RAP1GAP2

RAP1GAP2 is a Rap1-specific GTPase-activating protein that terminates Rap1 signaling to regulate integrin-mediated cell adhesion, platelet dense granule secretion, and axon outgrowth. In platelets, RAP1GAP2 GAP activity is modulated by phosphorylation: agonist-stimulated phosphorylation of Ser9 recruits 14-3-3 proteins to attenuate RAP1GAP2-mediated inhibition of adhesion, while cGKI/cAKI-mediated phosphorylation of the adjacent Ser7 disrupts 14-3-3 binding, restoring the capacity of RAP1GAP2 to suppress Rap1 (PMID:15632203, PMID:18039662). RAP1GAP2 forms a trimeric complex with synaptotagmin-like protein 1 (Slp1) and Rab27 through a C-terminal TKXT motif, and this interaction is required for RAP1GAP2-dependent augmentation of dense granule secretion in permeabilized platelets (PMID:19528539). In neurons, Rap1gap2 limits Rap1-dependent axon outgrowth and branching, as demonstrated by gain- and loss-of-function studies in olfactory sensory neurons (PMID:22732430).

1. 2005 High

   Identification of RAP1GAP2 as a platelet-expressed Rap1 GAP and discovery of its phosphorylation by cGKI on Ser7 established the first molecular link between cGMP-dependent inhibitory signaling and Rap1 inactivation in platelets.

   Evidence cDNA cloning from human platelet library, transfection-based GAP assays, in vitro kinase assay with cGKI, and site-directed mutagenesis of Ser7

   PMID:15632203

   Open questions at the time

   • Functional consequence of Ser7 phosphorylation on GAP activity or platelet function was not determined
   • The mechanism by which RAP1GAP2 is recruited to Rap1 at the platelet membrane was unknown
2. 2006 Medium

   Demonstrating that the GoLoco motif of RAP1GAP2 is functionally inactive toward Gαi/o ruled out a dual-function model in which RAP1GAP2 might simultaneously regulate both Rap1 and heterotrimeric G-protein signaling.

   Evidence In vitro GDI activity measurement and Lys75Arg mutagenesis

   PMID:16949794

   Open questions at the time

   • Only a single study; independent confirmation of GoLoco inactivity is lacking
   • Whether the GoLoco motif has any non-canonical function (e.g., protein–protein interaction scaffold) was not tested
3. 2007 High

   Discovery that 14-3-3 binds phospho-Ser9 of RAP1GAP2 and that Ser7 phosphorylation by cGKI/cAKI disrupts this interaction resolved how opposing platelet-activating and -inhibiting signals converge on RAP1GAP2 to control cell adhesion.

   Evidence Endogenous co-immunoprecipitation in platelets, in vitro phosphorylation, and cell adhesion assays with pharmacological inhibition of cGK/cAK

   PMID:18039662

   Open questions at the time

   • 14-3-3 binding did not alter catalytic GAP activity in vitro, leaving the mechanism by which it attenuates adhesion inhibition unresolved
   • In vivo relevance of the Ser7/Ser9 phospho-switch in thrombosis or hemostasis was not tested
4. 2009 High

   Identification of the RAP1GAP2–Slp1–Rab27 trimeric complex and its requirement for dense granule secretion revealed a GAP-independent effector function of RAP1GAP2 in platelet exocytosis.

   Evidence Yeast two-hybrid screen, endogenous co-IP in platelets, TKXT motif deletion mapping, and reconstituted secretion assay with purified proteins in permeabilized platelets

   PMID:19528539

   Open questions at the time

   • Whether GAP catalytic activity and Slp1 binding are coordinated or independent during platelet activation is unknown
   • Structural basis of the TKXT–C2A interaction has not been determined
5. 2012 Medium

   Demonstrating that Rap1gap2 restricts Rap1-dependent axon outgrowth in olfactory sensory neurons extended its functional role beyond platelets to neuronal development.

   Evidence Overexpression and siRNA knockdown in Neuro-2a cells and in vivo olfactory sensory neuron model with neurite length quantification

   PMID:22732430

   Open questions at the time

   • Downstream effectors linking Rap1 inactivation to cytoskeletal remodeling in neurons were not identified
   • Findings are from a single lab; genetic knockout confirmation is lacking

• Key unresolved questions include the structural basis of RAP1GAP2 catalysis and regulation, the in vivo consequences of RAP1GAP2 loss for hemostasis and neuronal circuit formation, and whether the Slp1/Slp2-a interactions represent tissue-specific modes of RAP1GAP2 regulation.
• No knockout mouse phenotype has been reported
• No structural model of the GAP domain or regulatory phospho-switch exists
• Relative contributions of GAP activity versus Slp1-mediated secretion function in platelet physiology are unresolved