Activation and Signaling Mechanism of the RET Tyrosine Kinase Receptor

描述(由申请人提供)：该奖学金将支持波士顿大学禤浩焯·惠蒂教授实验室研究生的研究目标和培训。许多形式的癌症是由生长因子(GF)受体信号异常引起的。因此，更好地了解GF受体的激活和功能将继续促进癌症治疗方法的改进。然而，关于GF受体激活和信号转导的分子机制的许多基本问题仍未得到解答。我们建议开发新的方法来定量研究GF受体信号的机制，以多组分的RET受体为测试案例。RET是一种受体酪氨酸激酶，是神经营养因子神经营养因子(GDNF)家族的受体。RET信号的失控与多种癌症有关，包括多发性内分泌肿瘤(MEN)2A和-2B、甲状腺髓样癌、甲状旁腺增生和嗜铬细胞瘤。我们建议通过以下三个目标来研究RET激活和信号转导的分子机制：1)利用基因编码的荧光共振能量转移(FRET)构建一个报告系统，用于利用流式细胞术监测活细胞表面的RET结合事件。2)在目标1中开发的FRET报告结构将用于解决有关活细胞上RET激活机制的关键问题。这些问题包括解决长期存在的困难和不确定性，即GF受体是通过配体诱导的受体二聚化发挥作用，还是通过引起前相关受体二聚体或寡聚体的结构重排发挥作用。3)探讨RET与血管内皮细胞生长因子受体2(VEGFR2)信号串扰的机制和影响，以确定RET信号串扰发生在什么水平，以及对RET功能的影响如何依赖于VEGFR2刺激水平。总而言之，这三个目标将建立新的方法来阐明GF受体的激活机制，并将建立两个不同的GF受体家族之间的串扰机制和影响。这项研究将帮助这位研究员实现她的职业目标，即领导一个学术研究小组，解决与人类健康有关的重要生物学问题。她计划通过在分子、细胞和活体水平的系统复杂性方面的工作来解决这些问题，主要采用假设驱动和(在适当的情况下)定量的观点。她研究生工作中的培训计划将使她能够专注于发展她在分子和细胞方法方面的知识和能力，她的想法是，她将继续通过在合适的实验室进行博士后研究来涵盖体内技术和观点。通过共同努力，候选人和她的赞助商制定了一个非常有效的计划，以实现她将需要的培训，以实现她的长期职业目标。

DESCRIPTION (provided by applicant): This fellowship will support the research aims and training of a graduate student in the laboratory of Professor Adrian Whitty at Boston University. Many forms of cancer are caused by aberrant signaling of growth factor (GF) receptors. Consequently, achieving an improved understanding of GF receptor activation and function continues to give rise to improved approaches to cancer treatment. However, many basic questions about the molecular mechanisms of GF receptor activation and signaling remain unanswered. We propose to develop new methods for the quantitative mechanistic study of GF receptor signaling, using the multicomponent RE-arranged during Transfection (RET) receptor as a test case. RET is a receptor tyrosine kinase that is the receptor for the glial cell-line deried neurotrophic factor (GDNF) family of neurotrophins. Deregulation of RET signaling is causally involved in several types of cancer, including multiple endocrine neoplasias (MEN) 2A and -2B, medullary thyroid carcinoma, parathyroid hyperplasia, and phaeochromocytoma. We propose to characterize the molecular mechanisms of RET activation and signaling by pursuing following three aims: 1) genetically encoded fluorescence resonance energy transfer (FRET) constructs will be used to develop a reporter system for monitoring RET association events on the surface of live cells using flow cytometry. 2) The FRET reporter constructs developed in Aim 1, will be used to address key questions concerning the mechanism of RET activation on live cells. These questions include addressing the difficult and longstanding uncertainty concerning whether GF receptors function by ligand-induced receptor dimerization, or alternatively by causing a structural rearrangement within pre-associated receptor dimers or oligomers. 3) The mechanisms and impact of crosstalk between RET and vascular endothelial growth factor receptor 2 (VEGFR2) will be explored, to establish at what levels in RET signaling crosstalk occurs and how the effects on RET function depend on the level of VEGFR2 stimulation. Collectively, these three aims will establish new methods to elucidate mechanisms of GF receptor activation, and will establish the mechanisms and impact of crosstalk between two different families of GF receptors. This research will help the fellow achieve her career goal, which is to lead an academic research group that addresses important biological problems relating to human health. She plans to address these problems by working at the molecular, cellular and in vivo levels of systems complexity, employing primarily a hypothesis-driven and (where appropriate) a quantitative perspective. The training plan in her graduate work will enable her to focus on developing her knowledge and capabilities of molecular and cellular approaches, with the idea that she will go on to encompass in vivo techniques and perspectives through post doctoral study in a suitable laboratory. Working together, the candidate and her sponsor have developed a highly effective plan for how to achieve the training she will need to meet her long-term career objectives.