Chromatin modifications

We think of chromatin in 2 phases: Heterochromatin: non transcribed, condensed Euo-chromatin: Transcription, less condensed (early replication) Changes through cell cycle We think of nucleosomes as the basic repetitive mode of chromatin organization- simple “holders “ of chromatin structure- w/o sequence specificity H1, H2B, H2A, H3, H4 Nucleosome is a rigid, evolutionary conserved structure.

Histones variants More than what we have “guessed” Minor but important changes. Unlike H2A H2A.X is an indicator for Repair complexes after it is phosphorylated H2AZ is a destabilizing factor in highly transcribed regions, correlation with In order to have a functional kinetochore H3 variant (CENP-A ) is essential for

Cell Plasticity: Developmental process Embryonic stem cells (ESCs) within the inner cell mass of the early blastocyst have high levels of histones TH2A, TH2B, H2A.Z, and H3.3, and low levels of macroH2A. This is in contrast to differentiated cells H2A.Z appears crucial to both in establishing pluripotency and in facilitating differentiation TIGS 31, p516–527, 2015

Histones are modified post transcription Modifications are variable, not arbitrary, complex Specific loci Mostly on tails Sometimes together Reversible, dynamic Not all the possibilities are marked here S-Ser, K-Lys, R-Arg, T-Thr

The different modifications vary in size, polarity Have specific enzymes (modifying/demod.) Specific active AA tails (lysine) Options (methyl)

Context PTM occur within the chromatin structure (?) Writers: enzyme that modify: (Histone Acetylases HAT , Methylases HMT) Erasures: HDM, HDAC… Readers : proteins that recognize specific modification and have recognition specific domains/ PTM (bromodomain, PHD finger etc.) Mono-uniqitin is added but proteolyticly removed, Recruited by: histone modifications, Active proteins, DNA damage mechanism, RNA Recruit other factors, prevent binding Recruiting? Influence packaging , chromatin spatial-structure Enhance current signals: HDAC can enhance transcription repression (legends, CT and curcumin) HAT can enhance/ activate transcription

Analysis Which one of the former methodologies we discussed is feasible? Chip-Seq Mass analysis

Names and analysis Histone, localization of AA modifies, Modification

Language Histone tail Acetylation is correlated to transcription+ cumulative effect Happen on multiple Lysine residues Seen in correlation with repair Probably due to electrochemical charge Very short T1/2 Same enzyme HAT acts on many sites Responsive to many binding proteins through Bromodomain A cause to transcription of a by product? A signal enhancing that of other signals?

H3K4me3 modification is rich at transcribed promotors. KO of H3K4 methylate-transferase do not reduce transcription per-ce H3K36me in gene bodies correlates with transcribed regions but reduction has minimal effect. H3K9me3; H3K27me3 are correlated with silencing and heterochromatin formation probably by recruiting specific binding proteins

Turn- over through cell cycle Cellular memory During replication Nucleosomes are “falling off” and reassembled with the Pol machinery Special chaperones assist the reassembly Modified histones are maintained and reassembled Enzymes are recruited for the other newly formed nucleosomes Cell cycle dilution Is this is cellular memory?/ epigenetics?

Turn-over (localized) If you isolate highly active genes you can isolate both HAT and HDAC The idea is that the +/- modifications are highly dynamic Methylated histones (H3K4 H3K36 and H3K79) typical to transcription are enriched in the gene bodies + H2A.Z and H3.3: more dynamic nucleosomes structure. The appropriate enzymes and TF assist the process The inactivated X is enriched with H3K27me3. The Enzyme complex PRC2 that is recruited by the lncXist is defining the modification I Think that Histone modification are assisting / increasing signals of transcription but are not sole regulators

Human disorders Mutations changing a key player . Probably- non-viable Changes of typical PTM/ variant in cancer and cellular aging (including CCC) Coffin-Lowry mental retardation and dysmorphic phenotype is due to X-linked mutation of the RSK2 A kinase of CREB and Histone 3 A flux of reports regarding change of pattern in different disorders