HDAC4調控D型肝炎病毒反向基因體複製之研究

The regulation of HDV antigenomic RNA replication by HDAC4

陳慧茹(Hui-Ru Chen)

指導教授 ： 陳培哲

國立臺灣大學/醫學院/微生物學研究所/碩士(2011年)

https://doi.org/10.6342/NTU.2011.02245

摘要

D型肝炎病毒是B型肝炎病毒的衛星病毒，屬於單股負向的RNA病毒。它可以在細胞中複製自己的基因體，不需要依賴B型肝炎病毒。過去的研究已知，D型肝炎病毒的基因體複製是由宿主細胞的RNA polymerase II來執行，並且受到自己的小型D型肝炎抗原所活化。進一步地發現，小型D型肝炎抗原的後轉譯修飾，尤其是第177位置Serine的磷酸化、第72位置Lysine的乙醯化、第13位置Arginine的甲基化，在D型肝炎病毒反向基因體複製中扮演重要的角色。細胞中的histone acetylase p300被提出會對小型D型肝炎抗原乙醯化。由於乙醯化在Lysine的的 alpha-amino group 是高度動態且可逆的反應，細胞中也許有histone deacetylase會對小型D型肝炎抗原進行去乙醯化，並且調節D型肝炎病毒基因體複製。因此，本論文便針對是否有histone deacetylase參與D型肝炎病毒基因體複製、哪一個histone deacetylase負責執行、並探究其機制。首先，我們發現D型肝炎病毒基因體的量隨著histone deacetylase抑制劑的量增加而增加，而反向基因體的量則否，意味著細胞中可能有histone deacetylase參與調控D型肝炎病毒基因體量。進一步地，大量表達histone deacetylase 4可以顯著地降低D型肝炎病毒基因體的量，而其他的histone deacetylase則否。更進一步發現，小型D型肝炎抗原的乙醯化可以被大量表達的histone deacetylase 4所降低，並且小型D型肝炎抗原可以與內生性的histone deacetylase 4交互作用。而且，大量表達histone deacetylase 4會干擾小型D型肝炎抗原與RNA polymerase II的交互作用，而且增加了小型D型肝炎抗原第177位置Serine的磷酸化。總結，這些結果代表著D型肝炎病毒反向基因體的複製，至少有一部分，可能透過histone deacetylase 4與小型D型肝炎抗原形成複合體，調節小型D型肝炎抗原的乙醯化，並調控小型D型肝炎抗原與RNA polymerase II的交互作用。

關鍵字

D型肝炎病毒 ； 小型D型肝炎抗原 ； 複製 ； 乙醯化 ； histone deacetylase 4

並列摘要

Hepatitis delta virus (HDV) is a satellite virus of HBV and belongs to single-, negative-stranded RNA virus. It can replicate its genome by itself without the help of HBV. Previous studies have shown that HDV RNA genome replication is carried out by the host RNA polymerase II (RNAPII) and is activated by its small delta antigen (S-HDAg). Further, the post-translational modifications (PTMs) of S-HDAg, especially Ser-177 phosphorylation, Lys-72 acetylation,and Arg-13 methylation, play important roles in HDV antigenomic RNA replication. Cellular histone acetylase p300 is proposed to be one of enzymes that acetylate S-HDAg. Since acetylation of the alpha-amino group of lysine residues is highly dynamic and reversible, cellular deacetylases may deacetylate S-HDAg to modulate HDV RNA replication. Hence, this study focuses on issues as: whether cellular deacetylases were involved in the regulation of HDV RNA replication, which cellular deacetylase(s) was/were doing the job, and the mechanism by which the deacetylases(s) regulated HDV RNA replication. First, HDV genomic RNA level, but not HDV antigenomic RNA level, was increased by distinct class of HDAC inhibitors, suggesting that HDACs may participate in the regulation of HDV genomic RNA level. Second, ectopic expression of HDAC4, but not other class I and II HDACs, significantly decreased HDV genomic RNA level. The total acetylation level of S-HDAg was decreased by ectopic expression of HDAC4. S-HDAg was co-immunoprecipitated with endogenous HDAC4 in HDV-replicating cells. Moreover, ectopic expression of HDAC4 abrogated the association between S-HDAg and RNAP II, and enhanced Ser177 phosphorylation level. Taken together, these results suggested that HDAC4 might regulate HDV antigenomic RNA replication, at least in part, via forming complexes with S-HDAg, modulating the acetylation level of S-HDAg and regulating the interaction between S-HDAg and RNAP II.

並列關鍵字

hepatitis delta virus ； small hepatitis delta antigen ； replication ； acetylation ； histone deacetylase 4

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