Maternal control of the Drosophila embryonic Dorsal-Ventral axis

描述（由申请人提供）：果蝇胚胎中的背腹极性是在卵子发生过程中通过腹侧卵泡细胞中 Pipe 磺基转移酶的表达而启动的，这通过未知的机制导致胚胎膜和蛋壳之间的卵周间隙中丝氨酸蛋白酶级联的局部激活。我们的长期目标是了解这种丝氨酸蛋白酶级联是如何进行空间调控的。本申请的目的是阐明丝氨酸蛋白酶级联的空间限制的实现机制，并确定毛囊细胞层中 Pipe 作用的目标。我们的中心假设是，腹侧卵泡细胞中的管道活动导致糖蛋白或糖脂的硫酸化，该糖蛋白或糖脂被分泌到腹侧卵周间隙并定位在腹侧卵周间隙内，从而引起丝氨酸蛋白酶级联的局部激活。拟议研究的基本原理是，它将极大地扩展我们对 DV 模式形成的理解，并且还将提供对调节影响人类健康的类似局部丝氨酸蛋白水解事件（例如参与血栓形成和分解的事件）的机制的见解。血液凝固和纤维蛋白溶解是血栓性疾病（例如心脏病和中风）病因学中的关键因素，这些疾病是全世界导致死亡的主要原因。因此，拟议的研究与美国国立卫生研究院的使命相关，即获取可能减轻疾病负担的基础知识。我们将追求两个具体目标：1）确定丝氨酸蛋白酶级联的活性在多大程度上受蛋白酶本身的定位、其空间限制的激活或活性、或通过调节其活性的彼此物理相互作用的调节。蛋白酶的功能性标记版本将用于检查其加工和空间分布，并研究野生型和各种遗传背景中的蛋白质-蛋白质相互作用。 2)鉴定参与腹侧滤泡细胞中Pipe磺基转移酶活性靶标合成的基因。将进行基因筛选，以确定导致突变雌性产生背侧化胚胎的突变。拟议的研究意义重大，因为它将对调节丝氨酸蛋白酶作用的机制产生新的见解，从而可能有助于开发改进的治疗剂来调节影响人类健康的丝氨酸蛋白酶。

DESCRIPTION (provided by applicant): Dorsal-ventral polarity in the Drosophila embryo is initiated during oogenesis by expression of the Pipe sulfotransferase in the ventral follicle cells which, by an unknown mechanism, leads to the localized activation of a serine protease cascade in the perivitelline space between the embryonic membrane and the eggshell. Our long-term goal is to understand how this serine protease cascade is spatially regulated. The objectives of this application are to elucidate the mechanism through which spatial restriction of the serine protease cascade is implemented and to identify the target of Pipe action in the follicle cell layer. Our central hypothesis is that Pipe activity in the ventral follicle cells results in the sulfation of a glycoprotein or glycolipid that is secreted into and localized within the ventral perivitelline space where it brings about the localized activation of the serine protease cascade. The rationale for the proposed research is that it will greatly expand our understanding of DV pattern formation and will also provide insight into mechanisms that regulate analogous localized serine proteolytic events that influence human health such as the ones involved in the formation and breakdown of blood clots. Blood coagulation and fibrinolysis are critical factors in the etiology of thrombotic diseases such as heart attack and stroke which represent leading causes of death worldwide. Thus, the proposed research is relevant to the mission of the NIH to obtain fundamental knowledge that will potentially reduce the burden of disease. We will pursue two Specific Aims: 1) To determine the extent to which the activity of the serine protease cascade is regulated by the localization of the proteases themselves, by their spatially restricted activation or activities, or by physical interactions with one another that modulate their activities. Functional, tagged versions of the proteases will be used to examine their processing and spatial distribution and to investigate protein-protein interactions in wildtype and various genetic backgrounds. 2) To identify genes involved in the synthesis of the target of Pipe sulfotransferase activity in the ventral follicle cells. Genetic screens will be carried out to identify mutations that lead to the production of dorsalized embryos by mutant females. The proposed research is significant as it will yield new insights into mechanisms that regulate serine protease action and thus may contribute to the development of improved therapeutic agents for the modulation of serine proteases that influence human health.