Elucidation of Novel Post-Translational Mechanisms That Regulate Notch Activity

描述(由申请人提供)：Notch信号的错误调节与许多生物体和人类疾病的发育缺陷有关，最明显的是T细胞急性淋巴细胞白血病(T-ALL)。这项研究将利用秀丽隐杆线虫来表征Notch的激活形式在发育背景下的调节。具体地说，这项研究将考察两种潜在的新形式的负面监管，它们在机制层面上没有被理解。这两种形式的调控都阻断了线虫LIN-12/Notch在细胞核中的功能。在一种情况下，与RAS-MAP激酶通路的串扰阻断了结构性激活的LIN-12/Notch的作用，特别是在RAS-MAP激酶被激活的地方。在第二种情况下，新的DNA结合蛋白，LIN-14，调节发育时间，在特定组织中阻止构成活性的LIN-12/Notch。被阻断的LIN-12的构成活性形式类似于在某些癌症中发现的突变，如T-ALL。在这项研究的过程中，我将表征两个必需的Notch辅助蛋白的调节，一个保守的转录因子(LAG-1)和一个共激活因子(SEL-8)，以及它们与Lin-12形成的转录复合体。此外，这些方法将被扩展到通过操纵LIN-12目标记者而发现的其他监管形式的特征。通过表征这两种机制，本提案旨在增加对关键Notch途径如何调节的理解，并有可能为T-ALL等疾病提供新的治疗策略。

DESCRIPTION (provided by applicant): The misregulation of Notch signaling has been implicated in developmental defects in many organisms and in human diseases, most notably T-Cell Acute Lymphoblastic Leukemia (T-ALL). This research will characterize the regulation of activated forms of Notch in developmental contexts using Caenorhabditis elegans. Specifically, this research will examine two potentially novel forms of negative regulation that are not understood at the mechanistic level. Both forms of regulation block C. elegans LIN-12/Notch function in the nucleus. In one case, crosstalk with the Ras-MAP kinase pathway blocks the effect of constitutively active LIN-12/Notch specifically where Ras-MAP kinase is activated. In the second case, the novel DNA-binding protein, LIN-14, which regulates developmental timing, blocks constitutively active LIN-12/Notch in a particular tissue. The constitutively active forms o LIN-12 blocked is analogous to mutations found in certain cancers such as T-ALL. In the course of this research, I will characterize the regulation of two required Notch accessory proteins, a well-conserved transcription factor (LAG-1) and a coactivator (SEL-8), and the transcriptional complex they form with LIN-12. Additionally, these methods will be expanded to characterize additional forms of regulation found through manipulation of LIN-12 target reporters. By characterizing these two mechanisms, this proposal aims to increase the understanding of how the critical Notch pathway is regulated and potentially provide novel therapeutic strategies for diseases such as T-ALL.