–1555.
Elliot MA, Bibb MJ, Buttner MJ & Leskiw BK (2001) BldD is a
direct regulator of key developmental genes in Streptomyces
coelicolor A3(2). Mol Microbiol 40: 257–269.
Elliot MA, Karoonuthaisiri N, Huang JQ, Bibb MJ, Cohen SN,
Kao CM & Buttner MJ (2003) The chaplins: a family of
hydrophobic cell-surface proteins involved in aerial mycelium
formation in Streptomyces coelicolor. Genes Dev 17: 1727–1740.
Embley T & Stackebrandt E (1994) The molecular phylogeny and
systematics of the actinomycetes. Annu Rev Microbiol 48:
257–289.
Errington J (2003) Regulation of endospore formation in Bacillus
subtilis. Nat Rev Microbiol 1: 117–126.
Fahey R (2001) Novel thiols of prokaryotes. Annu Rev Microbiol
55: 333–356.
Fernandez-Moreno M, Martin-Triana AJ, Martinez E, Niemi J,
Kieser H, Hopwood D & Malpartida F (1992) abaA, a new
pleiotropic regulatory locus for antibiotic production in
Streptomyces coelicolor. J Bacteriol 174: 2958–2967.
Flardh K & van Wezel GP (2004) Cell division during growth and
development of Streptomyces. Rec Dev Bacteriol 1: 71–90.
Flardh K, Findlay KC & Chater KF (1999) Association of early
sporulation genes with suggested developmental decision
points in Streptomyces coelicolor A3(2). Microbiology 145:
2229–2243.
Flardh K, Leibovitz E, Buttner MJ & Chater KF (2000)
Generation of a non-sporulating strain of Streptomyces
coelicolor A3(2) by the manipulation of a developmentally
controlled ftsZ promoter. Mol Microbiol 38: 737–749.
Fuglsang A (2005) Intragenic position of UUA codons in
streptomycetes. Microbiology 151: 3150–3152.
Fujita M & Losick R (2005) Evidence that entry into sporulation
in Bacillus subtilis is governed by a gradual increase in the level
and activity of the master regulator Spo0A. Genes Dev 19:
2236–2244.
Funa N, Funabashi M, Ohnishi Y & Horinouchi S (2005)
Biosynthesis of hexahydroxyperylenequinone melanin via
oxidative aryl coupling by cytochrome P-450 in Streptomyces
griseus. J Bacteriol 187: 8149–8155.
Gebbink MF, Claessen D, Bouma B, Dijkhuizen L & Wosten HA
(2005) Amyloids – a functional coat for microorganisms. Nat
Rev Microbiol 3: 333–341.
Gehring A, Nodwell J, Beverley S & Losick R (2000) Genomewide
insertional mutagenesis in Streptomyces coelicolor reveals
additional genes involved in morphological differentiation.
Proc Natl Acad Sci USA 97: 9642–9647.
Guijarro J, Santamaria R, Schauer A & Losick R (1988) Promoter
determining the timing and spatial localization of
transcription of a cloned Streptomyces coelicolor gene encoding
a spore-associated polypeptide. J Bacteriol 170: 1895–1901.
Heckman D, Geiser DM, Eidell BR, Stauffer RL, Kardos NL &
Hedges SB (2001) Molecular evidence for the early
colonization of land by fungi and plants. Science 293:
1129–1133.
Hendrix R (2005) Bacteriophage evolution and the role of phages
in host evolution. Phages: Their Role in Bacterial Pathogenesis
and Biotechnology (Waldor M, Friedman D & Adhya S, eds), pp
55–65. ASM Press, Washington DC.
Hesketh AR, Chandra G, Shaw AD, Rowland JJ, Kell DB, Bibb MJ
& Chater KF (2002) Primary and secondary metabolism, and
post-translational protein modifications, as portrayed by
proteomic analysis of Streptomyces coelicolor. Mol Microbiol 46:
917–932.
