Genetic and Molecular Studies of Drosophila Chromatin Remodeling Factors

核小体和染色质的其他成分通过以下方式作为真核转录的有效抑制因子 阻断转录因子和其他调节蛋白对DNA的访问。染色质抑制是 通过两种一般机制调节：依赖于ATP的染色质重塑和共价修饰 核小体组蛋白。这些过程的中断会导致各种人类疾病，包括 癌症。尽管在确定染色质的作用机制方面取得了巨大的进展- 重塑因子和组蛋白修饰酶，对它们在 真核转录与人类疾病。最近的研究表明，染色质重塑 因子可以调节高阶染色质结构，但它们在这一过程中的作用仍然很差。 明白了。为了解决这些重要问题，我们实验室使用了黑腹果蝇作为模型 系统研究三种不同的染色质重塑因子：BRM、KIS-L和ISWI。我们最近发现， Brm和Kis-L在核糖核酸聚合酶II的转录中起着全局作用。 BRM和KIS-L在转录中，我们将表征由于它们功能丧失而导致的缺陷，并绘制 它们在体内与之相互作用的活性基因的区域。我们还将检查特定地点是否 组蛋白尾部的甲基化影响BRM或KIS-L与其染色质底物相互作用的能力。 与BRM和KIS-L不同，ISWI在染色质压缩和转录抑制方面发挥着全球性的作用， 可能是通过促进连接物组蛋白H1与染色质的结合。为了检验这一假设，我们将 鉴定由ISWI功能丧失引起的染色体结构缺陷，并研究 使用RNAi和设计的显性-负性突变降低组蛋白H1功能的后果。至 确定参与调控高阶染色质结构的其他因素，我们将筛选 导致染色体缺陷的突变与ISWI突变体中观察到的突变类似。通过同时 在单个生物体中研究三种不同的染色质重塑因子，我们将获得更好的 了解它们在转录和其他过程中的作用。 与果蝇类似的染色质重塑因子-包括BRM、KIS-L和ISWI-存在于 在人类，它们在基因表达和发育中扮演着高度保守的角色。这些基因的突变 因子与多种癌症有关，包括横纹肌样瘤、乳腺癌和白血病。 因此，我们对遗传模式生物体中染色质重塑因子的研究应该有助于 这些人类癌症和其他疾病背后的分子机制。

Nucleosomes and other components of chromatin act as potent repressers of eukaryotic transcription by blocking the access of transcription factors and other regulatory proteins to DNA. Chromatin repression is regulated via two general mechanisms: ATP-dependent chromatin remodeling and the covalent modification of nucleosomal histones. The disruption of these processes leads to a variety of human diseases, including cancer. In spite of tremendous progress toward determining the mechanism of action of chromatin- remodeling factors and histone-modifying enzymes, much remains to be learned about their roles in eukaryotic transcription and human disease. Recent studies have suggested that chromatin-remodeling factors can regulate higher-order chromatin structure, but their role in this process remains poorly understood. To address these important issues, our laboratory uses Drosophila melanogaster as a model system to study three different chromatin-remodeling factors: BRM, KIS-L and ISWI. We recently found that BRM and KIS-L play global roles in transcription by RNA Polymerase II. To determine the precise roles of BRM and KIS-L in transcription, we will characterize defects resulting from their loss of function and map the regions of active genes with which they interact in vivo. We will also examine whether the site-specific methylation of histone tails affects the ability of BRM or KIS-L to interact with their chromatin substrates. Unlike BRM and KIS-L, ISWI plays a global role in chromatin compaction and transcriptional repression, possibly by promoting the association of the linker histone H1 with chromatin. To test this hypothesis, we will characterize defects in chromosome structure resulting from the loss of ISWIfunction and investigate the consequences of reducing histone H1 function using RNAi and engineered dominant-negative mutations. To identify other factors involved in the regulation of higher-order chromatin structure, we will screen for mutations that cause chromosome defects similar to those observed in ISWI mutants. By simultaneously studying three different chromatin-remodeling factors in a single organism, we will gain a much better understanding of their roles in transcription and other processes. Counterparts of Drosophila chromatin-remodeling factors - including BRM,KIS-L and ISWI - are present in humans, where they play highly conserved roles in gene expression and development. Mutations in these factors are associated with a variety of cancers, including rhabdoid tumors, breast cancer, and leukemias. Our studies of chromatin-remodeling factors in a genetic model organism should therefore shed light on the molecular mechanisms underlying these human cancers and other diseases.