G2A is an oncogenic G protein-coupled receptor

Abstract

G2A is a heptahelical cell surface protein that has recently been described as a potential tumor suppressor, based on its ability to counteract transformation of pre-B cells and ®broblasts by Bcr-Abl, an oncogenic tyrosine kinase. We have isolated cDNAs encoding G2A in the course of screening libraries for clones that cause oncogenic transformation of NIH3T3 ®broblasts. When expressed at high levels in NIH3T3 cells by retroviral transduction, G2A induced a full range of phenotypes characteristic of oncogenic transformation, including loss of contact inhibition, anchorage-independent survival and proliferation, reduced dependence on serum, and tumorigenicity in mice. When expressed by transfection, G2A greatly enhanced the ability of a weakly oncogenic form of Raf-1 to transform NIH3T3 cells. These results demonstrate that G2A is potently oncogenic both on its own and in cooperation with another oncogene. Expression of G2A in ®broblasts and endothelial cells resulted in changes in cell morphology and cytoskeleton structure that were equivalent to those induced by the G protein subunit Ga 13 . Transformation of NIH3T3 cells via G2A expression was completely suppressed by co-expression of LscRGS, a GTPase activating protein that suppresses signaling by Ga 12 and Ga 13 . Hyperactivity of Ga 12 or Ga 13 has previously been shown to result in activation of Rho GTPases. G2A expression resulted in activation of Rho, and transformation via G2A was suppressed by a dominant negative form of RhoA. These results indicate that G2A may be directly coupled to Ga 13 , and that it is the activation of this Rho-activating Ga protein which is responsible for the ability of G2A to transform ®bro-blasts.

The thrombin receptor, PAR-1, causes transformation by activation of Rho-mediated signaling pathways

“…Although the transforming activity of each of these receptors requires Rho function, the particular substrate that is activated by the receptors can di er. Mas transformation occurs in a Racl dependent manner (Zohn et al, 1998a), while G2A transformation is mediated by RhoA (Zohn et al, 2000). In the case of PAR-1, the identi®cation of a substrate that can mediate its transforming activity in native receptor expressing cells seems particularly important.…”

“…For these analyses, we utilized an expression vector that encoded the isolated RGS domain of Lsc, which has been shown to be a GAP for Ga 12 , and Ga 13 . Expression of LscRGS speci®-cally blocks transformation mediated by Ga 12 or Ga 13 , and also blocks transformation via G2A, a GPCR that causes transformation by activation of RhoA (Zohn et al, 2000). Co-expression of Lsc RGS strongly suppressed focus formation by both mouse and human PAR-1 (Figure 7b).…”

“…The human PAR-1 cDNA was provided by Ellen Van Obberghen-Schilling (Centre de Biochimie, CNRS, France). The LscRGS construct has been described previously (Zohn et al, 2000). It was inserted into a pCTV derivative vector and stably expressed by infection.…”

“…Furthermore there is increasing evidence that some GPCRs harbor oncogenic potential. For example, the mas and g2a oncogenes, both of which encode GPCRs, were isolated based on their transforming activity (Young et al, 1988;Zohn et al, 2000). The serotonin 5HT1b receptor (Julius et al, 1989) and the M 1 , M 3 and M 5 subtypes of the acetyl cholinergic receptors (Gutkind et al, 1991) were found to cause agonistdependent transformation, and constitutively-activated mutants of the a1B-adrenergic receptor have been shown to increase focus formation in NIH3T3 cell assays (Allen et al, 1991).…”

RGS-containing RhoGEFs: the missing link between transforming G proteins and Rho?

“…In this regard, p115-RhoGEF, PDZ-RhoGEF and LARG are themselves focus-forming when overexpressed in NIH3T3 cells (Figure 2), and a recent report indicates that the transformation of these cells induced by G2A, an oncogenic GPCR, can be suppressed by co-expression of the RGS domain of Lsc, the murine homolog of p115 RhoGEF (Zohn et al, 2000). Thus, given the importance of Rho in cell growth regulation, it is possible to speculate that the activation of RGS-containing RhoGEFs may represent a key component of the mitogenic and transforming pathway utilized by many GPCRs.…”