Heterochromatin

Darkly stained and condensed

Transcriptionally silent and silences adjacent genes

Present at centromeres and telomeres

HP1 interacts with H3 only when K9 is methylated

Repressive structure can be propagated

Euchromatic gene placed in heterochromatin is repressed

from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-33

Histone Modifications Associated with Heterochromatin and Euchromatin

from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-33

Initiation of Heterochromatin Assembly

from Grewal and Gia, Nature Rev.Genet. 8, 35 (2007)

Transcription factors and RNAi machinery bind to specific sequences or repetitive elements to recruit histone modifying enzymes

Boundary elements prevent further heterochromatin spread

HP1 recruits histone modifying enzymes to facilitate heterochromatin spread

Modified histones recruit HP1

from Bannister et al., Nature 410, 120 (2001)

Mechanism of Heterochromatin Spreading

HP1 binds to H3K9me3

HP1 recruits SUV39H1 methylase

SUV39H1 methylates H3K9 on neighboring nucleosomes

Heterochromatin spreading is restricted by boundary elements

H1 recruits Su(var)3-9 to heterochromatin resulting in transposon silencing

Propagation of Heterochromatin

from Maison and Almounzi, Nature Rev.Mol.Cell Biol. 5, 296 (2004)

Passage of the replication fork releases parental modified nucleosomes

Nucleosome binding sites are created by recruitment of CAF1 by PCNA

CAF1-bound HP1 recruits Suv39h, Dnmt1, and HDAC

Methylated histones provide new HP1 binding sites

Structural RNA associates

from Grewal and Gia, Nature Rev.Genet. 8, 35 (2007)

Heterochromatin Functions

DNA or H3 methylation recruits adaptors such as HP1

Adaptors recruit effectors that are involved in chromosome segregation, gene silencing, transcriptional activation, and histone modification

Role of RNAi in Heterochromatin Formation in S. pombe

dsRNA is transcribed from centromeric repeats or synthetic hairpin RNAs

dsRNA is processed to siRNA

siRNA promotes H3K9 methylation by Clr4

Methylated H3K9 recruits Swi6 to form silenced chromatin

Transcription of the top strand of centromeric repeats is repressed

Rdp1 activity ensures continuous dsRNA synthesis

Recruitment of Clr4 by Swi6 chromatin leads to spread of heterochromatinfrom Schramke and Allshire, Science 301, 1069 (2003)

Formation of Telomeric Heterochromatin

from Grunstein, Cell 93, 325 (1998)

RAP1 binds to C1-3A repeats

Recruits Sir proteins

Overexpression of Sir3 causes spread of telomeric heterochromatin

Silencing decreases exponentially with distance

Mechanism of Silencing at Telomeres

Sir2 deacetylates histones

Sir3,4 binds deacetylated histones and recruits additional Sir2

from Lodish et al., Molecular Cell Biology, 6th ed. Fig 7-35

Insulators Prevent the Progression of Condensed Chromatin

from West et al, Genes Dev. 16, 271 (2002)

Insulators protect genes from inappropriate signals

Insulators block the action of distal enhancers

Insulators prevent the spreading of heterochromatin

CTCF is the Main Insulator Protein in Vertebrates

Cohesin colocalizes with CTCF and is essential for CTCF function

CTCF mediates long-range interactions between genomic sequences

CTCF establishes domains in which genes are coregulated and targets regulatory sequences to their promoters

from Ong and Corces, Nature Rev.Genet. 15, 234 (2014)

gypsy Retrotransposon Contains an Insulator

gypsy protects a transgene from position effects

su(Hw) is necessary for enhancer blocking activity

gypsy contains a su(Hw) binding site

su(Hw) blocks the process that brings enhancer and promoter together

Formation of insulator bodies at the nuclear periphery to divide the chromosome into looped domains

Multiple su(Hw) binding sites can inhibit enhancer blocking activity

Models for Heterochromatin Barrier Formation

from Donze and Kamakaka, BioEssays 24, 344 (2002)

Stable block interrupts propagation of heterochromatin

Active barrier recruits a complex containing chromatin remodeling activity

BRCA1 Modifies Pericentric Heterochromatin

BRCA1 promotes enrichment of Ub-H2A in pericentric heterochromatin

Loss of BRCA1 triggers transcription of satellite-DNA in pericentric heterochromatin

