Histone Modification II PDF

Epigenetic code : Histone modification II BSMS214 Epigenetics Week 4/2 Lecturer: Taejeong Bae, PhD exclusive email for the class: [email protected] Learning objects Recap of DNA packaging More about Histone modifications Complexity of Histone code DNA, Histone, and Ch...

Epigenetic code : Histone modification II BSMS214 Epigenetics Week 4/2 Lecturer: Taejeong Bae, PhD exclusive email for the class: [email protected] Learning objects Recap of DNA packaging More about Histone modifications Complexity of Histone code DNA, Histone, and Chromatin Diameter of nucleus = up to 10 m Eukaryotic genomic DNA is compacted more than 10,000-fold by highly basic proteins = HISTONES The result is a highly structured entity termed CHROMATIN. Nucleosome core particle consists of 147 bp of super helical DNA wrapped in 1.75 turns around a histone octamer 2 m of DNA in a nucleus core with a diameter of 5-10 m Centrally located histone (H3/H4)2 tetramer is assembled with two histone H2A/H2B dimers Histone octamer + DNA = Nucleosome Multiple Levels of Chromatin Folding Chromatin compaction influences activity of DNA in transcription Euchromatin: Less densely compact, transcriptionally active chromatin Heterochromatin: Highly compact, transcriptionally silent chromatin Changing chromatin to allow transcription Opening and closing the chromatin template Changing the chromatin-template to help transcription factors Chromatin remodeling Opening of chromatin through directed nucleosome mobilization SWI/ Histone SNF ATP-dependent process RSC modification NURF HDACs Positioning of nucleosomes creates promoters CHRAC with different requirement for remodeling ACF HATs Opening of chromatin through directed modification of histone tails Basis for the histone code hypothesis 1. **Opening of chromatin through directed nucleosome mobilization**: - **ATP-dependent process**: The chromatin is opened up by chromatin remodelers, which are proteins that use energy from ATP hydrolysis to move or reposition nucleosomes (DNA wrapped around histone proteins). By shifting nucleosomes along the DNA, certain regions, such as promoters, become more accessible to transcription factors and the transcription machinery. 2. **Positioning of nucleosomes creates promoters with different requirements for remodeling**: - The specific arrangement of nucleosomes at a gene promoter affects how easily that region can be accessed. Some promoters are tightly bound by nucleosomes and need extensive remodeling to be accessible, while others may be more loosely packed and easier to access Histone Code Code - a system of signals or symbols for communication (Webster-Meriam online) Requirements from a code: Consistent Combinatorial (Kurdistany & Grunstein, 2003) The Histone Code Various postransla5onal modifications: Acetylation Sumoylation Methylation Deimination Phosphorylation ADP Ribosylation Ubiquitination etc. Patterns of these modifications form the "histone code" Specific modifications can be recognized by specific proteins, amplifying the effect of an activator or a repressor Histones are among the most conserved proteins known in evolution, but.....are also among the most variable in post-translational modifications. The substrates: Histone tails - multiple modifications Mechanisms that HPTM can affect genome regulation and function: 1. Cis effects Affect structure and can prevent contacts that facilitate chromatin conformations or higher order structures. 2. Trans effects Disrupt binding of proteins that associate with chromatin or histones. Provide altered binding surfaces that recruit effector proteins, having an activating or repressive outcome on transcription. Histone Post-translational Modifications (HPTM) May regulate transcription via combinatorial patterns, in temporal sequences. Can be established over short and long-range distances. Establish different functional outcomes - some transient, others stable and epigenetically heritable. Histone Phosphorylation Phosphorylation Best known HPTM, histones among first proteins found to be phosphorylated - due to P-H3S10 Kinases for P-H3S10: Msk1/2 and Rsk2 (mammals), SNF1 (S. cerevisiae) Phosphorylation H3S10 important phosphorylation site H1 phosphorylation may also affect transcriptional control Methyl-phos binary switch correlating with chromosome condensation in mitosis and meiosis P-H3S10ph like a temporal switch: ejects HP1 bound at H3K9. May act in concert with P-H3S28 and P-H3T11 by recruiting condensin complex and the mitotic spindle. Phosphorylation and Models for Effect on Chromatin 1st model: The negative charge resulting from phosphorylation of H1 affects its affinity to bind DNA therefore making it more accessible to transcriptional machinery. 2nd model: Proteins bound to DNA are dislodged by phosphorylation. -H3S10 3rd model 14-3-3 adapter protein (phospho-binder protein) recognizes P-H3S10 at promoters of inducible genes Histone Acetylation Acetylation and Deacetylation N-term tails reversible acetylated in Lys, particularly in H3+H4 Acetylation and Models for Effect on Chromatin Structural changes of Acetylation ve charge reduces strength of DNA binding to histones opens binding sites Can decompact nucleosome arrays opens chromatin for gene activation Removal of histone tails results in nucleosomal arrays that cannot condense past the 11nm fiber Binding of Proteins to regulate DNA-templated processes Bromodomain binds acetylated lysines. Present in HATs. Allow complexes with these proteins to bind chromatin A link histone acetylation - transcription Acetylated histone-tails enriched in transcribed chromatin Immuno-fractionated chromatin: acetylated fraction enriched in active chromatin Neutralizing mutations in histone-tails affect transcription Change in level of acetylation using inhibitors of deacetylation enhance transcription Chromatin immunoprecipitations (ChIP) show enrichment of acetylated histones in active promoters Acetylation makes nucleosomal DNA more