P-Rex1 Links Mammalian Target of Rapamycin Signaling to Rac Activation and Cell Migration

Abstract

Polarized cell migration results from the transduction of extracellular cues promoting the activation of Rho GTPases with the intervention of multidomain proteins, including guanine exchange factors. P-Rex1 and P-Rex2 are Rac GEFs connecting G␤␥ and phosphatidylinositol 3-kinase signaling to Rac activation. Their complex architecture suggests their regulation by protein-protein interactions. Novel mechanisms of activation of Rho GTPases are associated with mammalian target of rapamycin (mTOR), a serine/ threonine kinase known as a central regulator of cell growth and proliferation. Recently, two independent multiprotein complexes containing mTOR have been described. mTORC1 links to the classical rapamycin-sensitive pathways relevant for protein synthesis; mTORC2 links to the activation of Rho GTPases and cytoskeletal events via undefined mechanisms. Here we demonstrate that P-Rex1 and P-Rex2 establish, through their tandem DEP domains, interactions with mTOR, suggesting their potential as effectors in the signaling of mTOR to Rac activation and cell migration. This possibility was consistent with the effect of dominant-negative constructs and short hairpin RNA-mediated knockdown of P-Rex1, which decreased mTOR-dependent leucine-induced activation of Rac and cell migration. Rapamycin, a widely used inhibitor of mTOR signaling, did not inhibit Rac activity and cell migration induced by leucine, indicating that P-Rex1, which we found associated to both mTOR complexes, is only active when in the mTORC2 complex. mTORC2 has been described as the catalytic complex that phosphorylates AKT/PKB at Ser-473 and elicits activation of Rho GTPases and cytoskeletal reorganization. Thus, P-Rex1 links mTOR signaling to Rac activation and cell migration.P-Rex1 and P-Rex2 are Rac guanine exchange factors connecting G protein-coupled receptors, through G␤␥ and phosphatidylinositol 3-kinase, to Rac activation. In particular, P-Rex1 has been associated with the activation of Rac2, generating reactive oxygen species in neutrophils. P-Rex2 (showing two splice variants) is similarly regulated by G␤␥ and phosphatidylinositol 3-kinase. Northern blot assays revealed a differential distribution of the two members of the P-Rex 3 family, suggesting that they exert equivalent functions in different cellular populations (1-3, 7-11). The complex architecture of this family of proteins, constituted by a catalytic DH domain, followed by a phosphatidylinositol 3,4,5-trisphosphate-sensitive pleckstrin homology domain, two DEP and two PDZ domains in tandem, and a long carboxyl terminus (except for P-Rex2b, which is the short version, having a reduced carboxyl terminus), suggests that these Rac guanine exchange factors might be regulated by diverse protein-protein interactions modulating signal transduction pathways associated with the activation of Rac. In fact, in the developing brain, P-Rex1 is associated with neuronal migration in response to nerve growth factor (12, 13).The mammalian target of rapamycin, mTOR, a highly conserved serine-threonine k...

MHC class I and integrin ligation induce ERK activation via an mTORC2-dependent pathway

“…mTORC1 phosphorylates p70 S6 Kinase and eukaryotic initiation factor 4E binding protein 1 which activate translational machinery to increase cell proliferation [6,7]. mTORC2 is composed of mTOR binding to rapamycin insensitive companion of mTOR (Rictor), GβL, stress-activated protein kinase-interacting protein 1 (Sin1) and the newly discovered associations with PRoline-Rich protein 5 (PRR5) or the synonymous protein observed with Rictor 1 (Protor-1), and P-REX1 [8][9][10][11][12]. mTORC2 activates Akt at Ser473, Rac GTPase, cell migration and effects reorganization of the actin cytoskeleton [11][12][13][14].…”

“…mTORC2 is composed of mTOR binding to rapamycin insensitive companion of mTOR (Rictor), GβL, stress-activated protein kinase-interacting protein 1 (Sin1) and the newly discovered associations with PRoline-Rich protein 5 (PRR5) or the synonymous protein observed with Rictor 1 (Protor-1), and P-REX1 [8][9][10][11][12]. mTORC2 activates Akt at Ser473, Rac GTPase, cell migration and effects reorganization of the actin cytoskeleton [11][12][13][14]. Recent work utilizing mouse knockout models emphasizes the importance of mTOR for growth and proliferation in development [15,16].…”

T cell dependence on mTOR signaling

“…Recently, the Rac GEFs P-Rex 1 and 2 have been shown to associate with either mTORC1 or mTORC2 and participate in Rac-mediated migration of fibroblasts and tumorderived cell lines. 26 Short-term rapamycin treatment of particular tumor cell lines did not inhibit migration, indicating P-Rex association with the less sensitive mTORC2 complex is likely important for migratory responses. 26 However, the cell lines examined in this study have been previously shown to maintain mTORC2 activity after prolonged rapamycin treatment, 8 rendering them more rapamycinresistant than most T cell subsets.…”

“…26 Short-term rapamycin treatment of particular tumor cell lines did not inhibit migration, indicating P-Rex association with the less sensitive mTORC2 complex is likely important for migratory responses. 26 However, the cell lines examined in this study have been previously shown to maintain mTORC2 activity after prolonged rapamycin treatment, 8 rendering them more rapamycinresistant than most T cell subsets. It may be of interest to determine whether P-Rex 1 and 2 are expressed in T cells and whether they associate with mTORC1 or mTORC2 to regulate starvation-mediated migration.…”

mTOR Signaling in Lymphangioleiomyomatosis

“…132,133 mTORC2 directly interacted with and regulated Prex1, a Rac1 guanine nucleotide exchange factor that controls cytoskeletal rearrangement. 134 Although this discussion has been restricted to direct mTOR signaling interactions (Fig. 3), certain indirect effects of mTOR on biological processes relevant to LAM are worthy of mention.…”