RASopathy mutations provide functional insight into the BRAF cysteine-rich domain and reveal the importance of autoinhibition in BRAF regulation

• Published in: Molecular cell Vol. 82; no. 22; pp. 4262 - 4276.e5
• Main Authors: Spencer-Smith, Russell, Terrell, Elizabeth M., Insinna, Christine, Agamasu, Constance, Wagner, Morgan E., Ritt, Daniel A., Stauffer, Jim, Stephen, Andrew G., Morrison, Deborah K.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 17.11.2022
• Subjects: autoinhibition; BRAF; CRAF; CRD; Cysteine - genetics; cysteine-rich domain; development; Humans; Mutation; phosphatidylserine; Protein Domains; Proto-Oncogene Proteins B-raf - genetics; RAF kinases; RAS; RASopathies; Syndrome; phosphatidylserine; RAF kinases; development; CRD; RAS; BRAF; CRAF; cysteine-rich domain; RASopathies; autoinhibition
• ISSN: 1097-2765; 1097-4164; 1097-4164
• DOI: 10.1016/j.molcel.2022.10.016

BRAF is frequently mutated in human cancer and the RASopathy syndromes, with RASopathy mutations often observed in the cysteine-rich domain (CRD). Although the CRD participates in phosphatidylserine (PS) binding, the RAS-RAF interaction, and RAF autoinhibition, the impact of these activities on RAF function in normal and disease states is not well characterized. Here, we analyze a panel of CRD mutations and show that they increase BRAF activity by relieving autoinhibition and/or enhancing PS binding, with relief of autoinhibition being the major factor determining mutation severity. Further, we show that CRD-mediated autoinhibition prevents the constitutive plasma membrane localization of BRAF that causes increased RAS-dependent and RAS-independent function. Comparison of the BRAF- and CRAF-CRDs also indicates that the BRAF-CRD is a stronger mediator of autoinhibition and PS binding, and given the increased catalytic activity of BRAF, our studies reveal a more critical role for CRD-mediated autoinhibition in BRAF regulation. [Display omitted] •RASopathy BRAF-CRD mutations enhance membrane binding and/or relieve autoinhibition•Relief of autoinhibition is the major determinant of BRAF-CRD mutant severity•CRD-mediated autoinhibition is required to prevent aberrant BRAF signaling•BRAF-CRD has greater autoinhibitory and membrane binding activity than the CRAF-CRD Spencer-Smith et al. demonstrate that RASopathy mutations in the BRAF cysteine-rich domain (CRD) increase BRAF biological activity by relieving autoinhibition and/or enhancing membrane binding. Importantly, they show that CRD-mediated autoinhibition is essential for preventing aberrant BRAF signaling, whereas the CRAF-CRD plays a lesser role in autoinhibition and membrane binding.