METAMORPHOSIS OF CHROMATIN

染色质变态

• 批准号: 3303648
• 负责人: JUDITH WILLIS
• 金额: $ 12.02万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3303648
• 关键词: DNA binding protein; DNA footprinting; Lepidoptera; binding proteins; chromatin; complementary DNA; deoxyribonuclease I; developmental genetics; endonuclease; gel mobility shift assay; gene expression; genetic manipulation; genetic regulation; genetic regulatory element; genome; homeobox genes; hormone receptor; hormone regulation /control mechanism; invertebrate cuticle; laboratory mouse; metamorphosis; molecular genetics; nucleic acid hybridization; nucleic acid probes; nucleic acid sequence; polymerase chain reaction; protein biosynthesis; protein sequence

Metamorphosis in moths involves a continuous population of cells changing their synthetic activities to build new types of cuticle under the direct influence of two hormones, ecdysteroids and juvenoids. The changes that occur and our ability to manipulate them in vivo and in vitro presents a useful model for analyzing developmental events in our parasites and ourselves. We propose to learn how metamorphosis is regulated at the nomic level. To this end we shall isolate and characterize genes and transcripts for four cuticular proteins selected for their provocative spatial and temporal distributions. We plan to analyze chromatin to identify the regulatory domains that encompass these genes. Putative cis-acting regulatory regions will be located by using DNase I to map hypersensitive sites and by using dimethylsulfate (DMS) for "in vivo footprinting". Putative regulatory regions identified by these techniques will be used in gel retardation assays where the presence of regulatory proteins can be recognized since they reduce the mobility of DNA to which they are specifically bound. We shall test nuclear extracts from different anatomical regions and metamorphic stages as well as known trans-acting regulatory factors to sort out the nature (spatial, temporal, hormonal) of the information impinging on the cis-acting regulatory sequences of a single gene during the course of metamorphosis. We hope by these analyses to learn if hormones control metamorphosis by acting directly ont he genes that code for structural proteins. The moth epidermis presents an extremely favorable model for analyzing the fundamental issue of how chromatin structure is altered as a continuous population of cells plays out a development program that involves changes in the state of determination and activity for individual genes.

蛾子的变态包括不断变化的细胞数量 他们的合成活动在指导下构建新型角质层 两种激素的影响，蜕皮激素和幼年激素。这些变化 我们在体内和体外操纵它们的能力呈现出一种 用于分析寄生虫发育事件的有用模型和 我们自己。我们建议了解变态是如何在生物体中被调节的。 水平。为此，我们将分离和鉴定基因和转录本 四种角质层蛋白被选为具有挑衅性的空间和 时间分布。我们计划分析染色质以确定 包含这些基因的调节域。推定的顺势作用 将通过使用DNase I来定位调控区域以映射超敏 并使用二甲基硫酸盐(DMS)进行“体内足迹”。 这些技术确定的假定监管区域将用于 凝胶滞留分析是指调节蛋白的存在 因为它们降低了它们所针对的DNA的流动性 明确捆绑在一起。我们将测试不同来源的核提取液 解剖区域和变质阶段以及已知的反式作用 理清自然(空间、时间、荷尔蒙)的调控因素 影响顺式作用的调控序列的信息 变态过程中的单基因。我们希望通过这些分析 了解荷尔蒙是否通过直接作用于基因来控制变态 这是结构蛋白的密码。蛾的表皮呈现出一种 非常有利的模型，用于分析如何 随着细胞数量的不断增加，染色质结构也会发生变化 制定一项涉及国家状况变化的发展计划 单个基因的测定和活性。