Maternal control of the Drosophila embryonic Dorsal-Ventral axis

描述（由申请人提供）：果蝇胚胎的背腹极性在卵子发生期间通过腹侧卵泡细胞中Pipe磺基转移酶的表达而启动，其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空间调节的。本申请的目的是阐明丝氨酸蛋白酶级联的空间限制的机制，并确定在卵泡细胞层中的Pipe作用的目标。我们的中心假设是，在腹侧卵泡细胞中的Pipe活动导致糖蛋白或糖脂的硫酸化，糖蛋白或糖脂分泌到腹侧卵黄周间隙内并定位在腹侧卵黄周间隙内，在那里它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，它将大大扩展我们对DV模式形成的理解，并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的机制的见解，例如参与血栓形成和分解的机制。血液凝固和纤维蛋白溶解是血栓性疾病如心脏病发作和中风的病因学中的关键因素，这些疾病代表了世界范围内的主要死亡原因。因此，拟议的研究与NIH的使命相关，即获得可能减少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、受其空间限制的活化或活性、或受调节其活性的彼此之间的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它们的加工和空间分布，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)确定腹侧卵泡细胞中参与Pipe磺基转移酶活性靶标合成的基因。将进行遗传筛选，以确定导致突变雌性产生背侧胚胎的突变。这项研究意义重大，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用于调节影响人类健康的丝氨酸蛋白酶的改进治疗剂。

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.