open access

https://doi.org/10.14943/doctoral.k14906

https://hdl.handle.net/2115/85255

ヘテロクロマチンにおけるヒストンバリアントH2A.Zの機能に関する研究

英語

2022-03-24

Hokkaido University

81p

Heterochromatin, a silent chromatin is involved in many biological processes, such as gene silencing, development, chromosome segregation, and genome integrity. Post translational modifications of histone tails are crucial for heterochromatin formation. Methylation of histone H3 at Ly9 (H3K9me) is a well conserved heterochromatic modification. For the formation and maintenance of heterochromatin, RNAi-dependent and -independent pathways redundantly functions and recruits H3K9 methyltransferase Clr4 to heterochromatic region. HP1 recognizes H3K9me and forms highly condensed chromatin structure. HP1 recruits not only silencing factors but also anti-silencing factor Epe1, a H3K9me demethylase, to heterochromatic regions, suggesting that the competition between silencing factors and anti-silencing factors is important for proper maintenance of heterochromatin. As well as histone modifications, histone variants also play important roles. Especially, H2A.Z is one of the most evolutionally conserved histone H2A variant. H2A.Z has been implicated in many biological processes, including gene regulation, mammalian development, DNA replication, and stress response. In particular, the function of H2A.Z in euchromatic gene regulation is well studied. H2A.Z is highly enriched at gene promoter and is required for the proper transcription. However, its function and deposition mechanism in heterochromatin is still unclear. Here, I show that H2A.Z plays multiple roles in fission yeast heterochromatin. In fission yeast, H2A.Z is loaded into nucleosomes at TSS of euchromatic gene by the H2A.Z loader, SWR complex. While a small amount of H2A.Z localizes at pericentromeric heterochromatin compared to euchromatic gene promoter, loss of H3K9me induces the accumulation of H2A.Z, which is dependent on the SWR complex. The accumulated H2A.Z suppresses heterochromatic non coding RNA transcription. This transcriptional repression activity requires the N terminal tail of H2A.Z, which is involved in the regulation of euchromatic gene transcription. Additionally, RNAi defective cells, in which a substantial amount of H3K9me is retained by RNAi-independent heterochromatin assembly, also accumulate H2A.Z at pericentromeric heterochromatin. The additional loss of H2A.Z in these cells abolishes H3K9me. Moreover, loss of H2A.Z in RNAi defective cells decreased H3K9me at the mat locus heterochromatin and subtelomere heterochromatin, where RNAi is dispensable. However, H2A.Z is not required for the maintenance of artificially formed ectopic heterochromatin which is maintained by RNAi-independent mechanism in the absence of Epe1, suggesting the indirect effects of H2A.Z for RNAi-independent heterochromatin assembly. As Epe1, an eraser of H3K9me is responsible for the decrease of H3K9me in RNAi mutants, loss of Epe1 restored the H3K9me in RNAi and H2A.Z double deletion mutants. These results suggest that H2A.Z facilitates RNAi-independent heterochromatin assembly by antagonizing the demethylation activity of Epe1. Furthermore, loss of H2A.Z caused silencing defects at subtelomeric heterochromatin without affecting H3K9me. Since Epe1 also has a transcription activation domain at N-terminus, overexpression of Epe1 results in transcriptional activation at pericentromere without affecting H3K9me. Loss of H2A.Z in Epe1- overexpressig cells synergistically increases the transcription, and the subtelomeric silencing defect in H2A.Z deletion mutant is suppressed by loss of Epe1. These results suggests that H2A.Z suppresses Epe1-mediated transcriptional activation, which is required for subtelomeric gene repression. Taken together, this study provides novel evidence that H2A.Z plays diverse roles in chromatin silencing.

2022-03-24

甲第14906号

博士(理学)

(主査) 教授 坂口 和靖, 特任教授 髙木 睦, 教授 髙岡 晃教, 教授 村上 洋太

総合化学院（総合化学専攻）

博士論文

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