‘‘mobile’ DNA: transposable elementsmobile’ DNA: transposable elements

Transposable elementsTransposable elements

Discrete sequences in the genome that have Discrete sequences in the genome that have

the ability to translocate or copy itself the ability to translocate or copy itself

across to other parts of the genome across to other parts of the genome without without

any requirement for sequence homology any requirement for sequence homology byby

using a self-encoded recombinase called using a self-encoded recombinase called

transposase transposase

Transposable elements move from Transposable elements move from place to place in the genomeplace to place in the genome

1930s Marcus Rhoades and 1950s 1930s Marcus Rhoades and 1950s Barbara McClintock Barbara McClintock – transposable – transposable elements in corn elements in corn

1983 McClintock received Nobel 1983 McClintock received Nobel PrizePrize

Found in all organismsFound in all organisms Most 50 – 10,000 bpMost 50 – 10,000 bp May be present hundreds of times in May be present hundreds of times in

a genomea genome

TEs can generate mutations in adjacent genesTEs can generate mutations in adjacent genes

TEs in MaizeTEs in Maize

Fig 15.19 Genes VII by Fig 15.19 Genes VII by B. LewinB. Lewin

Common mechanism of transpositionCommon mechanism of transposition Transposons encode transposases that Transposons encode transposases that

catalyse transposition eventscatalyse transposition events Regulation of transposase expression Regulation of transposase expression

essentialessential

Fig13.24a: Hartwell

Common mechanism of transpositionCommon mechanism of transposition

Common mechanism of transpositionCommon mechanism of transposition

2 sequential steps2 sequential steps

Site specific cleavage of Site specific cleavage of DNA at the end of TEDNA at the end of TE

Complex of transposase-Complex of transposase-element ends element ends (transpososome)(transpososome) brought to DNA target brought to DNA target where strand transfer is where strand transfer is carried out by covalent carried out by covalent joining of 3’end of TE to joining of 3’end of TE to target DNA target DNA

transpososome

Common mechanism of transpositionCommon mechanism of transposition transposase (blue) binds and assembles a paired end complex transposase (blue) binds and assembles a paired end complex

(PEC) by dimerization, a process that might involve divalent (PEC) by dimerization, a process that might involve divalent metal ions (Memetal ions (Me2+2+). ).

PEC is then active for the cleavage reactions that remove PEC is then active for the cleavage reactions that remove flanking donor DNA (thin black lines) and transfer of the flanking donor DNA (thin black lines) and transfer of the transposon ends into target DNA (black dotted line).transposon ends into target DNA (black dotted line).

Trends in Microbiology 2005 Vol13(11) pp 543-549

Catalytic domain of transposase involved in a transphosphorylation Catalytic domain of transposase involved in a transphosphorylation reaction that initiates DNA cleavage & strand transferreaction that initiates DNA cleavage & strand transfer

Fig 15.14

Fig 15.10GenesVII Lewin

RNA intermediatesRNA intermediates Class I TEsClass I TEs – –

Use a ‘copy & paste’ Use a ‘copy & paste’ mechanismmechanism

DNA intermediatesDNA intermediates Class II TEsClass II TEs Use a ‘cut and paste’ mechanism Use a ‘cut and paste’ mechanism Generally short sequencesGenerally short sequences

Transposition can occur viaTransposition can occur via

See interspersed repeats from the repetitive elements lecture

How transposons moveHow transposons move

Classes of transposable elementsClasses of transposable elements

Science 12 March 2004: Vol. 303. no. 5664, pp. 1626 - 1632

Interspersed repeats (transposon-derived)Interspersed repeats (transposon-derived)

classclass familyfamily sizesize Copy Copy numbnumb

erer

% % genomgenom

e*e*LINELINE L1 (Kpn family)L1 (Kpn family)

