Abstract LB029: Comprehensive structure-function evaluation of the SHOC2 holophosphatase reveals disease mechanisms and therapeutic opportunities

Abstract

Abstract Receptor tyrosine kinase (RTK)-RAS signaling through the downstream mitogen activated protein kinase (MAPK) cascade regulate cell proliferation and survival. The SHOC2 holophosphatase functions as a key regulator of RTK-RAS signaling by removing an inhibitory phosphorylation on RAF family proteins to potentiate MAPK signaling. SHOC2 forms a ternary complex with MRAS and PP1C, and human germline gain-of-function mutations of this complex result in congenital RASopathy syndromes. However, the structure and the assembly of this complex are poorly understood. Here, we use cryogenic electron microscopy (cryo-EM) to resolve the structure of the SHOC2-PP1C-MRAS holophosphatase to 2.9Å resolution. Furthermore, we define the biophysical principles of SHOC2 holoenzyme interactions, elucidate the assembly order of the complex, and systematically interrogate the functional consequences of nearly all possible missense variants of SHOC2 through deep mutational scanning. We demonstrate that SHOC2 binds PP1C and MRAS through a concave surface created by the leucine-rich repeat region and further engages PP1C through the N-terminal disordered region containing a degenerate RVxF motif. Initial complex formation is mediated by SHOC2-PP1C interactions and is stabilized by binding of GTP-loaded MRAS. These observations define how clinical mutants in Noonan-like syndrome and cancer stabilize interactions of complex members to enhance holophosphatase activity. Together, this integrative structure-function model comprehensively defines key binding interactions within the SHOC2 holophosphatase, informing therapeutic development. Citation Format: Jason Kwon, Behnoush Hajian, Yuemin Bian, Alvaro Amor, James Fuller, Cara Fraley, Laura Baker, Jonathan So, David Mayhew, Abbey Sykes, Nicole Persky, Xiaoping Yang, David Root, Charles Perry, Christopher Lemke, William Hahn, Andrew Aguirre. Comprehensive structure-function evaluation of the SHOC2 holophosphatase reveals disease mechanisms and therapeutic opportunities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB029.