Hopwood D (1967) Genetic analysis and genome structure in
Streptomyces coelicolor. Bacteriol Rev 31: 373–403.
Hopwood DA, Wildermuth H & Palmer HM (1970) Mutants of
Streptomyces coelicolor defective in sporulation. J Gen Microbiol
61: 397–408.
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
669Evolution of Streptomyces development
Horinouchi S (2002) A microbial hormone, A-factor, as a master
switch for morphological differentiation and secondary
metabolism in Streptomyces griseus. Front Biosci 7:
D2045–D2057.
Hunt AC, Servin-Gonzalez L, Kelemen GH & Buttner MJ (2005)
The bldC developmental locus of Streptomyces coelicolor
encodes a member of a family of small DNA-binding proteins
related to the DNA-binding domains of the MerR family. J
Bacteriol 187: 716–728.
Ikeda H, Ishikawa J, Hanamoto A, Shinose M, Kikuchi H, Shiba
T, Sakaki Y, Hattori M & Omura S (2003) Complete genome
sequence and comparative analysis of the industrial
microorganism Streptomyces avermitilis. Nat Biotechnol 21:
526–531.
Ishikawa J & Hotta K (1999) FramePlot: a new implemetation of
the Frame analysis for predicting protein-coding regions in
bacterial DNA with a high G plus C content. FEMS Microbiol
Lett 174: 251–253.
Izumikawa M, Shipley PR, Hopke JN, O’Hare T, Xiang L, Noel JP
& Moore BS (2003) Expression and characterization of the
type III polyketide synthase 1,3,6,8-tetrahydroxynaphthalene
synthase from Streptomyces coelicolor A3(2). J Ind Microbiol
Biotechnol 30: 510–515.
Jakimowicz P, Cheesman MR, Bishai WR, Chater KF, Thomson
AJ & Buttner MJ (2005) Evidence that the Streptomyces
developmental protein WhiD, a member of the WhiB family,
binds a 4Fe-4S cluster. J Biol Chem 280: 8309–8315.
Jakimowicz D, Mouz S, Zakrzewska-Czerwinska Z & Chater KF
(2006) Regulation of developmentally controlled parAB
promoter leading to formation of ParB complexes in
Streptomyces coelicolor aerial hyphae. J Bacteriol 188:
1710–1720.
Jiang H & Kendrick KE (2000) Characterization of ssfR and ssgA,
two genes involved in sporulation of Streptomyces griseus. J
Bacteriol 182: 5521–5529.
Kato J, Suzuki A, Yamazaki H, Ohnishi Y & Horinouchi S (2002)
Control by A-factor of a metalloendopeptidase gene involved
in aerial mycelium formation in Streptomyces griseus. J
Bacteriol 184: 6016–6025.
Kato JY, Ohnishi Y & Horinouchi S (2005) Autorepression of
AdpA of the AraC/XylS family, a key transcriptional activator
in the A-factor regulatory cascade in Streptomyces griseus. J Mol
Biol 350: 12–26.
Kawamoto S & Ensign JC (1995) Cloning and characterization of
a gene involved in regulation of sporulation and cell division
of Streptomyces griseus. Actinomycetologica 9: 136–151.
Kawamoto S, Watanabe H, Hesketh A, Ensign JC & Ochi K
(1997) Expression analysis of the ssgA gene product, associated
with sporulation and cell division in Streptomyces griseus.
Microbiology 143: 1077–1086.
Keijser B, van Wezel G, Canters G & Vijgenboom E (2002)
Developmental regulation of the Streptomyces lividans ram
genes: involvement of RamR in regulation of the ramCSAB
operon. J Bacteriol 184: 4420–4429.
Keijser B, Noens E, Kraal B, Koerten H & van Wezel G (2003) The
Streptomyces coelicolor ssgB gene is required for early stages of
sporulation. FEMS Microbiol Lett 225: 59–67.
