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

LIN-31蛋白是转录因子翼螺旋家族的成员，在C. elegans发育 这种蛋白质被认为在发育过程中起两种作用：（1）促进外阴细胞命运（当被信号转导蛋白MAP激酶磷酸化时）和（2）促进非外阴细胞命运（当与另一种转录因子LIN-1异二聚化时）。本项目旨在通过对LIN-31的功能分析，进一步研究LIN-31在外阴发育过程中的细胞命运调控作用，具体目的如下：（1）有翼障碍转录因子LIN-31的功能分析（定点诱变实验将功能分配给LIN-31蛋白的不同区域），（2）LIN- 31的鉴定：LIN-1异源二聚化元件（将使用免疫共沉淀实验来确定LIN-31蛋白的酸性区域是否是与LIN-1蛋白（已知的异源二聚化配偶体）异源二聚化所需的），和（3）高亲和力LIN-31结合序列的鉴定（将使用顺序选择技术来鉴定以高亲和力结合LIN-31 DNA结合结构域的寡核苷酸-然后将使用那些序列来鉴定LIN-31 DNA结合结构域的实际靶。该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认识，提供对基因调控如何控制发育的更好理解。此外，通过鉴定该转录因子的潜在靶点，本实验也将阐明C.线虫外阴细胞的命运途径，这是不是很清楚。该项目的更广泛影响包括对教学、培训和学习的重大贡献。本科生将是该项目的一个组成部分，参与的P.I.在研究将提高她的教学，和教学博士后的培训将提供教育和研究的关键培训，除了加强在SCU不断发展的“科学学者社区”。此外，在迄今为止在P.I.进行研究的23名本科生中，在该实验室，17名（74%）是女性，其中大多数已经进入研究生课程。这些数字表明，在征聘和教育代表性不足群体的成员方面取得了成功。 SCU也有一个重要的拉丁裔人口，从研究学生将继续招募。

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.