Shcc Function In Growth, Development And Cancer

工作总结：我们的小组有兴趣了解ShcC衔接蛋白在受体酪氨酸激酶（RTK）功能中的作用，特别是神经发育。ShcC是衔接蛋白Shc家族的成员。该家族由三个成员（ShcA、ShcB和ShcC）组成，每个成员的特征在于存在氨基末端磷酸酪氨酸结合结构域（PTB）、中央富含Gly和Pro的效应子区（CH1）和羧基末端Src同源2（SH2）结构域。尽管该蛋白质家族缺乏任何内在酶活性，但Shc蛋白充当组装信号传导复合物的支架，所述信号传导复合物调节RTK以及多种受体蛋白的功能，所述受体蛋白包括整联蛋白、G蛋白偶联受体（GPCR）和造血受体。此外，其他实验室最近的工作表明，ShcA的某些亚型可能在细胞对氧化应激的反应和生物体的寿命中发挥重要作用。在与Tony Pawson博士的合作中，我们已经证明，消除ShcC不会影响神经系统的发育，这表明ShcC可能对功能而不是发育更重要。使用技术的组合，我们已经表明，ShcC表达仅限于神经系统，特别是神经元。为了解决ShcC在神经系统中的重要性，我们已经开始更精确地定义ShcC在发育过程中的时间和空间表达模式。除了确定ShcC的表达模式外，我们还开始了确定整个动物中ShcC磷酸化状态的实验。我们已经开发了针对活化形式的ShcC的磷酸特异性抗体。使用这些抗体，我们希望不仅能确定ShcC在何处和何时表达，还能确定它在何处和何时被激活。此外，我们正在利用质谱法，以确定在体内磷酸化的具体网站。这些方法也将证明用于确定环境损伤是否影响ShcC功能。例如，神经毒物是否影响ShcC表达或酪氨酸磷酸化？为了进一步探索ShcC的重要性，我们正在使用该蛋白的突变体来干扰RTK信号传导。尽管ShcC具有两个与活化的RTK相互作用的结构域，但我们已经证明PTB结构域是ShcC结合活化的EGFR的主要方式。此外，我们的研究结果表明，SH2结构域可能是重要的结合到一个额外的细胞成分（S）必需的EGFR功能。我们目前正试图分离SH2相互作用的蛋白，希望能确定ShcC和EGFR的重要下游靶点。

SUMMARY OF WORK: Our group is interested in understanding the role of the ShcC adaptor protein in receptor tyrosine kinase (RTK) function in general and neural development in particular. ShcC is a member of the Shc family of adaptor proteins. This family consists of three members (ShcA, ShcB and ShcC), each of which is characterized by the presence of an amino-terminal phosphotyrosine binding domain (PTB), a central Gly- and Pro-rich effector region (CH1), and a carboxy-terminal Src homology 2 (SH2) domain. Although this family of proteins lacks any intrinsic enzymatic activity, Shc proteins act as scaffolds to assemble signaling complexes that regulate the function of RTKs as well as a wide variety of receptor proteins including integrins, G-protein coupled receptors (GPCRs) and hematopoietic receptors. In addition, recent work from other laboratories suggests that certain isoforms of ShcA may play an important role in the response of cells to oxidative stress and in the life span of an organism. In collaboration with Dr. Tony Pawson, we have shown that elimination of ShcC does not affect development of the nervous system suggesting that ShcC may be more important for function rather than development. Using a combination of techniques, we have shown that ShcC expression is restricted to the nervous system and to neurons in particular. To address the importance of ShcC in the nervous system, we have begun to more precisely define both the temporal and spatial expression pattern of ShcC during development. In addition to determining the expression pattern of ShcC, we have begun experiments to determine the phosphorylation status of ShcC in the whole animal. We have developed phosphospecific antibodies directed against the activated forms of ShcC. Using these antibodies we hope to determine not only where and when ShcC is expressed but also where and when it is activated. In addition, we are utilizing mass spectrometry in order to determine the specific sites of phosphorylation in vivo. These approaches will also prove useful for determining whether environmental insult affects ShcC function. For example, do neurotoxicants affect ShcC expression or tyrosine phosphorylation? To further explore the importance of ShcC, we are using mutant versions of the protein to interfere with RTK signaling. Although ShcC possesses two domains with the potential of interacting with activated RTKs, we have demonstrated that the PTB domain is the predominant means by which ShcC binds to the activated EGFR. Furthermore, our results suggested that the SH2 domain might be important for binding to an additional cellular component(s) necessary for EGFR function. We are currently attempting to isolate SH2 interacting proteins with the hope of identifying important downstream targets of both ShcC and the EGFR.