Role of MacroH2A in Estrogen-Regulated Gene Expression

描述(申请人提供)：为了使雌激素受体(ER)激活基因转录，它必须克服抑制染色质结构。ER通过招募启动子酶来实现这一点，这些酶要么共价修改染色质，要么将其重塑为更允许的状态。直到最近才变得明显的是，所有的染色质都不是同质的。在基因组的特定区域，组蛋白变体取代了它们的典型组蛋白对应。组蛋白变异对基因调控的影响在很大程度上是未知的。然而，初步数据表明，组蛋白变异体宏H_2A参与了ER调控的乳腺癌细胞的转录。MacroH2A还与X失活过程中发生的表观遗传转录沉默有关。最近发现，大分子H_2A能与烟酰胺腺嘌呤二核苷酸(NAD+)代谢产物ADP-核糖(ADPR)和O-乙酰-ADP-核糖(OAADPR)结合。这一发现表明，在两个依赖于NAD+的途径中，宏H2A是刺激信号事件的目标：聚(ADP-核糖)聚合酶(PARP)合成的聚(ADP-核糖)(PAR)链由聚(ADP-核糖)糖水解酶(PAR)翻转和蛋白质由SIRT1脱乙酰化。这些研究的目的是为了更好地理解雌激素靶基因(目标1)中大分子H_2A掺入的原因以及这些基因中大分子H_2A掺入的影响(目标2)。为此，将使用各种基于细胞的生化和生物信息学方法，包括染色质免疫沉淀(ChIP)、ChlP-芯片、表达分析、RNAi、蛋白质-蛋白质相互作用分析、酶分析和DMA基序寻找算法。雌激素通过内质网传递信号，在性发育和生育等正常生理过程中发挥重要作用。然而，雌激素也促进乳腺癌的生长和进展，并对心脏病、中风和骨质疏松症具有保护作用。因此，了解大分子H_2A调控ER靶基因转录的机制将有助于深入了解这些疾病的进展。

DESCRIPTION (provided by applicant): In order for estrogen receptor (ER) to activate gene transcription it must overcome a repressive chromatin structure. ER does this by recruiting to the promoter enzymes that either covalently modify chromatin or remodel it into a more permissive state. Only recently has it become apparent that all chromatin is not homogenous. Histone variants replace their canonical histone counterparts in specific regions of the genome. The effect of histone variants on gene regulation is largely unknown. However, preliminary data indicates that the histone variant macroH2A is involved in ER-regulated transcription in breast cancer cells. MacroH2A has also been implicated in the epigenetic silencing of transcription that occurs during X inactivation. Recently, macroH2A was shown to bind to the nicotinamide adenine dinucleotide (NAD+) metabolites ADP-ribose (ADPR) and O-acetyl-ADP-ribose (OAADPR). This finding implicates macroH2A as a target of exciting signaling events in two NAD+-dependent pathways: the turnover of poly(ADP-ribose) polymerase (PARP)-synthesized poly(ADP-ribose) (PAR) chains by poly(ADP-ribose) glycohydrolase (PARC) and deacetylation of proteins by SIRT1. The objective of these studies is to achieve a better understanding of both the causes of macroH2A incorporation at estrogen target genes (Aim 1) and effects of macroH2A incorporation at these genes (Aim 2). Toward this end, a variety of biochemical cell-based,and bioinformatic approaches will be used including Chromatin immunoprecipition (ChIP), ChlP-chip, expression analysis, RNAi, protein-protein interaction assays, enzyme assays, and DMA motif finding algorithms. Estrogen signaling through the ER plays important roles in normal physiological processes such as sexual development and fertility. However, estrogen also promotes the growth and progression of breast cancer and has a protective effect from heart disease, stroke and osteoporosis. Therefore, an understanding of the mechanism by which macroH2A regulates the transcription of ER target genes will provide insights into the progression of these diseases.