Kelemen GH & Buttner MJ (1998) Initiation of aerial mycelium
formation in Streptomyces. Curr Opin Microbiol 1: 656–662.
Kelemen GH, Brown GL, Kormanec J, Potuckova L, Chater KF &
Buttner MJ (1996) The positions of the sigma-factor genes,
whiG and sigF, in the hierarchy controlling the development of
spore chains in the aerial hyphae of Streptomyces coelicolor
A3(2). Mol Microbiol 21: 593–603.
Kelemen GH, Brian P, Flardh K, Chamberlin L, Chater KF &
Buttner MJ (1998) Developmental regulation of transcription
of whiE, a locus specifying the polyketide spore pigment in
Streptomyces coelicolor A3(2). J Bacteriol 180: 2515–2521.
Kelemen GH, Viollier PH, Tenor JL, Marri L, Buttner MJ &
Thompson CJ (2001) A connection between stress and
development in the multicellular prokaryote Streptomyces
coelicolor A3(2). Mol Microbiol 40: 804–814.
Kim I & Lee K (1995) Physiological roles of leupeptin and
extracellular proteases in mycelium development of
Streptomyces exfoliatus SMF13. Microbiology 141: 1017–1025.
Kim D-W, Chater K, Lee K-J & Hesketh A (2005) Changes in the
extracellular proteome caused by the absence of the bldA gene
product, a developmentally significant tRNA, reveal a new
target for the pleiotropic regulator AdpA in Streptomyces
coelicolor. J Bacteriol 187: 2957–2966.
Kodani S, Hudson ME, Durrant MC, Buttner MJ, Nodwell JR &
Willey JM (2004) The SapB morphogen is a lantibiotic-like
peptide derived from the product of the developmental gene
ramS in Streptomyces coelicolor. Proc Natl Acad Sci USA 101:
11448–11453.
Kodani S, Lodato MA, Durrant MC, Picart F & Willey JM (2005)
SapT, a lantionine-containing peptide involved in aerial
hyphae formation in the streptomycetes. Mol Microbiol 58:
1368–1380.
Kormanec J, Potuckova L & Rezuchova B (1994) The
Streptomyces aureofaciens homologue of the whiG gene
encoding a putative sigma factor essential for sporulation.
Gene 143: 101–103.
Kormanec J, Homerova D, Potuckova L, Novakova R &
Rezuchova B (1996) Differential expression of two
sporulation-specific sigma factors of Streptomyces aureofaciens
correlates with the developmental stage. Gene 181: 19–27.
Kormanec J, Sevcikova B, Sprusansky O, Benada O, Kofronova O,
Novakova R, Rezuchova B, Potuckova L & Homerova D
(1998) The Streptomyces aureofaciens homologue of the whiB
gene is essential for sporulation: its expression correlates with
the developmental stage. Folia Microbiol 43: 605–612.
Kormanec J, Novakova R, Homerova D & Sevcikova B (1999) The
Streptomyces aureofaciens homologue of the sporulation gene
whiH is dependent on rpoZ-encoded sigma factor. Biochim
Biophys Acta 1444: 80–84.
Kwak J, McCue LA & Kendrick KE (1996) Identification of bldA
mutants of Streptomyces griseus. Gene 171: 75–78.
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
670 K.F. Chater & G. Chandra
Lawlor EJ, Baylis HA & Chater KF (1987) Pleiotropic
morphological and antibiotic deficiencies result from
mutations in a gene encoding a transfer RNA-like product in
Streptomyces coelicolor A3(2). Genes Dev 1: 1305–1310.
Lee E, Karoonuthaisiri N, Kim H, Park J, Cha C, Kao C & Roe J
(2005) A master regulator sigma governs osmotic and
oxidative response as well as differentiation via a network of
sigma factors in Streptomyces coelicolor. Mol Microbiol 57:
1252–1264.