Satellite-DNA transcription is sufficient to induce genome instability after loss of BRCA1

from Venkitaraman, Nature 477, 169 (2011)

Epigenetics

Heritable changes in gene function that cannot be explained by changes in gene sequences

DNA methylation

Nucleosome positioning

Histone variants and modifications

Epigenetic Modifications During Development

Epigenetically imposed restrictions to plasticity are erased in the germ line

Early mammalian development is characterized by progressive restriction of cellular plasticity accompanied by acquisition of epigenetic modifications

Epigenetic modifications impose a cellular memory that accompanies and enables stable differentiation

from Zhang, Science 320, 489 (2008)

Epigenetic Modifications Within an Arabidopsis Chromosome

Heterochromatin correlates with epigenetic marks

DNA Methylation

Methylation at CpG residues

Sites of methylation

Inactive X

Imprinted loci

Transposon-derived sequences

CpG islands and CpG island shores

Methylation patterns are reproduced at each round of cell division

Methylated CpG Islands Inhibit Transcription

Promoters are usually unmethylated

More than half of human promoters contain CpG islands

Methylated DNA recruits methyl-CpG-binding domain proteins which recruit histone modifying and chromatin-remodelling complexes

Unmethylated CpG islands recruit Cfp1 which associates with a histone methyltransferase creating H3K4me3

from Portela and Esteller, Nature Biotechnol. 28, 1057 (2010)

Methylated CpG Islands Inhibit Transcription

Promoters are usually unmethylated

More than half of human promoters contain CpG islands

Methylated DNA recruits methyl-CpG-binding domain proteins which recruit histone modifying and chromatin-remodelling complexes

Unmethylated CpG islands recruit Cfp1 which associates with a histone methyltransferase creating H3K4me3

from Portela and Esteller, Nature Biotechnol. 28, 1057 (2010)

Methylation of Repetitive Sequences Stabilize Chromosomes

Unmethylated repetitive sequences cause reactivation of endoparasitic sequences

from Portela and Esteller, Nature Biotechnol. 28, 1057 (2010)

RNA-dependent DNA Methylation in Plants

Methylation occurs in transposons and repetitive elements

PolIV transcribes ssRNA which is converted to dsRNA by RDR2

siRNA is produced by DCL3 and loaded onto AGO4

PolV produces IGN transcripts and recruits AGO4

siRNA-IGN duplex is formed and recruits DRM2

from Law and Jacobsen, Nature Rev.Genet. 11, 204 (2010)

from Law and Jacobsen, Nature Rev.Genet. 11, 204 (2010)

De Novo DNA Methylation in Mammals

DNMT3L interacts with unmethylated H3K4

DNMT3A is recruited and activated and forms a tetrameric complex

Active sites are separated by 8-10 bp and methylates opposite DNA strands

Tetramer oligomerizes and results in 10 bp pattern of methylation on the same strand

from Cedar and Bergman, Nature Rev.Genet. 10, 295 (2009)

Establishment of DNA Methylation Pattern

Most CpGs are unmethylated before implantation

RNA pol II recruits H3K4 methyltransferase

DNMT3L only binds unmethylated H3K4 and recruits DNA methyltransferases

Propagation of DNA Methylation State

Newly synthesized methylated DNA is hemimethylated

NP95 links DNMT1to hemimethylated DNA

DNMT1 is a maintenance methyltransferase and binds PCNA

NP95 binds hemimethylated DNA

from Richly et al., BioEssays 32, 669 (2010)

Mechanisms for Repression Mediated by MBD Proteins

from Wade, BioEssays 23, 1131 (2001)

Rett Syndrome is linked to mutations in MECP2 on the X chromosome

MeCP2 binds CpG residues and silences target genes such as BDNF and corticotropin-releasing hormone

Neural activity triggers MeCP2 phosphorylation and target gene activation

MeCP2 Regulates Gene Expression in Response to Neural Activity

Hippocampal neurons grow dendrites with fewer branches when MeCP2 is blocked

from Miller, Science 314, 1356 (2006)

from Bienvenu and Chelly, Nature Rev.Genet. 7, 415 (2006)