THE METAMORPHOSIS OF CHROMATIN

蛾类的变态包括细胞群体的连续变化 他们的合成活动，以建立新类型的角质层下的直接 两种激素的影响，蜕皮类固醇和juvenoids。 的变化 我们在体内和体外操纵它们的能力， 分析寄生虫发育事件的有用模型， 我们自己. 我们建议了解如何变态调节在nomic 水平 为此，我们将分离和表征基因和转录本 对于四种表皮蛋白，选择它们的挑衅性空间和 时间分布 我们计划通过分析染色质来确定 包括这些基因的调控域。 推定顺式作用 调节区域将通过使用DNase I定位超敏反应， 位点并使用硫酸二甲酯（DMS）进行“体内足迹”。 通过这些技术鉴定的推定调控区将用于 凝胶阻滞测定法，其中调节蛋白的存在可以 因为它们降低了DNA的流动性， 具体绑定。 我们将测试不同的核提取物 解剖区域和变质阶段以及已知的反式作用 调节因素，以整理的性质（空间，时间，激素） 影响顺式作用调控序列的信息， 单基因在变态过程中。我们希望通过这些分析 了解激素是否通过直接作用于基因来控制变态 编码结构蛋白的基因 蛾的表皮呈现一种 这是一个非常有利的模型，用于分析 染色质结构随着细胞群的连续运动而改变， 制定一项发展计划， 确定和活性的个别基因。

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.