Leskiw BK, Lawlor EJ, Fernandez-Abalos JM & Chater KF (1991)
TTA codons in some genes prevent their expression in a class
of developmental, antibiotic-negative, Streptomyces mutants.
Proc Natl Acad Sci USA 88: 2461–2465.
Manteca A, Fernandez M & Sanchez J (2006) Cytological and
biochemical evidence for an early cell dismantling event in
surface cultures of Streptomyces antibibioticus. Res Microbiol
157: 143–152.
Mendez C & Chater KF (1987) Cloning of whiG, a gene critical
for sporulation of Streptomyces coelicolor A3(2). J Bacteriol
169: 5715–5720.
Merrick M (1976) A morphological and genetic mapping study of
bald colony mutants of Streptomyces coelicolor. J Gen Microbiol
96: 299–315.
Metsa-Ketela M, Halo L, Munukka E, Mantsala P & Ylihonko K
(2002) Molecular evolution of aromatic polyketides and
comparative sequence analysis of polyketide ketosynthase and
16S ribosomal DNA genes from various Streptomyces species.
Appl Environ Microbiol 68: 4472–4479.
Molle V & Buttner MJ (2000) Different alleles of the response
regulator gene bldM arrest Streptomyces coelicolor development
at distinct stages. Mol Microbiol 36: 1265–1278.
Molle V, Palframan WJ, Findlay KC & Buttner MJ (2000) WhiD
and WhiB, homologous proteins required for different stages
of sporulation in Streptomyces coelicolor A3(2). J Bacteriol 182:
1286–1295.
Moore BS & Piel J (2000) Engineering biodiversity with type II
polyketide synthase genes. Antonie van Leeuwenhoek 78:
391–398.
Morris R, Nguyen L, Gatfield J, et al. (2005) Ancestral antibiotic
resistance in Mycobacterium tuberculosis. Proc Natl Acad Sci
USA 102: 12200–12205.
Nguyen KT, Willey JM, Nguyen LD, Nguyen LT, Viollier PH &
Thompson CJ (2002) A central regulator of morphological
differentiation in the multicellular bacterium Streptomyces
coelicolor. Mol Microbiol 46: 1223–1238.
Nguyen KT, Tenor J, Stettler H, Nguyen LT, Nguyen LD &
Thompson CJ (2003) Colonial differentiation in Streptomyces
coelicolor depends on translation of a specific codon within the
adpA gene. J Bacteriol 185: 7291–7296.
Nodwell JR & Losick RM (1998) Purification of an extracellular
signaling molecule involved in production of aerial mycelium
by Streptomyces coelicolor. J Bacteriol 180: 1334–1337.
Nodwell JR, McGovern K & Losick R (1996) An oligopeptide
permease responsible for the import of an extracellular signal
governing aerial mycelium formation in Streptomyces
coelicolor. Mol Microbiol 22: 881–893.
Noens EE, Mersinias V, Traag BA, Smith CP, Koerten HK & van
Wezel GP (2005) SsgA-like proteins determine the fate of
peptidoglycan during sporulation of Streptomyces coelicolor.
Mol Microbiol 58: 929–944.
O’Connor TJ & Nodwell JR (2005) Pivotal roles for the receiver
domain in the mechanism of action of the response regulator
RamR of Streptomyces coelicolor. J Mol Biol 351: 1030–1047.
Ohnishi Y, Kameyama S, Onaka H & Horinouchi S (1999) The A-
factor regulatory cascade leading to streptomycin biosynthesis
in Streptomyces griseus: identification of a target gene of the A-
factor receptor. Mol Microbiol 34: 102–111.
Ohnishi Y, Yamazaki H, Kato JY, Tomono A & Horinouchi S
(2005) AdpA, a central transcriptional regulator in the A-
factor regulatory cascade that leads to morphological
development and secondary metabolism in Streptomyces
griseus. Biosci Biotechnol Biochem 69: 431–439.
