3 UTR CONTROL OF TRA-2 TRANSLATION IN C ELEGANS

3 线虫中 TRA-2 翻译的 UTR 控制

• 批准号: 2190580
• 负责人: ELIZABETH B. GOODWIN
• 金额: $ 19.73万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-01-01 至 1997-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2190580
• 关键词: Caenorhabditis elegans; RNA; alleles; developmental genetics; gel mobility shift assay; gene expression; gene mutation; genetic regulation; genetic regulatory element; genetic translation; molecular cloning; reporter genes; sex determination

How are some genes that specify cell fates translationally regulated so that cells are determined at the right time and place? We propose to study this fundamental question of developmental gene regulation by analyzing the translational regulation of the C. elegan sex determination gene, tra-2. We have shown that a direct repeat, located in the 3' untranslated region (3'UTR), is necessary for repressing the translation of tra-2 mRNA. This repression is required for the proper specification of sexual fates. We have identified an RNA binding factor that binds to the direct repeat. We have also identified a gene, laf-1, that may be involved in this regulation; mutations in laf-1 disrupt the transitional control of reporter constructs by the 3'UTR. During the next five years, we propose to investigate the mechanism by which the 3'UTR controls translation of tra-2 mRNA during development. First, using reporter constructs, we will delineate the necessary cis- acting sequences required for the 3'UTR regulation. In addition, we will ask if the position or orientation of these sequences is important for control. We will study whether the direct repeat controls translation by regulating the length of the poly(A) tail. Second, using RNA gel retardation analysis and reporter constructs, we will investigate whether the binding factor may be a translational inhibitor by correlating the ability of the factor to bind mutant 3'UTRs with the capability of these 3'UTRs to regulate translation. Third, we will genetically characterize the known laf-1 alleles and determine the laf-1 loss-of-function phenotype. This analysis will elucidate the role of laf-1 in the 3'UTR control, and is necessary for the cloning of the gene. Fourth, we will clone laf-1, which will give further insight into its part in the 3'UTR control, and give us the tools to analyze the 3'UTR regulation, biochemically. Fifth, using genetics, reporter constructs, and TRA-2 antibodies, we will study how the 3'UTR regulation changes during development, and identify genes that may control these changes. The health relatedness of this work derives from its contribution to an understanding of fundamental control of gene activity by translational regulation. This class of post-transcriptional regulation is being found to be common in many metazoans, including man.

某些指定细胞命运翻译调节的基因如何 该细胞在正确的时间和地点确定？我们建议 研究这个通过 分析C. leegan性别确定的翻译调节 Gene，Tra-2。我们已经表明，位于3'的直接重复 未翻译的区域（3'UTR）是压制翻译所必需的 Tra-2 mRNA。适当规格需要这种抑制 性命运。我们已经确定了与结合的RNA结合因子 直接重复。我们还确定了一个基因LaF-1，可能是 参与该法规； LAF-1中的突变破坏了过渡 通过3'UTR控制记者构造。 在接下来的五年中，我们建议通过 3'UTR控制开发过程中Tra-2 mRNA的翻译。 首先，使用记者构造，我们将描述必要的顺式 3'UTR调节所需的作用序列。此外，我们将 询问这些序列的位置或方向对于 控制。 我们将研究直接重复控制翻译是否 通过调节聚（a）尾的长度。第二，使用RNA凝胶 延迟分析和记者结构，我们将调查是否是否 结合因子可以通过将 因子将突变体3'UTR与这些功能结合的能力 3'Utrs调节翻译。第三，我们将在基因上表征 已知的LAF-1等位基因并确定LAF-1功能丧失 表型。该分析将阐明LAF-1在3'UTR中的作用 控制，对于基因的克隆是必需的。第四，我们会的 克隆LAF-1，它将进一步了解其在3'UTR中的一部分 控制，并为我们提供分析3'UTR法规的工具， 生化。第五，使用遗传学，记者构造和TRA-2 抗体，我们将研究3'UTR调节在 开发并确定可能控制这些变化的基因。 这项工作的健康相关性来自于其对 通过翻译对基因活性的基本控制 规定。正在发现这类的转录后法规 在包括人在内的许多后生动物中很常见。