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

细胞之间的信号传导对于决定后生动物的身体计划以及控制生长和分化是至关重要的。因此，大量家族的信号分子和它们的同源受体已经进化，使早期发育，器官发生和稳态的复杂性。这些家族中的几个在早期发育期间是重要的，特别是无翼（Wnt）和成纤维细胞生长因子（FGF）家族。它们在发育过程中发挥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，这些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。我们提出了机制研究，这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结合蛋白2（ESyt 2）在FGF信号传导中的作用。ESyt是一种多C2结构域磷脂结合膜蛋白，其作为FGF受体（FGFR）的内吞衔接子发挥作用。ESyt 2通过介导FGFR内吞作用调节MAP-激酶途径，并且在内质网-质膜（ER-PM）连接、钙库操纵的钙进入（SOCE）/Ca 2 +-信号传导和磷脂向PM的转运中可能发挥相关作用。为了了解它们的生理作用，我们已经产生了ESyt 1/2/3-null小鼠，建立了ESyt-null细胞系，并显示它们在Ca 2+信号传导中存在缺陷，并已开始对果蝇中独特的ESyt直系同源物进行遗传研究。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，并影响FGF和潜在的Wnt信号转导。Wnt信号转导已被确立为发育模式的关键参与者。然而，尽管有相当多的科学努力，我们的理解之间的串扰的经典β-连环蛋白，非经典平面细胞极性和钙离子信号通路仍然是基本的。我们最近发现，畸形非洲爪蟾（MSN）同系物TNIK和MINK信号激酶形成的经典和非经典Wnt途径的组成部分。我们的数据强烈表明，经典和非经典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。我们已经表明，这种切割是保守的，因为独特的果蝇Msn也在发育过程中被切割。我们提出了一系列的遗传，细胞和分子研究，以确定的发展和生理意义的Msn切割，从而更好地了解Wnt信号的复杂控制。

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.