Piret JM & Chater KF (1985) Phage-mediated cloning of bldA, a
region involved in Streptomyces coelicolor morphological
development, and its analysis by genetic complementation. J
Bacteriol 163: 965–972.
Potuckova L, Kelemen GH, Findlay KC, Lonetto MA, Buttner MJ
& Kormanec J (1995) A new RNA polymerase sigma factor,
sigma(F), is required for the late stages of morphological
differentiation in Streptomyces spp. Mol Microbiol 17: 37–48.
Ryding NJ (1995) Analysis of sporulation genes in Streptomyces
coelicolor A3(2). University of East Anglia, Norwich.
Ryding NJ, Kelemen GH, Whatling CA, Flardh K, Buttner MJ &
Chater KF (1998) A developmentally regulated gene encoding
a repressor-like protein is essential for sporulation in
Streptomyces coelicolor A3(2). Mol Microbiol 29: 343–357.
Ryding NJ, Bibb MJ, Molle V, Findlay KC, Chater KF & Buttner
MJ (1999) New sporulation loci in Streptomyces coelicolor
A3(2). J Bacteriol 181: 5419–5425.
Schneider D, Bruton CJ & Chater KF (2000) Duplicated gene
clusters suggest an interplay of glycogen and trehalose
metabolism during sequential stages of aerial mycelium
development in Streptomyces coelicolor A3(2). Mol Gen Genet
263: 543–553.
Shen Y, Yoon P, Yu TW, Floss HG, Hopwood D & Moore BS
(1999) Ectopic expression of the minimal whiE polyketide
synthase generates a library of aromatic polyketides of diverse
sizes and shapes. Proc Natl Acad Sci USA 96: 3622–3627.
Soliveri J, Brown KL, Buttner MJ & Chater KF (1992) Two
promoters for the whiB sporulation gene of Streptomyces
coelicolor A3(2) and their activities in relation to development.
J Bacteriol 174: 6215–6220.
Soliveri J, Vijgenboom E, Granozzi C, Plaskitt KA & Chater KF
(1993) Functional and evolutionary implications of a survey of
various actinomycetes for homologues of two Streptomyces
coelicolor sporulation genes. J Gen Microbiol 139: 2569–2578.
Soliveri JA, Gomez J, Bishai WR & Chater KF (2000) Multiple
paralogous genes related to the Streptomyces coelicolor
developmental regulatory gene whiB are present in
FEMS Microbiol Rev 30 (2006) 651–672 c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
671Evolution of Streptomyces development
Streptomyces and other actinomycetes. Microbiology 146:
333–343.
Steyn AJC, Collins DM, Hondalus MK, Jacobs WR Jr, Kawakami
RP & Bloom BR (2002) Mycobacterium tuberculosis WhiB3
interacts with RpoV to affect host survival but is dispensable
for in vivo growth. Proc Natl Acad Sci USA 99: 3149–3152.
Taguchi S, Kikuchi H, Suzuki M, Kojima S, Terabe M, Miura K,
Nakase T & Momose H (1993) Streptomyces subtilisin
inhibitor-like proteins are distributed widely in
streptomycetes. Appl Environ Microbiol 59: 4338–4341.
Takano E, Tao M, Long F, Bibb MJ, Wang L, Li W, Buttner MJ,
Deng ZX & Chater KF (2003) A rare leucine codon in adpA is
implicated in the morphological defect of bldA mutants of
Streptomyces coelicolor. Mol Microbiol 50: 475–486.
Takano E, Kinoshita H, Mersinias V, Bucca G, Hotchkiss G,
Nihira T, Smith CP, Bibb M, Wohlleben W & Chater K (2005)
A bacterial hormone (the SCB1) directly controls the
expression of a pathway-specific regulatory gene in the cryptic
type I polyketide biosynthetic gene cluster of Streptomyces
coelicolor. Mol Microbiol 56: 465–479.
