Structural and Energetic Mechanisms of Cooperative Autoinhibition and Activation of Vav1

Department of Biochemistry, Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8816, USA.
Cell (Impact Factor: 32.24). 01/2010; 140(2):246-56. DOI: 10.1016/j.cell.2009.12.033
Source: PubMed


Vav proteins are guanine nucleotide exchange factors (GEFs) for Rho family GTPases. They control processes including T cell activation, phagocytosis, and migration of normal and transformed cells. We report the structure and biophysical and cellular analyses of the five-domain autoinhibitory element of Vav1. The catalytic Dbl homology (DH) domain of Vav1 is controlled by two energetically coupled processes. The DH active site is directly, but weakly, inhibited by a helix from the adjacent Acidic domain. This core interaction is strengthened 10-fold by contacts of the calponin homology (CH) domain with the Acidic, pleckstrin homology, and DH domains. This construction enables efficient, stepwise relief of autoinhibition: initial phosphorylation events disrupt the modulatory CH contacts, facilitating phosphorylation of the inhibitory helix and consequent GEF activation. Our findings illustrate how the opposing requirements of strong suppression of activity and rapid kinetics of activation can be achieved in multidomain systems.

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Available from: Diana R. Tomchick, Mar 02, 2014

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Article: Structural and Energetic Mechanisms of Cooperative Autoinhibition and Activation of Vav1

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    • "N-terminal to these are a calponin homology (CH) domain and an acidic region that negatively regulate the GEF activity of Vav1 by binding to the DH and PH domains and sterically blocking binding of the GTPase substrates (Aghazadeh et al., 2000; Yu et al., 2010). Phosphorylation of tyrosine residues located within the acidic domain leads to dissociation of the CH and acidic domains from the DH-PH-C1 catalytic core of Vav1 and hence increased enzymatic activity (Aghazadeh et al., 2000; Yu et al., 2010). C-terminal to the DH-PH-C1 domains of Vav1, the protein contains one SH2 domain and two SH3 domains (SH3A, SH3B), suggesting that Vav1 may function as an adaptor as well as a GEF. "
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    ABSTRACT: The antigen-specific binding of T cells to antigen presenting cells results in recruitment of signaling proteins to microclusters at the cell-cell interface known as the immunological synapse (IS). The Vav1 guanine nucleotide exchange factor plays a critical role in TCR signaling, leading to the activation of multiple pathways. We now show that it is recruited to microclusters and to the IS in primary CD4(+) and CD8(+) T cells. Furthermore we show that this recruitment depends on the SH2 and C-terminal SH3 (SH3(B)) domains of Vav1, and on phosphotyrosines 112 and 128 of the SLP76 adaptor protein. Biophysical measurements show that Vav1 binds directly to these residues on SLP76 and that efficient binding depends on the SH2 and SH3(B) domains of Vav1. Finally, we show that the same two domains are critical for the phosphorylation of Vav1 and its signaling function in TCR-induced calcium flux. We propose that Vav1 is recruited to the IS by binding to SLP76 and that this interaction is critical for the transduction of signals leading to calcium flux.
    Journal of Cell Science 09/2012; 125(22). DOI:10.1242/jcs.105148 · 5.43 Impact Factor
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    • "However, this hypothesis is unlikely because we did not detect an increase in Vav2 tyrosine phosphorylation upon Wnt stimulation. Alternatively and more likely, it is possible that the interaction frees the Vav2 catalytic Dbl homology domain from the coordinated auto-inhibition by the Acidic and Calponin homology domains (Yu et al., 2010). Thus, p120-catenin interaction would mimic the effects of Vav2 phosphorylation in the Acidic domain. "
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    ABSTRACT: A role for Rac1 GTPase in canonical Wnt signalling has been recently demonstrated, being required for β-catenin translocation to the nucleus. In this article we have investigated the mechanism of Rac1 stimulation by Wnt. Up-regulation of Rac1activity by Wnt3a temporally correlates with enhanced p120-catenin binding to Rac1 and Vav2. Vav2 and Rac1 association with p120-catenin is modulated by phosphorylation of this protein: it is stimulated upon serine/threonine phosphorylation by CK1 and inhibited by tyrosine phosphorylation by Src or Fyn. Acting on these two post-translational modifications, Wnt3a induces the release of p120-catenin from E-cadherin, enables p120-catenin interaction with Vav2 and Rac1 and facilitates Rac1 activation by Vav2. Since p120-catenin depletion disrupts gastrulation in Xenopus, we analysed p120-catenin mutants for their ability to rescue this phenotype. In contrast to the wild-type protein or other controls, p120-catenin point mutants deficient in the release from E-cadherin or in Vav2- or Rac1-binding failed to rescue p120-catenin depletion. Collectively, these results indicate that p120-catenin binding to Vav2 and Rac1 is required for the activation of this GTPase upon Wnt signalling.
    Journal of Cell Science 09/2012; 125(22). DOI:10.1242/jcs.101030 · 5.43 Impact Factor
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    • "They all display a conserved tyrosine residue (Y174, Y172 and Y173 respectively) in the short N-terminal helix that gets phosphorylated by Src Family- Kinases (SFK), destructuring the helix to facilitate access to the DH domain. These phosphorylations are considered as signatures of the active state of those GEFs (Aghazadeh et al., 2000; Colomba et al., 2008; Han et al., 1997; Lopez-Lago et al., 2000; Yohe et al., 2007; Yu et al., 2010). Mutations or deletions of this N-terminal domain often give rise to constitutive activation (Colacios et al., 2011; Schuebel et al., 1998). "
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    ABSTRACT: Rho GTPases act as molecular switches central in cellular processes such as cytoskeleton dynamics, migration, cell proliferation, growth or survival. Their activation is tightly regulated downstream of cell surface receptors by Guanine nucleotide Exchange Factors (GEFs), that are responsible for the specificity, the accuracy, and the spatial restriction of Rho GTPases response to extracellular cues. Because there is about four time more RhoGEFs that Rho GTPases, and GEFs do not always show a strict specificity for GTPases, it is clear that their regulation depends on specific interactions with the subcellular environment. RhoGEFs bear a peculiar structure, highly conserved though evolution, consisting of a DH-PH tandem, the DH (Dbl homology) domain being responsible for the exchange activity. The function of the PH (Pleckstrin homology) domain known to bind phosphoinositides, however, remains elusive, and reports are in many cases rather confusing. This review summarizes data on the regulation of RhoGEFs activity through interaction of the PH-associated DH domain with phosphoinositides which are considered as critical players in the spatial organization of major signaling pathways.
    05/2012; 52(2):303-14. DOI:10.1016/j.jbior.2012.04.001
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