RUI: Analysis of the Role of CHD1 in Chromatin Structure and Transcription

染色质重塑因子利用三磷酸腺苷的能量调节染色质的结构，从而调节转录、复制和修复等过程。虽然这些ATPase在体外的作用机制已经取得了重大进展，但关于它们在体内的活性仍有许多未知之处。PI的长期目标是了解CHD1染色质重塑和组装因子在体内是如何调节和靶向的。果蝇是研究这种特殊蛋白作用的理想模型系统。智力优势：本研究的目的是利用果蝇遗传学的强大工具来研究CHD1，一种色域、解旋酶和DNA结合域蛋白的体内活性。CHD1是一种特殊的ATPase，它在体外既具有核小体重塑又具有核小体组装活性。这些实验解决了特定的假设，即CHD1在全球范围内发挥作用，重新组装活性基因上的染色质，这一活性导致转录抑制。这一假说将通过追求三个具体目标来解决：(1)通过检测功能获得和功能丧失的突变体的表型来研究chd1的体内功能。(2)确定每个保守蛋白质结构域的相对贡献率。(3)研究CHD1ATPase活性的增加和丧失对染色体结构和RNA聚合酶II的影响。PI将结合遗传和细胞生物学的方法来分析突变的表型、染色体形态和转录。这些成熟的技术适用于本科生的研究，并已被PI广泛用于她对相关染色质重塑因子Brahma和ISWI的研究。PI的初步结果和专业知识使她处于实现这些特定目标的独特位置，其结果应该为CHD1的功能和调节提供重要的见解，CHD1是一种高度保守但鲜为人知的ATPA。广泛影响：这项研究将主要由克莱蒙特·麦肯纳学院、斯克里普斯学院和匹泽学院的本科生进行。在她在联合科学部的头三年里，PI在实验室亲自培训了15名本科生(13名女性和2名男性)，其中包括两名拉丁裔第一代大学生。学生在研究实验室的平均时间为两年。在已毕业的PIS研究学生中，100%在研究生院或医学院继续他们的科学教育。在这项研究的三年时间里，PI将在研究实验室亲自培训12-15名本科生，为他们提供帮助他们在科学职业生涯中取得成功的技能。PI位于一个独特的系(联合科学系)，该系由三所顶级文科院校共享。斯克里普斯学院是一所全女子学院，而克莱蒙特·麦肯纳学院和皮策学院在种族多元化程度最高的文理学院中分别排名第五和第七。这种多样化的学生群体使PI能够鼓励来自历史上代表性不足的群体的理科学生。除了为本科生提供研究机会，这项研究还将允许本科生参加一年一度的果蝇研究会议，以及南加州本科生研究会议。PI的一名本科生是该实验室最新出版的《遗传学》的合著者。这项瑞的研究将为PI和她的学生提供资源，以继续他们目前使用现代分子遗传技术对染色质重塑蛋白进行的竞争性研究。

Chromatin remodeling factors use the energy of ATP to modulate the structure of chromatin, thereby regulating processes such as transcription, replication, and repair. While significant advances have been made in regards to the mechanism of action of these ATPases in vitro, much remains unknown regarding their activities in vivo. The PI's long-term goal is to understand how the CHD1 chromatin remodeling and assembly factor is regulated and targeted in vivo. Drosophila melanogaster is an ideal model system in which to study the actions of this unusual protein.Intellectual merit: The goal of this research is to use the powerful tools of Drosophila genetics to investigate the in vivo activities of CHD1, a chromodomain, helicase, and DNA-binding domain protein. CHD1 is an unusual ATPase in that it possesses both nucleosome remodeling and nucleosome assembly activities in vitro. These experiments address the specific hypothesis that CHD1 functions globally to reassemble chromatin on active genes, an activity that results in the inhibition of transcription. This hypothesis will be addressed by pursuing three specific aims (1) Investigate the in vivo function of CHD1 by examining phenotypes of both gain of function and loss of function chd1 mutants. (2) Determine the relative contributions of each of the conserved protein domains. (3) Examine the consequences of both gain and loss of CHD1 ATPase activity on chromosome structure and RNA Polymerase II. The PI will employ a combination of genetic and cell biological approaches to analyze mutant phenotypes, chromosome morphology, and transcription. These well-established techniques are amenable to undergraduate research, and have been extensively used by the PI in her studies of related chromatin remodeling factors Brahma and ISWI. The PI's preliminary result and expertise place her in a unique position to accomplish these specific aims, the results of which should provide significant insights into the function and regulation of CHD1, a highly conserved yet poorly understood ATPase.Broader impacts: The research will be carried out primarily by undergraduate science students from Claremont McKenna, Scripps, and Pitzer Colleges. During her first three years in the Joint Science Department, the PI personally trained 15 undergraduates in the laboratory (13 women and two men), including two Latina first-generation college students. Students spend an average of two years in the research laboratory. Of the PIs research students who have graduated, 100% are continuing their science education in graduate or medical schools. Over the three years of this study, the PI will personally train an additional 12-15 undergraduates in the research laboratory, providing them with skills to help them succeed in their careers in science. The PI is in a unique department (The Joint Science Department) that is shared by three top-tier liberal arts institutions. Scripps is an all-women college, while Claremont McKenna and Pitzer College are the 5th and 7th most ethnically diverse liberal arts colleges in the top tier. This diverse student body allows the PI to encourage science students from historically underrepresented groups. In addition to providing research opportunities for undergraduates, this research will allow undergraduates to attend the Annual Drosophila Research Conference in addition to the Southern California Conference for Undergraduate Research. One of the PI's undergraduate research students is a co-author on the lab's most recent publication in Genetics. This RUI research will provide resources for the PI and her students to continue their current, competitive research into chromatin remodeling proteins using modern molecular genetic techniques.