Tan HR, Yang HH, Tian YQ, Wu W, Whatling CA, Chamberlin
LC, Buttner MJ, Nodwell J & Chater KF (1998) The
Streptomyces coelicolor sporulation-specific sigma(WhiG)
form of RNA polymerase transcribes a gene encoding a ProX-
like protein that is dispensable for sporulation. Gene 212:
137–146.
Traag BA, Kelemen GH & van Wezel GP (2004) Transcription of
the sporulation gene ssgA is activated by the IclR-type
regulator SsgR in a whi-independent manner in Streptomyces
coelicolor A3(2). Mol Microbiol 53: 985–1000.
Trepanier NK, Jensen SE, Alexander DC & Leskiw BK (2002) The
positive activator of cephamycin C and clavulanic acid
production in Streptomyces clavuligerus is mistranslated in a
bldA mutant. Microbiology 148: 643–656.
Ueda K, Oinuma KI, Ikeda G, Hosono K, Ohnishi Y, Horinouchi
S & Beppu T (2002) AmfS, an extracellular peptidic
morphogen in Streptomyces griseus. J Bacteriol 184: 1488–1492.
Ueda K, Takano H, Nishimoto M, Inaba H & Beppu T (2005)
Dual transcriptional control of amfTSBA, which regulates the
onset of cellular differentiation in Streptomyces griseus. J
Bacteriol 187: 135–142.
Van Wezel GP, Van der Meulen J, Kawamoto S, Luiten RGM,
Koerten HK & Kraal B (2000) ssgA is essential for sporulation
of Streptomyces coelicolor A3(2) and affects hyphal
development by stimulating septum formation. J Bacteriol 182:
5653–5662.
Viollier PH, Nguyen KT, Minas W, Folcher M, Dale GE &
Thompson CJ (2001) Roles of aconitase in growth,
metabolism, and morphological differentiation of
Streptomyces coelicolor. J Bacteriol 183: 3193–3203.
Viollier PH, Kelemen GH, Dale GE, Nguyen KT, Buttner MJ &
Thompson CJ (2003) Specialized osmotic stress response
systems involve multiple SigB-like sigma factors in
Streptomyces coelicolor. Mol Microbiol 47: 699–714.
Wenner T, Roth V, Fischer G, Fourrier C, Aigle B, Decaris B &
Leblond P (2003) End-to-end fusion of linear deleted
chromosomes initiates a cycle of genome instability in
Streptomyces ambofaciens. Mol Microbiol 50: 411–425.
Willey J, Schwedock J & Losick R (1993) Multiple extracellular
signals govern the production of a morphogenetic protein
involved in aerial mycelium formation by Streptomyces
coelicolor. Genes Dev 7: 895–903.
Willey J, Willems A, Kodani S & Nodwell JR (2006)
Morphogenetic surfactants and their role in the formation of
aerial hyphae in Streptomyce coelicolor. Mol Microbiol 59:
731–742.
Yamazaki H, Ohnishi Y & Horinouchi S (2000) An A-factor-
dependent extracytoplasmic function sigma factor
(sigma(AdsA)) that is essential for morphological
development in Streptomyces griseus. J Bacteriol 182:
4596–4605.
Yamazaki H, Tomono A, Ohnishi Y & Horinouchi S (2004) DNA-
binding specificity of AdpA, a transcriptional activator in the
A-factor regulatory cascade in Streptomyces griseus. Mol
Microbiol 53: 555–572.
Yeo M & Chater K (2005) The interplay of glycogen metabolism
and differentiation provides an insight into the developmental
biology of Streptomyces coelicolor. Microbiology 151: 855–861.
FEMS Microbiol Rev 30 (2006) 651–672c� 2006 Federation of European Microbiological SocietiesPublished by Blackwell Publishing Ltd. All rights reserved
672 K.F. Chater & G. Chandra