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，响应于广泛的生理或病理刺激。这一过程在多个层面上受到非常严格的调控，从细胞膜到细胞核。许多上游信号通路和调节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，而且还包括组蛋白的重塑、翻译后改变和/或变体替换。虽然组蛋白的主要作用是结构性的，涉及染色质包装，但越来越明显的是，它在调节基因转录中也起着相互作用。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。我们以前曾报道，组蛋白H2 A，称为H2 Az，一个特定的异构体，在心肌肥厚过程中上调。我们的初步结果表明，在新生小鼠中，这种组蛋白在所有器官中的表达水平相对较高，但仅在正常成年心脏和骨骼肌中降至不可检测的水平。H2 Az的功能是发育所必需的、非冗余的和高度保守的，其机制仍有待于在哺乳动物细胞中研究。然而，在低等真核生物中，它特别涉及核小体重塑和转录激活。本提案将探讨其在心脏肥大过程中的作用。基于我们的初步结果，我们假设H2 Az通过直接调节生长相关基因的一个子集的表达对肥大性生长是必要的。该补助金的具体目标是：1.采用基因突变和RNA干扰技术，研究H2 Az在心肌肥厚中的作用和机制。2.利用染色质免疫沉淀（ChIP）和消减杂交鉴定受H2 Az调控的基因。3.利用酵母双杂交系统鉴定H2 Az调控/效应蛋白及其功能。4.表征心脏中过表达H2 Az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.