SNTS--DEVELOPMENTAL FUNCTIONS AND RECEPTOR INTERACTIONS

描述（改编自申请人摘要）：SNT （Suc1-binding Neurotrophic factor-induced tyrosine -phosphorylated target）蛋白是最近发现的衔接分子，在神经营养因子（NGF、BDNF、NT3）和fgf等生长因子刺激下，但不受胰岛素和EGF的刺激，它们会经历快速的酪氨酸磷酸化。它们与IRS有远亲关系。snt是肉豆酰化和膜锚定的，并具有n端磷酸酪氨酸结合（PTB）结构域，该结构域介导弱但功能显著的snt受体相互作用。snt的配体依赖性酪氨酸磷酸化使snt能够与至少两个不同的SH2结构域片段Grb2/Sos和Shp2相互作用。由于Grb2/Sos和随后的Ras激活以及Shp2磷酸酶的激活已被证明是FGF和神经营养因子诱导的广泛生物学反应所必需的，snt可能是激活两个生物学关键途径所必需的。在这一应用中提出了三个目的来阐明snt的生化和生物学特性。目的1是通过核磁共振和诱变方法的结合来确定SNT PTB结构域之间相互作用的性质。还将测试snt与另一种受体酪氨酸激酶Ret之间可能的直接相互作用。在具体目标2中，将讨论SNT1和SNT2的生物学功能。为此目的将采取两种办法：1.)snt的显性阴性突变体将在培养细胞中过表达。产生SNT基因null、半变形和条件null突变的小鼠。最后，在具体目标3中，将测试snt作为潜在生物特异性决定因素的想法。为此，嵌合的SNT-IRS以及胰岛素受体- trk杂交体将被制造出来，以确定它们是否能将胰岛素反应转化为神经营养因子或fgf的反应。

DESCRIPTION (Adapted from applicant's abstract): SNT (Suc1-binding Neurotrophic factor-induced Tyrosine-phosphorylated target) proteins are recently discovered adaptor molecules that undergo rapid tyrosine phosphorylation following stimulation by growth factors including neurotrophins (NGF, BDNF, NT3) and FGFs but not by insulin and EGF. They are distantly related to IRS. SNTs are myristoylated and membrane-anchored and bear an N-terminal phosphotyrosine-binding (PTB) domain that mediates weak, but functionally significant SNT-receptor interactions. The ligand- dependent tyrosine phosphorylation of SNTs enables SNTs to interact with at least two different SH2 domain-containing moieties, Grb2/Sos and Shp2. As activation of both the Grb2/Sos and the subsequent Ras activation as well as the Shp2 phosphatase has been shown to be required for a wide-range of FGF- and neurotrophin- induced biological responses, SNTs may be required for activation of two biologically critical pathways. Three aims are proposed in this application to elucidate the biochemical and biological properties of SNTs. Aim 1 is proposed to determine the nature of the interaction between SNT PTB domains by using a combination of NMR and mutagenesis approaches. A possible direct interaction between SNTs and another receptor tyrosine kinase Ret will also be tested. In specific aim 2, the biological function of SNT1 and SNT2 will be addressed. Two approaches will be undertaken for this purpose: 1.) Dominant negative mutants of SNTs will be overexpressed in cultured cells and 2.) Mice with null, hypmerorphs and conditional null mutations in SNT genes will be generated. Finally, in specific aim 3, the idea of SNTs serving as potential biological specificity determinants will be tested. To this end, chimeric SNT-IRS as well as insulin receptor-Trk hybrids will be produced to determine if they can turn insulin response to that of neurotrophins or FGFs.