E63-1 AND STEROID/CALCIUM-REGULATED CROSSTALK

类固醇激素是一种小的调节分子，它深刻地 影响发育中生长和动态平衡的基本过程 有机体。据推测，这发生在荷尔蒙诱导 基因表达模式的质变。因此， 识别和表征类固醇作用的基因靶点， 以及它们是如何转化为目标生物变化的 组织，是这项研究的主要目标。具体地说，这个提案 是这一意外发现的逻辑延伸 派的组织特异性(唾液腺)博士后研究 果蝇黑腹果蝇中的类固醇调节基因。 简要地说，E63-1，一个主要的(早期)目标 果蝇类固醇蜕皮激素，编码高亲和力的钙结合 蛋白。这一发现既新颖又令人兴奋，因为大多数 先前表征的早期目标被显示为编码 转录因子。这一发现提供了第一个联系 类固醇和钙离子调节的信号转导通路 果蝇。因此，人们获得了一个独特的机会来 研究这种可能的“串音”在一种遗传上容易驯服的 在一个已经很好描述的分子的背景下的有机体 层级结构。 正在讨论的总体假设是E63-1的功能是 协调类固醇和钙离子介导的调节通路 唾液腺。我们的目标是在 分子水平，研究E64-1编码一种 信号转导分子，并将该信息应用于其他 系统。具体目标如下： 1)从基因水平上操纵E63-1蛋白和 研究这些对组织生理状态的影响 (分泌活动，基因表达表现为泡芙， 细胞内钙离子变化)。 2)通过启动分子筛查进一步定义机制 针对E63-1的下游受体蛋白。

Steroid hormones are small regulatory molecules that profoundly influence basic processes of growth and homeostasis in developing organisms. Presumably, this occurs when the hormone induces a qualitative change in the pattern of gene expression. Thus, identifying and characterizing the gene targets for steroid action, and how they are translated into biological changes in the target tissue, is a primary goal of this research. Specifically, this proposal is a logical extension of the unexpected discovery made during the PI's post-doctoral studies on tissue-specific (salivary gland) steroid-regulated genes in the fruit fly Drosophila melanogaster. Briefly, it was shown that E63-1, a primary (early) target for the Drosophila steroid ecdysone, encodes a high-affinity Ca2+-binding protein. This discovery was novel and exciting because most of the previously characterized early targets were shown to encode transcription factors. This finding provides the first link between steroid-regulated and Ca2+-regulated signaling pathways in Drosophila. Thus, one is afforded a unique opportunity to investigate this possible "crosstalk" in a genetically tractable organism in the context of an already well characterized molecular hierarchy. The overall hypothesis being addressed is that E63-1 functions to coordinate a steroid- and calcium-mediated regulatory pathway in the salivary gland. The goals are to dissect this pathway at the molecular level, investigate the possibility that E64-1 encodes a signal transduction molecule, and apply this information to other systems. The specific aims are as follows: 1) Genetically manipulate the levels of E63-1 protein and investigate these effects on the physiological state of the tissue (secretory activity, gene expression manifested as puffs, intracellular Ca2+ changes). 2) Further define the mechanism by initiating a molecular screen for downstream acceptor proteins of E63-1.