Transcriptional Activation by Reorganizing Chromatin

描述(申请人提供)：染色质重塑可作为 多种细胞过程中的功能关键，其中之一是调节 通过促进转录复合体的形成和 转录复合体的伸长。有几个有很好记录的 染色质重塑复合体与 基因特异的转录因子，使DNA可供 转录机器。此外，核小体结构是一种严重的 对生产所需基因重排的威慑 免疫球蛋白。染色质重塑显然是一种用来紧密 调节脊椎动物的免疫系统，可能是分子的关键 15年前提出的“可及性假说”背后的机制。 染色质重塑也参与细胞周期控制，并与 肿瘤抑制蛋白Rb或视网膜母细胞瘤蛋白。在理解如何 SWI/SNF和ISW2重塑核小体，重要的是要知道它确实是这样做的 不是在染色质上随机工作，但他们被招募或针对特定的 通过基因特定的转录因子或抑制物来定位。证据 表明染色质重塑可以与DNA紧密协调 DNA甲基化和DNA复制等修饰。这份名单 与染色质重塑有关的疾病继续增长，包括 横纹肌样肿瘤是一种侵袭性很强的儿科癌症， 乳腺癌、白血病、智力低下、威廉姆斯综合征和Rett 综合症。目前尚不清楚SWI/SNF的哪些亚基与 转录激活剂或其与核小体的相互作用 与不分青红皂白地与核小体结合不同。我们的 研究计划是检查结构及其与功能的关系 SWI/SNF染色质重塑复合体通过一系列方法 修饰的DNA或修饰的组蛋白八聚体。我们将获得3维的 用电子断层扫描研究SWI/SNF的结构并确定哪些区域 通过连接来自定点的数据与DNA和组蛋白八聚体相互作用 对结构进行光亲和标记和蛋白质降解。接下来，我们将 确定SWI/SNF和ISW2在招募到特定的 通过各自的“靶向”蛋白质在核小体阵列中定位。数据 这两种不同的染色质重塑复合体表明它们 在体内和以显著不同的方式调节染色质结构 在试管中。

DESCRIPTION (provided by applicant): Chromatin remodeling serves as a functional key in multiple cellular processes, one of them being the regulation of gene expression through promoting formation of the transcription complex and elongation of the transcription complex. There are several well-documented examples of chromatin remodeling complexes working in conjunction with gene-specific transcription factors to make the DNA accessible to the transcription machinery. In addition, the nucleosome structure is a severe deterrent to the rearrangement of genes required for the production of immunoglobulins. Chromatin remodeling is apparently a mechanism used to tightly regulate vertebrate immune systems and is probably the key to the molecular mechanism underlying the "accessibility hypothesis" proposed 15 years ago. Chromatin remodeling is also involved in cell cycle control and interacts with the tumor suppressor protein Rb or retinoblastoma protein. In understanding how SWI/SNF and ISW2 remodel the nucleosome, it is important to know that it does not work randomly on chromatin, but they are recruited or targeted to specific locations by gene-specific transcription factors or repressors. Evidence indicates that chromatin remodeling can be tightly coordinated with DNA modifications such as methylation of DNA and DNA replication. The list of diseases linked to chromatin remodeling continues to grow and includes such diseases as rhabdoid tumours, a very aggressive form of pediatric cancers, breast cancer, leukemia, mental retardation, Williams syndrome, and Rett syndrome. It is not known which subunits of SWI/SNF interact with the transcription activator or how its interaction with the nucleosome may be different when recruited versus indiscriminate binding to nucleosomes. Our research plan is to examine the structure and its relation to function of the SWI/SNF chromatin remodeling complex by a series of approaches that uses either modified DNA or modified histone octamers. We will obtain the 3-dimensional structure of SWI/SNF by electron tomography and determine which regions interact with DNA and histone octamer by linking data from site-directed photoaffinity labeling and proteolysis to the structure. Next, we will determine how SWI/SNF and ISW2 remodel chromatin when recruited to specific sites within nucleosomal arrays by their respective "targeting" proteins. Data on these two different chromatin remodeling complexes suggest that they modulate chromatin structure in significantly different ways both in vivo and in vitro.