L2 L2

~6.4kb~6.4kb 0.5x100.5x1066

0.3 x 100.3 x 1066

16.916.9

3.23.2

SINESINE AluAlu ~0.3kb~0.3kb 1.1x101.1x1066 10.610.6

LTRLTR e.g.HERVe.g.HERV ~1.3kb~1.3kb 0.3x100.3x1066 8.38.3

DNA DNA

transposontransposon

marinermariner ~0.25kb~0.25kb 1-2x101-2x1044 2.82.8

major types

* Updated from HGP publications HMG3 by Strachan & Read pp268-272

LINEs LINEs (long interspersed elements)

Science 12 March 2004: Vol. 303. no. 5664, pp. 1626 - 1632

Most ancient of eukaryotic genomesMost ancient of eukaryotic genomes Autonomous transposition (reverse Autonomous transposition (reverse

trancriptase)trancriptase) ~6-8kb long, located mainly in euchromatin~6-8kb long, located mainly in euchromatin Internal polymerase II promoter and 2 ORFsInternal polymerase II promoter and 2 ORFs 3 related LINE families in humans 3 related LINE families in humans

– – LINE-1, LINE-2, LINE-3.LINE-1, LINE-2, LINE-3.

LINE-1 still active (~17% of human genme)LINE-1 still active (~17% of human genme) Believed to be responsible for Believed to be responsible for

retrotransposition of SINEs and creation of retrotransposition of SINEs and creation of processed pseudogenesprocessed pseudogenes

LINEs

LINEs (long interspersed elements)

HMG3 by Strachan & Read pp268-272

SINEs SINEs (short interspersed elements)

Science 12 March 2004: Vol. 303. no. 5664, pp. 1626 - 1632

Non-autonomousNon-autonomous (successful freeloaders! (successful freeloaders! ‘borrow’ RT from other sources such as LINEs)‘borrow’ RT from other sources such as LINEs)

~100-300bp long~100-300bp long Internal polymerase III promoter Internal polymerase III promoter No proteinsNo proteins Share 3’ ends with LINEsShare 3’ ends with LINEs 3 related SINE families in humans 3 related SINE families in humans

– – active Alu, inactive MIR and Ther2/MIR3.active Alu, inactive MIR and Ther2/MIR3.

SINEs

100-300bp 1,500,000 13%

LINES and SINEs have preferred insertion sitesLINES and SINEs have preferred insertion sites

In this example, In this example, yellow represents the yellow represents the distribution of distribution of mysmys (a (a type of LINE) over a type of LINE) over a mouse genome where mouse genome where chromosomes are chromosomes are orange. There are orange. There are more more mysmys inserted in inserted in the sex (X) the sex (X) chromosomes. chromosomes.

Try the link below to do an online experiment Try the link below to do an online experiment which shows how an Alu insertion which shows how an Alu insertion polymorphism has been used as a tool to polymorphism has been used as a tool to reconstruct the human lineagereconstruct the human lineage

http://www.geneticorigins.org/http://www.geneticorigins.org/geneticorigins/pv92/intro.htmlgeneticorigins/pv92/intro.html

Long Terminal Repeats (LTR)

Science 12 March 2004: Vol. 303. no. 5664, pp. 1626 - 1632

Repeats on the Repeats on the same orientationsame orientation on both sides of element on both sides of element e.g. ATATATnnnnnnnnnnnnnnATATATe.g. ATATATnnnnnnnnnnnnnnATATAT• contain sequences that serve as transcription contain sequences that serve as transcription

promoters as well as terminators. promoters as well as terminators. • These sequences allow the element to code for an These sequences allow the element to code for an

mRNA molecule that is processed and mRNA molecule that is processed and polyadenylated. polyadenylated.

• At least two genes coded within the element to At least two genes coded within the element to supply essential activities for retrotransposition. supply essential activities for retrotransposition.

• RNA contains a specific primer binding site (PBS) for RNA contains a specific primer binding site (PBS) for initiating reverse transcription. initiating reverse transcription.

• small direct repeats formed at the site of integration. small direct repeats formed at the site of integration.

