Role of Clic in Epithelial Morphogenesis

描述（由申请人提供）：本申请的长期目标是阐明最近描述的称为Clics（氯细胞内通道）的蛋白质家族在上皮形态发生中的作用。虽然首字母缩略词“Clic”意味着这些蛋白质可能起离子通道的作用，但新出现的证据表明，它们也具有不同于离子转运的替代功能。待检验的中心假设是Clics是在上皮细胞形态发生期间控制细胞骨架活性和极性的调节蛋白。通过遗传学、生物化学和亚细胞定位研究，已确定Clics与肌动蛋白细胞骨架和信号转导相关;然而，它们的确切作用仍有待确定。最近的研究表明，Clic基因的破坏会导致C.小鼠感觉上皮细胞结构变性引起的线虫和耳聋。此外，人Clics已被认为是肿瘤形成和转移的诊断指标和致病因素。Clics的功能尚未经过严格测试。在这个提议中，果蝇将被用作一个模型系统，以解开Clics的功能意义和细胞作用。与有多个Clic基因和剪接变体的脊椎动物不同，果蝇只有一个Clic基因。我们的初步数据表明，果蝇Clic功能相互作用的细胞骨架蛋白和信号分子必不可少的苍蝇发展。主要目的是研究Clic在胚胎上皮细胞形态发生过程中的细胞骨架调节作用，并拓宽我们对Clic在胚胎发育过程中信号传导作用的观点，具体如下：1.研究Clic在胚胎上皮形态发生中的作用。我们将测试是否果蝇Clic是建立在早期胚胎发育的分割决定因素的位置所必需的。此外，我们将研究的作用，CLIC在上皮内陷和上皮片运动在胚胎发育后期。CLIC和已知的调节蛋白质之间的潜在相互作用的关键上皮极性将进行测试遗传和生化。2.确定在胚胎发育过程中与Clic相互作用的基因。我们将确定遗传修饰的CLIC，验证的相互作用，并评估细胞功能的CLIC修饰剂在胚胎发育过程中。此外，我们将测试候选人的调控基因的功能重要的相互作用与CLIC在形态发生运动的上皮片。由于许多蛋白质在脊椎动物和无脊椎动物物种中是功能保守的，因此很可能对果蝇中的Clic功能进行分析将有助于更好地了解人类中的Clic功能。 公共卫生部门：该提案的主要目标是确定在人类和昆虫之间高度保守的Clic蛋白如何控制细胞形状和运动。Clic蛋白与人类健康相关，因为它们对听力和平衡至关重要，并且与肿瘤形成和转移有关。果蝇将被用作模型系统，以了解Clic如何与其他基因和蛋白质一起工作，这些基因和蛋白质对胚胎发育期间的细胞形状和运动至关重要。

DESCRIPTION (provided by applicant): The long-term goal of this application is to elucidate the role of a recently described family of proteins termed Clics (chloride intracellular channels) in epithelial morphogenesis. Although the acronym "Clic" implies that these proteins may function as ion channels, emerging evidence indicates that they also have alternative functions distinct from ion transport. The central hypothesis to be tested is that Clics are regulatory proteins that control cytoskeletal activity and polarity during epithelial cell morphogenesis. Clics have been identified in association with the actin cytoskeleton and signal transduction through genetic, biochemical, and subcellular localization studies; however, their precise roles remain to be defined. Recent studies have shown that disruption of Clic genes causes tubular cyst formation in C. elegans and deafness due to structural degeneration of sensory epithelial cells in mice. In addition, human Clics have been implicated as diagnostic indicators and causative factors in tumor formation and metastasis. The function of Clics has not been rigorously tested. In this proposal, Drosophila will be used as a model system to unravel the functional significance and cellular roles of Clics. Unlike vertebrates, which have multiple Clic genes and splice variants, Drosophila has only one Clic gene. Our preliminary data indicate that Drosophila Clic functionally interacts with cytoskeletal proteins and signaling molecules essential to fly development. The major objectives are to examine the role of Clic in cytoskeletal regulation during embryonic epithelial morphogenesis and to broaden our perspective on the role of Clic in signaling during embryonic development as follows: 1. Investigate the role of Clic during embryonic epithelial morphogenesis. We will test whether Drosophila Clic is required for establishing the position of segmentation determinants in early embryogenesis. In addition, we will investigate the role of Clic in epithelial invagination and epithelial sheet movement late in embryogenesis. Potential interactions between Clic and known regulatory proteins key to epithelial polarity will be tested genetically and biochemically. 2. Identify genes which interact with Clic during embryonic development. We will identify genetic modifiers of Clic, verify the interactions, and assess the cellular functions of Clic modifiers during embryonic development. In addition, we will test candidate regulatory genes for functionally important interactions with Clic during morphogenetic movements of epithelial sheets. Since many proteins are functionally conserved among vertebrate and invertebrate species, it is likely that analysis of Clic function in Drosophila will lead to a better understanding of Clic function in humans. PUBLIC HEALTH REVELANCE: The main goal of this proposal is to determine how Clic proteins, which are highly conserved between humans and insects, control cell shape and movement. Clic proteins are relevant to human health because they are essential for hearing and balance and have been implicated in tumor formation and metastasis. The fruit fly will be used as a model system to understand how Clic works with other genes and proteins known to be important for cell shape and movement during embryonic development.