title: Mechanistic insight into GPCR-mediated activation of the microtubule-associated RhoA exchange factor GEF-H1.
abstract: The RhoGEF GEF-H1 can be sequestered in an inactive state on polymerized microtubules by the dynein motor light-chain Tctex-1. Phosphorylation of GEF-H1 Ser885 by PKA or PAK kinases creates an inhibitory 14-3-3-binding site. Here we show a new mode of GEF-H1 activation in response to the G-protein-coupled receptor (GPCR) ligands lysophosphatidic acid (LPA) or thrombin that is independent of microtubule depolymerization. LPA/thrombin stimulates disassembly of the GEF-H1:dynein multi-protein complex through the concerted action of Gα and Gβγ. Gα binds directly to GEF-H1 and displaces it from Tctex-1, while Gβγ binds to Tctex-1 and disrupts its interaction with the dynein intermediate chain, resulting in the release of GEF-H1. Full activation of GEF-H1 requires dephosphorylation of Ser885 by PP2A, which is induced by thrombin. The coordinated displacement of GEF-H1 from microtubules by G-proteins and its dephosphorylation by PP2A demonstrate a multistep GEF-H1 activation and present a unique mechanism coupling GPCR signalling to Rho activation.
authors.affiliation: Department of Biology, Technion Israel Institute of Technology, Haifa 32000, Israel.
authors.forename: David
authors.initials: D
authors.lastname: Meiri
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, University of Toronto, 101 College Street, Room 12-704, Toronto Medical Discovery Tower, Toronto, Ontario, Canada M5G 1L7.
authors.forename: Christopher B
authors.initials: CB
authors.lastname: Marshall
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, University of Toronto, 101 College Street, Room 12-704, Toronto Medical Discovery Tower, Toronto, Ontario, Canada M5G 1L7.
authors.forename: Daphna
authors.initials: D
authors.lastname: Mokady
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, University of Toronto, 101 College Street, Room 12-704, Toronto Medical Discovery Tower, Toronto, Ontario, Canada M5G 1L7.
authors.forename: Jose
authors.initials: J
authors.lastname: LaRose
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Avenue, Room 992A, Toronto, Ontario, Canada M5G 1X5.
authors.forename: Michael
authors.initials: M
authors.lastname: Mullin
authors.affiliation: 1] Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Avenue, Room 992A, Toronto, Ontario, Canada M5G 1X5 [2] Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada M5S 1A8.
authors.forename: Anne-Claude
authors.initials: AC
authors.lastname: Gingras
authors.affiliation: 1] Princess Margaret Cancer Centre, University Health Network, University of Toronto, 101 College Street, Room 12-704, Toronto Medical Discovery Tower, Toronto, Ontario, Canada M5G 1L7 [2] Department of Medical Biophysics, University of Toronto, 1 King's College Circle, Toronto, Ontario, Canada M5S 1A8.
authors.forename: Mitsuhiko
authors.initials: M
authors.lastname: Ikura
authors.affiliation: 1] Princess Margaret Cancer Centre, University Health Network, University of Toronto, 101 College Street, Room 12-704, Toronto Medical Discovery Tower, Toronto, Ontario, Canada M5G 1L7 [2] Department of Medical Biophysics, University of Toronto, 1 King's College Circle, Toronto, Ontario, Canada M5S 1A8 [3] Department of Medicine, University of Toronto, 1 King's College Circle, Toronto, Ontario, Canada M5S 1A8 [4] Department of Immunology, University of Toronto, 1 King's College Circle, Toronto, Ontario, Canada M5S 1A8 [5] Division of Rheumatology, St Michael's Hospital, 30 Bond Street, Toronto, Ontario, Canada M5B 1W8.
authors.forename: Robert
authors.initials: R
authors.lastname: Rottapel
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism; D000818:Animals; D004398:Dyneins / Q000378:metabolism*; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000187:drug effects / Q000378:metabolism; D044387:GTP-Binding Protein beta Subunits / Q000187:drug effects / Q000378:metabolism; D044388:GTP-Binding Protein gamma Subunits / Q000187:drug effects / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D008870:Microtubules / Q000378:metabolism*; D010766:Phosphorylation; D043562:Receptors, G-Protein-Coupled / Q000187:drug effects / Q000378:metabolism*; D064067:Rho Guanine Nucleotide Exchange Factors / Q000187:drug effects / Q000378:metabolism*; D013917:Thrombin / Q000494:pharmacology; D054462:p21-Activated Kinases / Q000378:metabolism
chemical_list: D048948:14-3-3 Proteins; C577402:Arhgef2 protein, mouse; C533386:Dynlt1b protein, mouse; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D008246:Lysophospholipids; D043562:Receptors, G-Protein-Coupled; D064067:Rho Guanine Nucleotide Exchange Factors; D054462:p21-Activated Kinases; D013917:Thrombin; D004398:Dyneins; D044365:GTP-Binding Protein alpha Subunits, G12-G13; C032881:lysophosphatidic acid