Long Terminal Repeats (LTR)

Autonomous or non-autonomousAutonomous or non-autonomous Autonomous LTR encode retroviral genes Autonomous LTR encode retroviral genes gag, polgag, pol

genes e.g HERVgenes e.g HERV Non-autonomous elements lack the Non-autonomous elements lack the polpol and and

sometimes the sometimes the gag gag genes e.g. MaLRgenes e.g. MaLR

Long Terminal Repeats (LTR)

Ancestral repeats (AR)Ancestral repeats (AR)‘transpositional fossils’

Comprise ~ 25% of the genome

~780 classes

Largely nonfunctional

Sporadic cases where AR have

acquired anew function after

insertion

MER121 is highly conserved among

mammals!!

Science 12 March 2004: Vol. 303. no. 5664, pp. 1626 - 1632

DNA transposonsDNA transposons

DNA intermediateDNA intermediateClass II TEsClass II TEs

IS elements and transposonsIS elements and transposons

bounded by bounded by invertedinverted terminal repeats (ITR)terminal repeats (ITR)

e.g. ATGCNNNNNNNNNNNCGTAe.g. ATGCNNNNNNNNNNNCGTA

DNA intermediateDNA intermediateClass II TEsClass II TEs

Prokaryotic IS elements (e.g. IS10, Prokaryotic IS elements (e.g. IS10, Ac/Ds, mariner) encode only transposase Ac/Ds, mariner) encode only transposase sequences sequences

eukaryotic transposons encode eukaryotic transposons encode additional genes such as antibiotic additional genes such as antibiotic resistance genesresistance genes

Some types of rearrangements mediated by DNA Some types of rearrangements mediated by DNA transposonstransposons

Gene (2005)345 pp91-100

Transposons move in different Transposons move in different waysways

Classified into 5 families on the basis of their Classified into 5 families on the basis of their transposition pathwaystransposition pathways

1)      DDE-transposases1)      DDE-transposases2)      RT/En transposases2)      RT/En transposases

(reverse transcriptase/endonuclease) (reverse transcriptase/endonuclease) 3)  Tyrosine (Y) transposases 3)  Tyrosine (Y) transposases 4)      Serine (S) transposases4)      Serine (S) transposases5) 5) Rolling circle (RC) or Y2 transposasesRolling circle (RC) or Y2 transposases

Nature Rev Mol. Cell Biol (Nov2003) 4(11):865-77Nature Rev Mol. Cell Biol (Nov2003) 4(11):865-77))

Transposons can be used to transfer Transposons can be used to transfer DNA between bacterial cellsDNA between bacterial cells

Transposons (pink) integrate into new sites on the chromosome or plasmids by non-homologous recombination. Integrons (dark green) use similar mechanisms to exchange single gene cassettes (brown).

Nature Reviews Microbiology 3, 722-732 (2005)

Some transposons can encode Some transposons can encode integronsintegrons

Integrons are assembly platforms — DNA Integrons are assembly platforms — DNA elements that acquire open reading frames elements that acquire open reading frames embedded in exogenous gene cassettes and convert embedded in exogenous gene cassettes and convert them to functional genes by ensuring their correct them to functional genes by ensuring their correct expression. expression.

e.g. bacterial Tn7 also encodes an integron — a e.g. bacterial Tn7 also encodes an integron — a DNA segment containing several cassettes of DNA segment containing several cassettes of antibiotic-resistance genes. These cassettes can antibiotic-resistance genes. These cassettes can undergo rearrangements in hosts that express a undergo rearrangements in hosts that express a related recombinase, leading to alternative related recombinase, leading to alternative combinations of antibiotic-resistance genes.combinations of antibiotic-resistance genes.

Mazel Nature Reviews Microbiology 4, 608–620 (August 2006)

IntegronsIntegrons

Mobile IntegronsMobile Integrons

SuperintegronsSuperintegrons

ReadingReading

1)1) Chapter 9 pp 265-268 Chapter 9 pp 265-268 HMG 3 by HMG 3 by Strachan and Strachan and

ReadRead

2)2) Chapter 10: pp 339-348Chapter 10: pp 339-348Genetics from genes to Genetics from genes to genomes by genomes by Hartwell et alHartwell et al (2/e) (2/e)