accessible for TF- binding - a ChIP: Chromatin Immunoprecipitation The enzymes: the HATs Histone acetyltransferases (HATs) Catalyze the transfer of acetyl groups from acetyl-CoA onto histone tails Two main groups: A-type and B-type HATs -Type HATs catalyze thioredoxin(trx)-related acetylations -Type HATs are cytoplasmic and catalyze acetylations linked to transport of newly synthesized histones from the cytoplasm to the nucleus Histone deacetylases (HDACs) HAT families GNAT family Gcn5-related N-acetyltransferase Many contains bromodomains MYST Named for its founding members: MOZ, Ybf2/Sas3, Sas2, Tip60 Several contain chromodomains CBP/p300 GTF-HATs TAFII250 Nuclear receptor linked HATs SRC1,ACTR Global versus targeted acetylation/deacetylation Global acetylation Bulk acetylation levels surprisingly high On average,13 of the 30 tail Lys residues in a histone octamer are acetylated. this steady-state level of acetylation is maintained by the opposing actions of HAT and HDAC complexes. Targeting of HAT and HDAC complexes to promoter regions then creates in specific patterns of hyper- and hypoacetylation in a background of global acetylation that correlate with transcription activation and repression, respectively. Recruitment was the missing link HATs were for a long time not considered interesting for transcription because they were assumed to act globally Recruitment changed this misconception HAT-activity in coactivator- complexes recruited to promoters by TF-interaction Histone Methylation Histone methylation Both Lys (K) and Arg (R) can be methylated asymmetric with more than one methyl-group symmetric Histone Methylation Methylation is a relatively stable modification with a slow turnover rate. Histone demethylases have been recently found. An ideal epigenetic mark for more long-term maintenance of chromatin states. Methylated residues are present both in eu- and heterochromatin. The Subtrates: Histone Tails - Multiple Methylations Methylation of Lysines Biological significance came to light with discovery of first HKMT All enzymes share catalytic active site, the SET domain and bind cofactor. (except Dot 1) 6 well characterized sites H3K4, H3K9, H3K27, H3K36, H3K79 H4K20 Activation: H3K4, H3K36, H3K79 DNA Repair: H3K79, H4K20 The enzymes: Histone lysine methyltransferases (HKMTs) The first HKMT, SUV39H1 was identified in 2000. It specifically methylates Lys 9 of histone H3. Later several other HMTs have been identified (see table below) A SET-domain is required for HKMT activity 5 lysines within H3 (K4, K9, K27, K36 and K79) and 1 lysine within H4 (K20) are methylated by specific HKMTs SET-domains Most HKMTs characterized by a conserved SET-domain Humans: >30 SET-domains Yeast genome: 6 SET-domains >300 found Function - Activation or Repression? 2 sites 2 HKMTs 2 Effects Histone methylation was traditionally linked to repression, but turns out to be linked also to activation H3K9 methylation correlates with heterochromatin formation. Me-K4 in H3 correlates with transcriptional activation Function of Histone Methylation Lysine Repression (H3-K9, H3-K27, H4-K20) Numerous reports on silencing effects linked to H3-K9 methylation Heterochromatin formation linked to H3-K9 methylation and HP1 binding Highly condensed centromeric regions related to methyl addition to H3-K9. H3-K9 and H3-K27 methylation are important for the X chromosome inactivation process. Activation (H3-K4, H3-K36, H3-K79) H3-K4 methylation is generally associated with trx active chromatin. di-methyl H3-K4 appears to be a global epigenetic mark in euchromatic regions and tri- methylation of H3-K4 correlates with active transcription both H3-K4 and H3-K79 participate in establishing euchromatic regions by preventing the spreading of heterochromatic regions. Role of histone lysine methylation in transcriptional elongation  Modification of H3 Lys9 - Eu- and Heterochromatin? Activating Effects of Methylation- 2 Levels Chromosome level - preventing spread of heterochromatin The methylation of H3-K4 specifically impairs methylation at H3-K9, thereby blocking a major pathway of heterochromatin formation binding of the histone deacetylase NuRD repression complex to the H3 N-terminal tail is precluded by methylation at K4, but not K9 Gene level di-methyl H3-K4 appears to be a global epigenetic mark in euchromatic regions and tri-methylation of H3-K4 correlates with active transcription Elongation Function - Arginine Methylation Methylation of specific arginines in histones H3 and H4 correlate with the active state of transcription. Ex: Methylation of Arg 3 of histone H4 facilitates H4 acetylation and enhances transcription activation by nuclear hormone receptors. Histone arginine methyltransferases (HRMTs) PRMTs Several Arg HMTases idenFfied: The PRMT domain The catalytic module that methylates specific arginines is known as the PRMT (protein R methyltransferase) domain. The PRMT domain transfers the methyl group from SAM to the guanidino group of arginines to produce monomethylarginine or dimethylarginine. Some PRMT modules specifically prepare symmetric dimethylarginine and others produce asymmetric dimethylarginine. Other Histone Modifications Ubiquitylation/Deubiquitylation and Sumoylation Large polypeptides Increase the size of the histone by 2/3 Ubiquitylation Activating or Repressing depending on site No histone-specific ubiquitin-binding protein has been identified Mediators may interact by 2 binding sites: to surface of ubiquitin and to histone sequence. Sumoylation Negative acting to prevent activating HPTM Directly block lysines Sumoylated histones may recruit HDAC PARP PARP recruitment During activation, PARP is recruited by TFs to condensed chromatin (green). Modification = Poly-ADP-ribosylation PARP induces dissociation of nucleosomes (beige) by adding long ADP-ribose tails to their histone proteins. Stays open PARP prevents rebinding of transcription factors to the DNA by adding ADP-ribose tails to them The Complexity of the Histone Code