ACETYLATION OF H4K4 IS CELL CYCLE REGULATED & MEDIATED BY HAT3 IN T BRUCEI

H4K4 的乙酰化受细胞周期调节

• 批准号: 7954099
• 负责人: GEORGE ALAN MARTIN CROSS
• 金额: $ 0.12万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7954099
• 关键词: Acetylation; Antibodies; Binding; Cell Cycle; Cell Nucleus; Cell Separation; Cells; Computer Retrieval of Information on Scientific Projects Database; Cycloheximide; Enzymes; Epigenetic Process; Eukaryota; Funding; Grant; Histone Acetylation; Histone Code; Histone H4; Histones; Horns; Immunofluorescence Microscopy; Individual; Institution; Lysine; Manuscripts; Masks; Mediating; Modification; Phase; Protein Synthesis Inhibitors; Publishing; Research; Research Personnel; Resources; Site; Source; Specific qualifier value; System; Transcriptional Regulation; Trypanosoma brucei brucei; United States National Institutes of Health; Western Blotting; histone acetyltransferase; histone modification; macromolecule

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Post-translational histone modifications have been studied intensively in several eukaryotes. It has been proposed that these modifications constitute a 'histone code' that specifies epigenetic information for transcription regulation. With a limited number of histone-modifying enzymes, implying less redundancy, Trypanosoma brucei represents an excellent system in which to investigate the function of individual histone modifications and histone-modifying enzymes. In this study, we characterized the acetylation of lysine 4 of histone H4 (H4K4), the most abundant acetylation site in T. brucei histones. Because of the large sequence divergence of T. brucei histones, we generated highly specific antibodies to acetylated and unmodified H4K4. Immunofluorescence microscopy and Western blots with sorted cells revealed a strong enrichment of unmodified H4K4 in S phase and suggested a G1/G0-specific masking of the site, owing to non-covalently binding factors. Finally, we showed that histone acetyltransferase 3 (HAT3) is responsible for H4K4 acetylation and that treatment of cells with the protein synthesis inhibitor cycloheximide led to an almost instantaneous loss of unmodified H4K4 sites. As HAT3 is located inside the nucleus, our findings suggest that newly synthesized histone H4 with an unmodified K4 is imported rapidly into the nucleus, where it is acetylated, possibly irreversibly. A manuscript describing these results has been published: Acetylation of histone H4K4 is cell cycle regulated and mediated by HAT3 in Trypanosoma brucei. Siegel TN, Kawahara T, Degrasse JA, Janzen CJ, Horn D, Cross GA. Mol Microbiol. 2008 Feb;67(4):762-71.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 翻译后组蛋白修饰已在几种真核生物中得到深入研究。有人提出，这些修饰构成了“组蛋白密码”，指定了转录调控的表观遗传信息。由于组蛋白修饰酶的数量有限，意味着冗余较少，布氏锥虫代表了一个研究个体组蛋白修饰和组蛋白修饰酶功能的优秀系统。在这项研究中，我们表征了组蛋白 H4 (H4K4) 赖氨酸 4 的乙酰化，这是布氏锥虫组蛋白中最丰富的乙酰化位点。由于布氏锥虫组蛋白的序列差异较大，我们生成了针对乙酰化和未修饰的 H4K4 的高度特异性抗体。免疫荧光显微镜和分选细胞的蛋白质印迹显示，S 期未修饰的 H4K4 大量富集，并表明由于非共价结合因子，该位点存在 G1/G0 特异性掩蔽。最后，我们发现组蛋白乙酰转移酶 3 (HAT3) 负责 H4K4 乙酰化，并且用蛋白质合成抑制剂放线菌酮处理细胞导致未修饰的 H4K4 位点几乎瞬时丢失。由于 HAT3 位于细胞核内部，我们的研究结果表明，新合成的具有未修饰的 K4 的组蛋白 H4 会快速输入细胞核，并在细胞核中被乙酰化，可能是不可逆的。描述这些结果的手稿已发表： 布氏锥虫中组蛋白 H4K4 的乙酰化是由 HAT3 调节和介导的细胞周期。西格尔 TN、川原 T、德格拉斯 JA、詹森 CJ、霍恩 D、克罗斯 GA。 摩尔微生物。 2008 年 2 月；67(4)：762-71。