Role of Misshappen (NIK/Msn) kinases and Extended-Synaptotagmins (E-Syts) in Wnt and FGF intracellular signaling pathways.

细胞间的信号传导是决定后生动物身体计划和控制生长和分化的基础。因此，大量的信号分子家族及其同源受体已经进化到能够实现早期发育、器官发生和体内平衡的复杂性。其中一些家族在早期发育中很重要，特别是无翼细胞（Wnt）和成纤维细胞生长因子（fgf）家族。它们在发育过程中发挥着独立但相互关联的作用，并通过不同的和共享的转导途径发挥作用。我们已经确定了FGF和Wnt信号通路的新元件，为信号传导和通路间串扰的重要方面提供了潜在的窗口。我们提出的机制研究将为细胞内信号传导的复杂网络提供新的见解。******我们首先确定了扩展突触塔蛋白2 （ESyt2）在FGF信号传导中的作用。ESyts是一种多c2结构域磷脂结合膜蛋白，作为FGF受体（fgfr）的内吞适配器。ESyt2通过介导FGFR内吞作用调节map -激酶途径，并可能在内质网-质膜（ER-PM）连接、储存操作钙离子(SOCE)/Ca2+信号传导和磷脂转运到PM中发挥相关作用。为了了解它们的生理作用，我们产生了esyt1 /2/3缺失的小鼠，建立了ESyt-null细胞系，并证明它们在Ca2+信号传导中存在缺陷，并开始了果蝇独特的ESyt同源基因的遗传研究。我们建议使用这些工具来确定ESyts的体外和体内功能，以及它们如何影响磷脂代谢，影响FGF和潜在的Wnt信号转导。******Wnt信号已被确定为发育模式的关键参与者。然而，尽管付出了相当大的科学努力，我们对规范β -连环蛋白、非规范平面细胞极性和Ca2+信号通路之间的串扰的理解仍然是初级的。我们最近发现，Xenopus Misshapen （Msn）的同源物TNIK和MINK信号激酶是典型和非典型Wnt通路的组成部分。我们的数据强烈表明，典型和非典型途径之间的平衡是由TNIK和MINK的蛋白水解控制的，这决定了它们的激酶活性和亚细胞位置。我们已经证明这种切割是保守的，因为独特的果蝇Msn在发育过程中也被切割。我们提出了一系列的遗传学、细胞和分子研究来确定Msn切割的发育和生理意义，从而更好地理解Wnt信号传导的复杂控制。

Signalling between cells is fundamental to determining the metazoan body plan and in controlling growth and differentiation. Hence, a large number of families of signalling molecules and their cognate receptors have evolved to enable the complexities of early development, organogensis and homeostasis. Several of these families are of importance during early development, in particular the Wingless (Wnt) and Fibroblast Growth Factors (FGFs) families. These play separate, but interrelated, roles during development and act through distinct as well as shared transduction pathways. We have identified novel elements of the FGF and Wnt signalling pathways that provide potential windows into important aspects of signalling and inter-pathway crosstalk. We propose mechanistic studies that will provide new insight into the complex network of intracellular signalling.******We were first to identify a role for the Extended Synaptotagmin 2 (ESyt2) in FGF signalling. The ESyts are a multi-C2-domain phospholipid-binding membrane protein, that function as endocytic adapters for the FGF Receptors (FGFRs). ESyt2 regulates the MAP-kinase pathway by mediating FGFR endocytosis, and plays probably related roles in endoplasmic reticulum - plasma membrane (ER-PM) junctioning, Store Operated Calcium Entry (SOCE)/Ca2+-signaling, and transport of phospholipids to the PM. To understand their physiological roles, we have generated ESyt1/2/3-null mice, established ESyt-null cell lines and shown them to have defects in Ca2+ signalling, and have begun genetic studies of the unique ESyt ortholog in Drosophila. We propose to use these tools to determine the in vitro and in vivo functions of the ESyts and how they impinge on phospholipid metabolism, and affect FGF and potentially Wnt signal transduction.******Wnt signalling has been established as a key player in developmental patterning. However, despite considerable scientific effort, our understanding of crosstalk between the canonical beta-catenin, non-canonical Planar Cell Polarity and Ca2+ signalling pathways remains rudimentary. We recently discovered that the Xenopus Misshapen (Msn) homologs TNIK and MINK signalling kinases form integral components of both the canonical and non-canonical Wnt pathways. Our data strongly suggest the balance between the canonical and non-canonical pathways is controlled by proteolysis of TNIK and MINK, which determines both their kinase activities and subcellular locations. We have shown that this cleavage is conserved, since the unique Drosophila Msn is also cleaved during development. We propose a series of genetic, cell and molecular studies to determine the developmental and physiological significance of Msn cleavage, and hence to better understand the complex control of Wnt signalling.