title: Serine-71 phosphorylation of Rac1 modulates downstream signaling.
abstract: The Rho GTPases Rac1 and Cdc42 regulate a variety of cellular functions by signaling to different signal pathways. It is believed that the presence of a specific effector at the location of GTPase activation determines the route of downstream signaling. We previously reported about EGF-induced Ser-71 phosphorylation of Rac1/Cdc42. By using the phosphomimetic S71E-mutants of Rac1 and Cdc42 we investigated the impact of Ser-71 phosphorylation on binding to selected effector proteins. Binding of the constitutively active (Q61L) variants of Rac1 and Cdc42 to their specific interaction partners Sra-1 and N-WASP, respectively, as well as to their common effector protein PAK was abrogated when Ser-71 was exchanged to glutamate as phosphomimetic substitution. Interaction with their common effector proteins IQGAP1/2/3 or MRCK alpha was, however, hardly affected. This ambivalent behaviour was obvious in functional assays. In contrast to Rac1 Q61L, phosphomimetic Rac1 Q61L/S71E was not able to induce increased membrane ruffling. Instead, Rac1 Q61L/S71E allowed filopodia formation, which is in accordance with abrogation of the dominant Sra-1/Wave signalling pathway. In addition, in contrast to Rac1 transfected cells Rac1 S71E failed to activate PAK1/2. On the other hand, Rac1 Q61L/S71E was as effective in activation of NF-kappaB as Rac1 Q61L, illustrating positive signal transduction of phosphorylated Rac1. Together, these data suggest that phosphorylation of Rac1 and Cdc42 at serine-71 represents a reversible mechanism to shift specificity of GTPase/effector coupling, and to preferentially address selected downstream pathways.
authors.affiliation: Department of Toxicology, Hannover Medical School, Hannover, Germany.
authors.forename: Janett
authors.initials: J
authors.lastname: Schwarz
authors.forename: Julia
authors.initials: J
authors.lastname: Proff
authors.forename: Anika
authors.initials: A
authors.lastname: Hävemeier
authors.forename: Markus
authors.initials: M
authors.lastname: Ladwein
authors.forename: Klemens
authors.initials: K
authors.lastname: Rottner
authors.forename: Britta
authors.initials: B
authors.lastname: Barlag
authors.forename: Andreas
authors.initials: A
authors.lastname: Pich
authors.forename: Helma
authors.initials: H
authors.lastname: Tatge
authors.forename: Ingo
authors.initials: I
authors.lastname: Just
authors.forename: Ralf
authors.initials: R
authors.lastname: Gerhard
mesh_terms: D000818:Animals; D004789:Enzyme Activation; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D050505:Mutant Proteins / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009479:Neuropeptides / Q000172:deficiency / Q000378:metabolism*; D010641:Phenotype; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism*; D011485:Protein Binding; D011554:Pseudopodia / Q000378:metabolism; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D054462:p21-Activated Kinases / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000172:deficiency / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein
chemical_list: D050505:Mutant Proteins; D016328:NF-kappa B; D009479:Neuropeptides; C484602:Rac1 protein, mouse; D010768:Phosphoserine; D054462:p21-Activated Kinases; D020764:cdc42 GTP-Binding Protein; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein