RUI: Functional Analysis of the C. elegans Winged-Helix Transcription Factor, LIN-31

RUI：线虫翼螺旋转录因子 LIN-31 的功能分析

• 批准号: 0315072
• 负责人: Leilani Miller
• 金额: $ 31万
• 依托单位: Santa Clara University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0315072&HistoricalAwards=false
• 关键词: RUI; Functional; Analysis; C.; elegans

0315072MillerThe LIN-31 protein, a member of the winged-helix family of transcription factors, is required for the proper specification of vulval cell fates during C. elegans development. This protein is believed to play two roles during development: (1) to promote vulval cell fates (when phosphorylated by the signal transduction protein MAP kinase) and (2) to promote non-vulval cell fates (when heterodimerized with another transcription factor, LIN-1). This project aims to further investigate LIN-31's role in cell fate specification during vulval development by continuing a functional analysis of this multifaceted protein.The Specific Aims of this proposal are (1) Functional Analysis of the Winged Helix Transcription Factor LIN-31 (site-directed mutagenesis experiments will assign functions to different regions of the LIN-31 protein), (2) Identification of LIN- 31::LIN-1 Heterodimerization Elements (co-immunoprecipitation experiments will be used to determine if the acidic region of the LIN-31 protein is required for heterodimerization with the LIN-1 protein, a known heterodimerization partner), and (3) Identification of High-Affinity LIN-31 Binding Sequences (a sequential selection technique will be used to identify oligonucleotides that bind the LIN-31 DNA binding domain with high affinity--those sequences will then be used to identify actual targets of the LIN-31 transcription factor).The Intellectual Merit of this project is to provide a better understanding of how gene regulation controls development by advancing our knowledge of a transcription factor at the end of a signaling cascade that determines cell fates. In addition, by identifying potential targets of this transcription factor, the experiments described in this proposal will also illuminate a part of the C. elegans vulval cell fate pathway that is not well understood. The Broader Impacts of this project include significant contributions to teaching, training and learning. Undergraduate students will be an integral part of the project, involvement of the P.I. in research will enhance her teaching, and training of a Teaching Post-Doc will provide critical training in education and research, in addition to enhancing the evolving "community of science scholars" at SCU. Furthermore, of the 23 undergraduate students who have thus far conducted research in the P.I.'s laboratory, 17 (74%) were women, most of which have gone on to post-graduate programs. These numbers indicate established success in recruiting and educating members of underrepresented groups. SCU also has a significant Latino population, from which research students will continue to be recruited.

LIN-31蛋白是有翼螺旋转录因子家族中的一员，在线虫发育过程中对外阴细胞命运的正确指定是必需的。该蛋白被认为在发育过程中扮演两个角色：(1)促进外阴细胞的命运(当被信号转导蛋白MAP激酶磷酸化时)和(2)促进非外阴细胞的命运(当与另一个转录因子LIN-1异源二聚体时)。本项目旨在通过继续对这种多方面蛋白的功能分析，进一步研究LIN-31‘S在外阴发育过程中对细胞命运的调控作用。本项目的具体目标是(1)有翼螺旋转录因子LIN-31的功能分析(定点突变实验将把功能分配给LIN-31蛋白的不同区域)，(2)LIN-31：：LIN-1异二聚化元件的鉴定(将使用免疫共沉淀实验来确定LIN-31蛋白的酸性区域是否需要与已知的异二聚体LIN-1蛋白进行异源二聚反应)，以及(3)高亲和力LIN-31结合序列的鉴定(顺序选择技术将被用于识别以高亲和力结合LIN-31 DNA结合域的寡核苷酸-这些序列将被用于识别LIN-31转录因子的实际靶标)。这个项目的智力价值是通过提高我们对决定细胞命运的信号级联末端的转录因子的知识，更好地理解基因调控如何控制发育。此外，通过确定该转录因子的潜在靶点，本提案中描述的实验还将阐明线虫外阴细胞命运途径中尚未被很好了解的一部分。该项目的更广泛影响包括对教学、培训和学习的重大贡献。本科生将是该项目不可或缺的一部分，P.I.参与研究将加强她的教学，培训教学博士后将提供教育和研究方面的关键培训，此外还将加强SCU不断发展的“科学学者社区”。此外，到目前为止，在P.I.S实验室进行研究的23名本科生中，有17名(74%)是女性，其中大多数人已经攻读了研究生课程。这些数字表明，在招收和培养代表性不足的群体成员方面取得了既定的成功。南加州大学也有大量的拉丁裔学生，研究生会继续从他们那里招生。