Rawson, ShaunKim, Byeong-WonSharif, HumayunJeon, HyesungPark, EunyoungLi, KunhuaFicarro, ScottGonzalez-De Pino, GonzaloMarto, JarrodEck, Michael2022-07-272019-10-03Park, Eunyoung, Shaun Rawson, Kunhua Li, Byeong-Won Kim, Scott B. Ficarro, Gonzalo Gonzalez-Del Pino, Humayun Sharif, Jarrod A. Marto, Hyesung Jeon, and Michael J. Eck. 2019. Architecture of Autoinhibited and Active BRAF-MEK1-14-3-3 Complexes. Nature 575, no. 7783: 545-50.0028-08361476-4687https://nrs.harvard.edu/URN-3:HUL.INSTREPOS:37372897RAF family kinases are RAS-activated switches that initiate signaling through the MAP kinase cascade to control cellular proliferation, differentiation and survival1-3. RAF activity is tightly regulated and inappropriate activation is a frequent cause of cancer4-6. At present, the structural basis for RAF regulation is poorly understood. Here we describe autoinhibited and active state structures of full-length BRAF in complexes with MEK1 and a 14-3-3 dimer, determined using cryo electron microscopy (cryo-EM). A 4.1Å resolution cryo-EM reconstruction reveals an inactive BRAF/MEK1 complex restrained in a cradle formed by the 14-3-3 dimer, which binds the phosphorylated S365 and S729 sites that flank the BRAF kinase domain. The BRAF cysteine-rich domain (CRD) occupies a central position that stabilizes this assembly, but the adjacent RAS-binding domain (RBD) is poorly ordered and peripheral. The 14-3-3 cradle maintains autoinhibition by sequestering the membrane-binding CRD and blocking dimerization of the BRAF kinase domain. In the active state, these inhibitory interactions are released and a single 14-3-3 dimer rearranges to bridge the C-terminal pS729 binding sites of two BRAFs, driving formation of an active, back-to-back BRAF dimer. Our structural snapshots provide a foundation for understanding normal RAF regulation and its mutational disruption in cancer and developmental syndromes.en-USMultidisciplinaryJournal Article2022-07-2710.1038/s41586-019-1660-y