The C-Terminal SH3 Domain Contributes to the Intramolecular Inhibition of Vav Family Proteins

Abstract

Vav proteins are phosphorylation-dependent guanine nucleotide exchange factors (GEFs) that catalyze the activation of members of the Rho family of guanosine triphosphatases (GTPases). The current regulatory model holds that the nonphosphorylated, catalytically inactive state of these GEFs is maintained by intramolecular interactions among the amino-terminal domains and the central catalytic core, which block the binding of Vav proteins to GTPases. We showed that this autoinhibition is mechanistically more complex, also involving the bivalent association of the carboxyl-terminal Src homology 3 (SH3) region of Vav with its catalytic and pleckstrin homology (PH) domains. Such interactions occurred through proline-rich region-independent mechanisms. Full release from this double-locked state required synergistic weakening effects from multiple phosphorylated tyrosine residues, thus providing an optimized system to generate gradients of Vav GEF activity depending on upstream signaling inputs. This mechanism is shared by mammalian and Drosophila melanogaster Vav proteins, suggesting that it may be a common regulatory feature for this protein family.

Integrated mate-pair and RNA sequencing identifies novel, targetable gene fusions in peripheral T-cell lymphoma

“…We focused on RAC1 because mutation of the VAV1 CSH3 domain strongly activates RAC1, whereas its effects on the r GTPases CDC42 and RHOA are more modest. 27 RAC1 also has been shown to be activated by NPM-ALK in ALCL cells, and a Rac small molecule inhibitor blocked lymphoma development and dissemination. 50,51 We showed that cells expressing VAV1-GSS were exquisitely sensitive to azathioprine, which inhibits activation of RAC1 but not of CDC42 or RHOA, 33,34 Other clinically available drugs that inhibit RAC1 and have been proposed to be repurposed as anticancer agents include metformin, statins such as simvastatin, and R-ketorolac.…”

“…Because the VAV1 CSH3 domain mediates autoinhibition of VAV1 GEF activity, 27 we hypothesized that VAV1 fusions, which lack this domain, promote VAV1 activation. Because VAV1 mediates cytoskeletal remodeling, 28 we first performed immunofluorescence for actin in adherent NIH-3T3 fibroblasts transfected with VAV1-GSS, wild-type (wt)VAV1, or an empty control vector.…”

Genetic Dissection of the Vav2-Rac1 Signaling Axis in Vascular Smooth Muscle Cells

“…To this end, we used standard genomic recombination techniques to insert an ectopic cassette containing a cDNA fragment encoding a truncated, hemagglutinin (HA)-tagged Vav2 ⌬1-184 mutant protein (here termed OncoVav2) downstream of the first exon of the mouse Vav2 locus. We have shown before that this type of truncated Vav protein shows phosphorylation-independent, constitutive exchange activity due to the removal of the N-terminal inhibitory domains (14,(27)(28)(29). To make possible the inducible expression of this protein, this cassette contained 5=-located Stop-Neo R DNA sequences flanked by loxP sites (Fig.…”

Oncoproteínas Vav: de la regulación del citoesqueleto a funciones fisiológicas y patológicas

“…Sin embargo, su fosforilación tras la estimulación celular hace que estas interacciones intramoleculares se rompan y las proteínas Vav adquieran una conformación "abierta" plenamente activa [3]. Los dominios estructurales y sitios de fosforilación implicados en este proceso han sido descubiertos recientemente [4,5]. Hay que destacar que estas proteínas ejercen pluriempleo a nivel celular, puesto que pueden estimular vías de señalización alternativas independientemente de su actividad enzimática (Figura 1).…”