Structure and Function of the SHOC2 Holophosphatase Complex in RAS-driven Cancer

项目摘要/摘要： 在癌症的临床病例中，发现RAS蛋白频繁突变。有效的靶向治疗 针对RAS及其下游通路一直备受追捧。在RAS效应器中， RAF/丝裂原活化蛋白激酶(MAPK)级联信号通路已被证明对繁殖很重要 RAS下游的致癌信号。许多有希望的靶向疗法已经被开发出来 直接抑制突变的KRAS和MAPK成员，但由于抗性机制而受到限制。 最近发现SHOC2在各种RAS突变体中是一个强大的合成致死漏洞 背景，包括RAS驱动的白血病/黑色素瘤，并促进癌症对多种RAS/MAPK的耐药性 抑制剂。SHOC2是一种结合RAS和PP1C形成三元全磷酸酶复合体的支架，该复合体 使抑制RAF蛋白的‘S259’去磷酸化并启动MAPK信号转导。将SHOC2实现为 治疗靶点和提示今后努力合理设计抑制剂，针对复杂、深入 需要了解管理复杂装配和功能的SHOC2结构特征。我们有 现在解决了SHOC2的复杂结构，并概述了几乎所有 通过深层突变扫描，可能在SHOC2的每个位置发生氨基酸替换。 这些发现突出了SHOC2的新结构特征，这些结构特征可能对治疗靶点很重要 在RAS突变的癌症中。本提案中概述的研究将以这项工作为基础，并旨在(1)界定和 验证SHOC2全磷酸酶复合体对RAF底物的特异性结构特征，以及(2) 确定调节SHOC2功能的新的翻译后修饰。我在K99接受的训练 阶段将为我提供结构生物学、蛋白质组学和泛素生物学方面的必要专业知识。 因为我有教学/实验室管理经验，所以我可以有效地领导我自己的独立 研究计划。K99/R00将有助于我实现我的长期目标，建立我的 实验室专注于研究RAS相关的新脆弱性和耐药性机制，通过 细胞信号和蛋白质结构-功能的系统方法。

Project Summary/Abstract: RAS proteins are found to be frequently mutated in clinical cases of cancer. Effective targeted therapies against RAS and its downstream pathways have been highly sought after. Among the RAS effectors, the RAF/mitogen-activated protein kinase (MAPK) cascade has been shown to be important for propagating oncogenic signaling downstream of RAS. Many promising targeted therapies have been developed that directly inhibit mutant KRAS and MAPK members but have been limited due to resistance mechanisms. SHOC2 has been recently found to be a potent synthetic lethal vulnerability in various RAS mutant contexts, including RAS-driven leukemia/melanoma and facilitates cancer resistance to numerous RAS/MAPK inhibitors. SHOC2 is a scaffold that binds RAS and PP1C to form a ternary holophosphatase complex which dephosphorylates inhibitory ‘S259’ on RAF proteins and enables MAPK signaling. To actualize SHOC2 as a therapeutic target and inform future efforts to rationally design inhibitors against the complex, a deep understanding of SHOC2 structural features that govern complex assembly and function is needed. We have now resolved the SHOC2 complex structure and have profiled the functional consequence of nearly all possible amino acid substitutions at every position of SHOC2 through deep mutational scanning. These findings highlight novel structural features of SHOC2 that may be important to therapeutically target in RAS mutant cancers. The studies outlined in this proposal will build on this work and aim to (1) define and validate the structural features of SHOC2 holophosphatase complex specificity toward RAF substrate, and (2) identify novel post-translational modifications that regulate SHOC2 function. The training I receive in the K99 phase will provide me the necessary expertise in structural biology, proteomics, and ubiquitin biology as well as equip me with teaching/lab management experience so that I can effectively lead my own independent research program. The K99/R00 will be instrumental in helping me achieve my long-term goal, to establish my laboratory focused on studying novel RAS-associated vulnerabilities and drug resistance mechanisms via systematic approaches in cell signaling and protein structure-function.