title: Distinct requirement for an intact dimer interface in wild-type, V600E and kinase-dead B-Raf signalling.
abstract: The dimerisation of Raf kinases involves a central cluster within the kinase domain, the dimer interface (DIF). Yet, the importance of the DIF for the signalling potential of wild-type B-Raf (B-Raf(wt)) and its oncogenic counterparts remains unknown. Here, we show that the DIF plays a pivotal role for the activity of B-Raf(wt) and several of its gain-of-function (g-o-f) mutants. In contrast, the B-Raf(V600E), B-Raf(insT) and B-Raf(G469A) oncoproteins are remarkably resistant to mutations in the DIF. However, compared with B-Raf(wt), B-Raf(V600E) displays extended protomer contacts, increased homodimerisation and incorporation into larger protein complexes. In contrast, B-Raf(wt) and Raf-1(wt) mediated signalling triggered by oncogenic Ras as well as the paradoxical activation of Raf-1 by kinase-inactivated B-Raf require an intact DIF. Surprisingly, the B-Raf DIF is not required for dimerisation between Raf-1 and B-Raf, which was inactivated by the D594A mutation, sorafenib or PLX4720. This suggests that paradoxical MEK/ERK activation represents a two-step mechanism consisting of dimerisation and DIF-dependent transactivation. Our data further implicate the Raf DIF as a potential target against Ras-driven Raf-mediated (paradoxical) ERK activation.
authors.affiliation: Spemann Graduate School of Biology and Medicine, Albert-Ludwigs-University of Freiburg, Freiburg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Röring
authors.forename: Ricarda
authors.initials: R
authors.lastname: Herr
authors.forename: Gina J
authors.initials: GJ
authors.lastname: Fiala
authors.forename: Katharina
authors.initials: K
authors.lastname: Heilmann
authors.forename: Sandra
authors.initials: S
authors.lastname: Braun
authors.forename: Anja E
authors.initials: AE
authors.lastname: Eisenhardt
authors.forename: Sebastian
authors.initials: S
authors.lastname: Halbach
authors.forename: David
authors.initials: D
authors.lastname: Capper
authors.forename: Andreas
authors.initials: A
authors.lastname: von Deimling
authors.forename: Wolfgang W
authors.initials: WW
authors.lastname: Schamel
authors.forename: Darren N
authors.initials: DN
authors.lastname: Saunders
authors.forename: Tilman
authors.initials: T
authors.lastname: Brummer
mesh_terms: D001557:Benzenesulfonates / Q000494:pharmacology; D018938:Caco-2 Cells; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D009536:Niacinamide / Q000031:analogs & derivatives; D010671:Phenylurea Compounds; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D055503:Protein Multimerization / Q000187:drug effects / Q000502:physiology*; D048493:Proto-Oncogene Proteins B-raf / Q000378:metabolism*; D011725:Pyridines / Q000494:pharmacology; D000077157:Sorafenib; D013449:Sulfonamides / Q000494:pharmacology
chemical_list: D001557:Benzenesulfonates; D007211:Indoles; C528407:PLX 4720; D010671:Phenylurea Compounds; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D013449:Sulfonamides; D009536:Niacinamide; D000077157:Sorafenib; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf