title: Autophosphorylation on S614 inhibits the activity and the transforming potential of BRAF.
abstract: The BRAF proto-oncogene serine/threonine-protein kinase, known as BRAF, belongs to the RAF kinase family. It regulates the MAPK/ERK signalling pathway affecting several cellular processes such as growth, survival, differentiation, and cellular transformation. BRAF is mutated in ~8% of all human cancers with the V600E mutation constituting ~90% of mutations. Here, we have used quantitative mass spectrometry to map and compare phosphorylation site patterns between BRAF and BRAF V600E. We identified sites that are shared as well as several quantitative differences in phosphorylation abundance. The highest difference is phosphorylation of S614 in the activation loop which is ~5fold enhanced in BRAF V600E. Mutation of S614 increases the kinase activity of both BRAF and BRAF V600E and the transforming ability of BRAF V600E. The phosphorylation of S614 is mitogen inducible and the result of autophosphorylation. These data suggest that phosphorylation at this site is inhibitory, and part of the physiological shut-down mechanism of BRAF signalling.
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany.
authors.forename: Layal
authors.initials: L
authors.lastname: Dernayka
authors.affiliation: Systems Biology Ireland, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Nora
authors.initials: N
authors.lastname: Rauch
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany.
authors.forename: Mohamed-Ali
authors.initials: MA
authors.lastname: Jarboui
authors.affiliation: Systems Biology Ireland, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Armin
authors.initials: A
authors.lastname: Zebisch
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany.
authors.forename: Yves
authors.initials: Y
authors.lastname: Texier
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany.
authors.forename: Nicola
authors.initials: N
authors.lastname: Horn
authors.affiliation: Systems Biology Ireland, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: David
authors.initials: D
authors.lastname: Romano
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany; Deutsches Zentrum für Neurodegenerative Erkrankungen e. V., Otfried-Müller Strasse 23, 72076 Tübingen, Germany.
authors.forename: Christian Johannes
authors.initials: CJ
authors.lastname: Gloeckner
authors.affiliation: Systems Biology Ireland, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Alex von
authors.initials: AV
authors.lastname: Kriegsheim
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany.
authors.forename: Marius
authors.initials: M
authors.lastname: Ueffing
authors.affiliation: Systems Biology Ireland, University College Dublin, Belfield, Dublin 4, Ireland; Conway Institute of Biomolecular & Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland; School of Medicine and Medical Science, University College Dublin, Belfield, Dublin 4, Ireland. Electronic address: walter.kolch@ucd.ie.
authors.forename: Walter
authors.initials: W
authors.lastname: Kolch
authors.affiliation: Medical Proteome Center, Division for Experimental Ophthalmology, Institute for Ophthalmic Research, University of Tuebingen, Tuebingen, Germany. Electronic address: karsten.boldt@uni-tuebingen.de.
authors.forename: Karsten
authors.initials: K
authors.lastname: Boldt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000378:metabolism* / Q000473:pathology*; D006801:Humans; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism*; D011485:Protein Binding; D000090063:Proto-Oncogene Mas; D048493:Proto-Oncogene Proteins B-raf / Q000737:chemistry / Q000378:metabolism*; D051381:Rats
chemical_list: C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D010768:Phosphoserine; D048493:Proto-Oncogene Proteins B-raf; D020929:Mitogen-Activated Protein Kinase Kinases
keywords: BRAF; Inhibition of kinase activity and transformation; S614 autophosphorylation