Epigenetic regulation of cardiac MHC gene locus

描述（由申请人提供）：心脏MHC基因（1和2）的对立调控在甲状腺状态改变、糖尿病、压力过载和发育过程中是高度协调的，但这种调控的机制尚不清楚。在之前的研究中，我们发现了一种天然存在的反义2 RNA转录物，它从2和1基因之间的基因间隔（IGS）中间开始，向上游延伸到2MHC基因启动子区域，完全重叠于2基因。这种来自IGS的反义转录先前被认为在正常啮齿动物心脏中协调心脏MHC基因表达，并对甲状腺状态改变、糖尿病和压力过载做出反应。最近，通过链特异性RT-PCR对基因间RNA进行了更全面的分析，揭示了基因间RNA在意义方向上的存在，其转录方向指向1MHC基本启动子区，并继续通过1MHC基因（见图1）。这些结果（以及启动子报告分析的初步结果）有力地支持了IGS在啮齿动物心脏中双向转录活性的新概念。低链的转录，向上游的2MHC基因进行，产生反义RNA：一个可能干扰2基因转录的过程。上链的转录，从1MHC基因TSS上游约2kb开始，通过1启动子进入1基因内部，这一过程可能会增强1基因的转录。因此，我们假设基因间双向转录控制了相邻1和2 MHC基因的协调对立调节。本研究的目的是通过表观遗传机制，包括DNA甲基化、染色质重塑和组蛋白修饰，在改变心脏MHC基因位点上两个相邻基因表达的背景下，研究双向基因间转录的体内调控。为了比较的目的和作为我们理解这个基因位点的基因调控方法的一部分，研究还将涉及心房和慢骨骼肌。这些组织的独特之处在于前者主要表达1，而后者只表达微量的1，以2为主。因此，本研究将探索基因调控的新领域，并研究一个有趣的非编码基因间RNA调控机制，以及通过双向基因间转录对心脏MHC基因位点的表观遗传调控。

DESCRIPTION (provided by applicant): The antithetical regulation of the cardiac MHC genes (1 and 2) is highly coordinated in response to altered thyroid state, diabetes, pressure overload, and during development, yet the mechanism underlying this regulation is poorly understood. In previous studies, we discovered a naturally occurring antisense 2 RNA transcript that starts in the middle of the intergenic spacer (IGS) between the 2 and 1 genes and extends upstream to the 2MHC gene promoter region, fully overlapping the 2 gene. This antisense transcription originating from the IGS was previously proposed to coordinate cardiac MHC gene expression in normal rodent hearts and in response to altered thyroid state diabetes, and pressure overload. Recently, more comprehensive analyses of intergenic RNA via strand specific RT-PCR revealed the existence of intergenic RNA in the sense direction that is transcribed toward the 1MHC basic promoter region and continues through the 1MHC gene (see figure 1). These results (and preliminary results on promoter reporter assays) strongly support the novel concept that the IGS is transcriptionally active in both directions in rodent heart. Transcription of the lower strand, which proceeds upstream toward the 2MHC gene, produces antisense RNA: a process that may interfere with 2 gene transcription. Transcription of the upper strand, which starts from ~2kb upstream from the 1MHC gene TSS and proceeds through the 1 promoter to within the 1 gene: a process that may enhance 1 gene transcription. Thus, we hypothesize that the intergenic bidirectional transcription controls the coordinated antithetical regulation of adjacent 1 and 2 MHC genes. The goal of this proposed research is to examine the in vivo regulation of the bidirectional intergenic transcription in the context of altering the expression of the two adjacent genes on the cardiac MHC gene locus via an epigenetic mechanism which involves DNA methylation, chromatin remodeling and histone modification. For comparative purposes and as part of our approach to understanding the gene regulation on this locus, studies will also involve the atria and slow skeletal muscle. These tissues are unique in that the former expresses predominantly 1, while the latter expresses only traces of 1, with 2 being predominant. Consequently, this research will explore a new area of gene regulation and investigate an intriguing regulatory mechanism involving non-coding intergenic RNA, and epigenetic regulation of the cardiac MHC gene locus via bidirectional intergenic transcription.