Structure-function analysis of the SHOC2-MRAS-PP1C holophosphatase complex

  • Nature. 2022 Sep;609(7926):408-415. doi: 10.1038/s41586-022-04928-2.
Jason J Kwon  #  1  2  3 ,  Behnoush Hajian  #  4 ,  Yuemin Bian  #  4 ,  Lucy C Young  #  5 ,  Alvaro J Amor  4 ,  James R Fuller  6 ,  Cara V Fraley  4 ,  Abbey M Sykes  3  4 ,  Jonathan So  1  2  3 ,  Joshua Pan  1  2  3 ,  Laura Baker  4 ,  Sun Joo Lee  1  2  3 ,  Douglas B Wheeler  1  2 ,  David L Mayhew  1  2  3 ,  Nicole S Persky  7 ,  Xiaoping Yang  7 ,  David E Root  7 ,  Anthony M Barsotti  8 ,  Andrew W Stamford  8 ,  Charles K Perry  4 ,  Alex Burgin  4 ,  Frank McCormick  5  9 ,  Christopher T Lemke  10 ,  William C Hahn  11  12  13  14 ,  Andrew J Aguirre  15  16  17  18
Affiliations
  • 1. Cancer Program, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
  • 2. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA.
  • 3. Harvard Medical School, Boston, Massachusetts, USA.
  • 4. Center for the Development of Therapeutics, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
  • 5. Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, CA, USA.
  • 6. Helix Biostructures, Indianapolis, IN, USA.
  • 7. Genetic Perturbation Platform, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
  • 8. Deerfield Discovery and Development, Deerfield Management, New York, NY, USA.
  • 9. NCI RAS Initiative, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD, USA.
  • 10. Center for the Development of Therapeutics, Broad Institute of MIT and Harvard, Cambridge, MA, USA. [email protected].
  • 11. Cancer Program, Broad Institute of MIT and Harvard, Cambridge, MA, USA. [email protected].
  • 12. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA. [email protected].
  • 13. Harvard Medical School, Boston, Massachusetts, USA. [email protected].
  • 14. Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA. [email protected].
  • 15. Cancer Program, Broad Institute of MIT and Harvard, Cambridge, MA, USA. [email protected].
  • 16. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, USA. [email protected].
  • 17. Harvard Medical School, Boston, Massachusetts, USA. [email protected].
  • 18. Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA. [email protected].
  • # Contributed equally.
PMID: 35831509 DOI: 10.1038/s41586-022-04928-2
Abstract

Receptor tyrosine kinase (RTK)-RAS signalling through the downstream mitogen-activated protein kinase (MAPK) cascade regulates cell proliferation and survival. The SHOC2-MRAS-PP1C holophosphatase complex functions as a key regulator of RTK-RAS signalling by removing an inhibitory phosphorylation event on the Raf family of proteins to potentiate MAPK signalling1. SHOC2 forms a ternary complex with MRAS and PP1C, and human germline gain-of-function mutations in this complex result in congenital RASopathy syndromes2-5. However, the structure and assembly of this complex are poorly understood. Here we use cryo-electron microscopy to resolve the structure of the SHOC2-MRAS-PP1C complex. We define the biophysical principles of holoenzyme interactions, elucidate the assembly order of the complex, and systematically interrogate the functional consequence of nearly all of the possible missense variants of SHOC2 through deep mutational scanning. We show that SHOC2 binds PP1C and MRAS through the concave surface of the leucine-rich repeat region and further engages PP1C through the N-terminal disordered region that contains a cryptic RVXF motif. Complex formation is initially mediated by interactions between SHOC2 and PP1C and is stabilized by the binding of GTP-loaded MRAS. These observations explain how mutant versions of SHOC2 in RASopathies and Cancer stabilize the interactions of complex members to enhance holophosphatase activity. Together, this integrative structure-function model comprehensively defines key binding interactions within the SHOC2-MRAS-PP1C holophosphatase complex and will inform therapeutic development .