Molecular identification and functional characterization of fried, a gene require

描述(由申请人提供)： 我的实验室的长期目标是提高我们对果蝇发育过程中的生长、新陈代谢和细胞周期调节的理解。本研究主要对油炸食品的分子鉴定和功能鉴定进行了研究。这三个油炸突变体是在相同的EMS克隆筛选中鉴定的，该克隆筛选鉴定了GATA中的突变，我们鉴定的该基因符合NIH R15授权的GM074735-01的特定目的。与GATA一样，卵巢生殖细胞中的油炸突变克隆会导致哺育细胞和卵母细胞中的染色体形态缺陷，随后卵室停止和降解(Morris，Navarro等人)。2003年)。此外，两个突变体都显示出生长缺陷和发育停滞，尽管油炸幼虫在大小上比GATA更可变，并且油炸突变体阻止羽化后的发育，而GATA突变体阻止幼虫(Morris，Bergman等人)。2008)，对FORED的研究具有非常重要的意义，因为了解FORED将可能有助于控制生长和细胞周期调节。这笔拨款中提出的计划具有额外的重要意义，因为它们将极大地加强林肯中心福特汉姆学院的研究计划，并将使6-10名本科生能够接受遗传学和细胞生物学研究方面的培训。没有其他实验室正在研究油炸，所以这里提出的工作不太可能在其他地方进行。具体目标1：描述油炸EMS等位基因的生长和停滞表型，并描述细胞周期缺陷。具体目的2：通过候选基因测序和转基因拯救，鉴定突变的油炸等位基因，对这些基因进行分子和遗传学特征分析，并通过RT-PCR和原位杂交确定油炸等位基因的表达模式。具体目的3：通过Western blotting和免疫组织化学方法分析FUND在FRED突变体中表达和激活生长调节因子的功能，并通过遗传分析来确定哪些途径与FRED突变相互作用。 公共卫生相关性： 项目叙述缺乏油炸基因功能的果蝇生长不良，发育受阻。适当的生长调节是发育最基本的方面之一，果蝇的许多生长调节因子在人类身上也具有重要的功能。我们建议鉴定FRED突变体中的哪个基因被破坏，并确定FRED基因在野生型果蝇中的功能。

DESCRIPTION (provided by applicant): PROJECT ABSTRACT My laboratory's long-term goal is to improve our understanding of growth, metabolism, and cell cycle regulation during Drosophila development. This proposal focuses on the molecular identification and functional characterization of fried. The three fried mutants were identified in the same EMS clonal screen that identified mutants in gatA, the gene that we characterized in meeting the specific aims of NIH R15 grant GM074735-01. As with gatA, mutant clones of fried in the germ cells of the ovary result in chromosome morphology defects in the nurse cells and oocyte followed by egg chamber arrest and degradation (Morris, Navarro et al. 2003). In addition, both mutants show growth defects and developmental arrest, though the fried larvae are more variable in size than gatA, and fried mutants arrest development post-pupariation while gatA mutants arrest as larvae (Morris, Bergman et al. 2008) Study of fried is highly significant because understanding fried will likely shed light on control of growth and cell cycle regulation. The initiatives proposed in this grant are additionally significant because they will greatly enhance the research program at Fordham College at Lincoln Center and will enable the training of 6-10 undergraduates in genetics and cell biology research. No other lab is studying fried, so the work proposed here is very unlikely to be performed elsewhere. Specific Aim 1: To characterize the growth and arrest phenotypes and to describe cell cycle defects in the fried EMS alleles. Specific Aim 2: To identify the gene defined by the mutant fried alleles by candidate gene sequencing and transgenic rescue, to molecularly and genetically characterize these alleles, and to determine the fried expression pattern by RTPCR and in situ hybridization. Specific Aim 3: To analyze the function of fried by using Western Blotting and Immunohistochemistry to expression and activation of growth regulators in fried mutants and by using genetic assays to determine which pathways interact with fried mutations. PUBLIC HEALTH RELEVANCE: PROJECT NARRATIVE Drosophila lacking fried gene function grow poorly and arrest development. Appropriate regulation of growth is one of the most fundamental aspects of development, and many regulators of growth in Drosophila also have essential functions in humans. We propose to identify which gene is disrupted in fried mutants and to determine the function of fried in wild type Drosophila.