The role of histone H2Az in cardiac gene expression

描述（由申请人提供）：细胞基因表达谱的变化，是其表型变化的基础，是对广泛的生理或病理刺激的反应。从细胞膜到细胞核，这一过程在多个层面上都受到严格的调控。许多上游信号通路和调控转录机制的第二信使继续被大量研究。最终结果不仅包括转录因子的修饰，还包括重塑、翻译后改变和/或组蛋白的变异替代。虽然组蛋白的主要作用是建筑，涉及染色质包装，但越来越明显的是，它在调节基因转录方面也起着相互作用。然而，在哺乳动物细胞中，特别是在心肌细胞中，所涉及的机制仍然知之甚少。我们以前报道过，组蛋白H2A的一种特定亚型，称为H2Az，在心脏肥厚期间上调。我们的初步结果表明，在新生小鼠的所有器官中，这种组蛋白的表达水平相对较高，但仅在正常成年小鼠的心脏和骨骼肌中，其表达水平降至无法检测的水平。H2Az的功能对发育至关重要，非冗余且高度保守，其机制仍有待于在哺乳动物细胞中研究。在低等真核生物中，它与核小体重塑和转录激活有关。本研究将探讨其在心脏肥厚过程中的作用。基于我们的初步结果，我们假设H2Az通过直接调节一部分生长相关基因的表达是肥厚生长所必需的。这项奖助金的具体目的是：1。采用诱变和RNA干扰的方法研究H2Az在心肌肥厚中的作用及机制。2. 利用染色质免疫沉淀（ChIP）和减法杂交技术鉴定受H2Az调控的基因。3. 利用酵母双杂交系统鉴定H2Az调控/效应蛋白及其功能。4. 描述在心脏中过表达H2Az的转基因小鼠模型。

DESCRIPTION (provided by applicant): Changes in the gene expression profile of a cell, underlie changes in its phenotype, in response to a broad range of physiological or pathological stimuli. This process is very tightly regulated at multiple levels, from the membrane to the nucleus. Many of the upstream signaling pathways and second messengers that regulate the transcriptional machinery continue to be heavily investigated. The end result does not only include modification of transcription factors, but in addition, remodeling, post-translational alterations, and/or variant replacements of histones. While the main role of histones is architectural, involving chromatin packaging, it is becoming increasingly evident that it also plays an interactive role in regulating gene transcription. The mechanisms involved, though, remain poorly understood in mammalian cells in general or in cardiac myocytes in particular. We have previously reported that a specific isoform of histone H2A, termed H2Az, is upregulated during cardiac hypertrophy. Our preliminary results show that in the neonatal mouse this histone is expressed at relatively high levels in all organs but drop to undetectable amounts in the normal adult heart and skeletal muscle only. The functions of H2Az are essential for development, non-redundant, and highly conserved, the mechanisms of which remain to be investigated in mammalian cells. In lower eukaryotes though, it has been specifically implicated in nucleosomal remodeling and transcriptional activation. This proposal will investigate its role in the heart during cardiac hypertrophy. Built on our preliminary results, we hypothesize that H2Az is necessary for hypertrophic growth through directly regulating the expression of a subset of growth-related genes. The specific aims of this grant are: 1. To determine the role and mechanism of function of H2Az in cardiac hypertrophy, using mutagenesis and RNA interference. 2. To identify genes that are regulated by H2Az, using chromatin immunoprecipitation (ChIP) and subtractive hybridization. 3. To identify H2Az regulatory/effector proteins and their functions, using the yeast two-hybrid system. 4. Characterize a transgenic mouse model over-expressing H2Az in the heart.