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Shelley Berger The Wistar Institute University of Pennsylvania

Bio 483 October 9 , 2012. Overview of Histone Variants. Shelley Berger Biology; CDB; Genetics Penn Epigenetics Program bergers@upenn.edu. Shelley Berger The Wistar Institute University of Pennsylvania. Lecture Outline. What are histone variants?

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Shelley Berger The Wistar Institute University of Pennsylvania

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  1. Bio 483 October 9, 2012 Overview of Histone Variants Shelley Berger Biology; CDB; Genetics Penn Epigenetics Program bergers@upenn.edu Shelley Berger The Wistar Institute University of Pennsylvania

  2. Lecture Outline What are histone variants? Review nuclesome structure wrt replacement Discuss H2A: macroH2A, H2A.Bbd, H2Ax, H2AZ H3: CENPA, H3.3 testes histone variants Consideration of following issues: - sequence variation - nucleosome structure - genome function - location within the genome - physiological function

  3. Core vs. variant histones H3 globular N C H4 H2A H2B Are there additional histones?

  4. H3 H4 H2A H2B H1

  5. H3 H4 H2A H3 ‘tail’ H2B Luger and Richmond, Nature 1997

  6. Newnucleosome formation: after replication H3/H4 tetramer binds to DNA 2 dimers H2A/H2B bind H3/H4 tetramer

  7. How to replace? During replication assemble with variant histones H2A/H2B dimer exchange Disassembly of nucleosome during DNA damage repair, transcription

  8. DNA repair Gene repression X chrominact Gene activation H2A H2B Gene activation H3 H4 Centromere identity

  9. H2A

  10. Chromatin compaction by macroH2A

  11. Chromatin destabilization by H2A.Bbd

  12. H2A H2Ax Histone H2Ax in DNA damage pathway

  13. DNA damage leads to complex repair pathways

  14. Molecular details of DNA repair

  15. H2Ax evolutionary conservation

  16. H2Ax phosphorylation is an early step in the repair process

  17. Histone H2AZ in gene regulation HistoneH2AZ (= HTZ1): What is its localization in the genome? What genomic process does it regulate? How does it affect NCP structure and positioning?

  18. TSS Genome-wide view of H2AZ at promotors TSS Gene ORF Promoter H2AZ tends to occur in one nuc up- and one nuc down-stream of the TSS Does it regulate transcription? Nuc “Depleted” Region

  19. H2AZ is released as genes are activated Its release is required for full activation

  20. Is there a difference in nucleosome stability? Association of H2A/H2AZ with chromatin H2AZ is more loosely bound to chromatin (although NCP structure very similar)

  21. What is the localization of H2AZ around active/inactive genes? Active genes less enriched than inactive

  22. H2AZ What is its localization in the genome? Inactive promoters What genomic process does it regulate? Poises genes for activation How does it affect NCP structure and positioning? DestablizesNCP: correlates with depletion of one NCP over the TSS How is it incorporated? …a digression

  23. ATP-dependent Nucleosome Remodeling Nucleosomes can be changed in detailed association with DNA, position, presence

  24. ADP + Pi ATP Activities of ATP-dependent chromatin remodeling enzymes ATP Remodeler

  25. Evolutionary Tree of SWI2/SNF2 Remodelers INO80 ino80-sc ISW1-Sc ISW2-Sc ISWI SNF2L-Hs SNF2H-Hs ISWI-Dm CHD5-Hs CHD4-Hs CHD3-Hs MI2-Dm CHD CHD1-Sc CHD2-Hs CHD1-Hs CHD-Dm STH1-Sc SWI/SNF SWI2-I Brahma-Dm BRG-1-Hs BRM-Hs

  26. ATP dependent remodelers and Histone Variant deposition Swr1 H2AZ Start with H2A NCPs

  27. What do ATP-dependent remodeling complexes do? 1. Directly alter NCP position 2. Exchange histones and histonevariants: New paradigm!

  28. Model for H2AZ Biochem. Cell Biolvol 84 2006

  29. Instability of H2AZ NCPs allows high temp to turn on/off genes in plants

  30. Histone H3 and its Variants Replication Coupled deposition (only S phase) CenpA Replication independent Deposition (not strictly during S phase)

  31. Histone H3.1 and H3.3 H3/H3.1 H3.3 H4

  32. H3.3 is very similar to H3.1 H3.3 is more ancient

  33. What genomic processes do H3.1/H3.3 regulate? Replication Dependent: Either H3.1 or H3.3 Replication Independent: Only H3.3

  34. H3.3 localizes with RNAPII and K4me H3.3 H3.1 K4me2 RNAP genes

  35. How are H3.1 and H3.3 deposited in RC and RI pathways? H3.1 H3.3 p150 p60 HIRA IP-Western Two different chaperone/ assembly factors

  36. Nucleosome assembly RC or RI

  37. Histone CENPA H3 CENPA H4

  38. What are centromeres?

  39. CENPA, an H3 variant, is in a specialized NCP at centromeres in all eukaryotes Sequence complexity

  40. Targeting of CENPA to centromeres Mapping in vitro and in vivo defined a region in the histone fold as “CATD”

  41. Chaperone for Centromeric Localization Cell line expressing CENPA-TAP

  42. HJURP required for CENPA targeting to centromeres 1. OE CENPA did not restore targeting 2. H3-CATD recruited by HJURP HJURP Knock Down cell line No targeting of CENPA in HJURP KD

  43. Controversy regarding structure of the centromeric CENPA NCP Proposed structures:

  44. Is there a distinct structure? (CENPA-H4)2 is more compact

  45. Structure of CENPA NCP Structural features map to CATD! It forms a well behaved octamer

  46. Summary: H3 variants and their functions CenpA HJURP HIRA CAF1

  47. Spermatogenesis and chromatin compaction: key role for histone variants

  48. Testes-specific histone variants replace core histones following meiosis

  49. All histones (except H4) have testes variants

  50. A remarkable variety of histone variants to regulate the genome via location-specific nuclesomefuntion H3 H4 H2A H2B H1

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