Breast Cancer Migration and Invasion Depend on Proteasome Degradation of Regulator of G-Protein Signaling 4

Abstract

Aberrant signaling through G-protein coupled receptors promotes metastasis, the major cause of breast cancer death. We identified regulator of G-protein signaling 4 (RGS4) as a novel suppressor of breast cancer migration and invasion, important steps of metastatic cascades. By blocking signals initiated through G i -coupled receptors, such as proteaseactivated receptor 1 and CXC chemokine receptor 4, RGS4 disrupted Rac1-dependent lamellipodia formation, a key step involved in cancer migration and invasion. RGS4 has GTPaseactivating protein (GAP) activity, which inhibits G-protein coupled receptor signaling by deactivating G-proteins. An RGS4 GAP-deficient mutant failed to inhibit migration and invasion of breast cancer cells in both in vitro assays and a mouse xenograft model. Interestingly, both established breast cancer cell lines and human breast cancer specimens showed that the highest levels of RGS4 protein were expressed in normal breast epithelia and that RGS4 down-regulation by proteasome degradation is an index of breast cancer invasiveness. Proteasome blockade increased endogenous RGS4 protein to levels that markedly inhibit breast cancer cell migration and invasion, which was reversed by an RGS4-targeted short hairpin RNA. Our findings point to the existence of a mechanism for posttranslational regulation of RGS4 function, which may have important implications for the acquisition of a metastatic phenotype by breast cancer cells. Preventing degradation of RGS4 protein should attenuate aberrant signal inputs from multiple G i -coupled receptors, thereby retarding the spread of breast cancer cells and making them targets for surgery, radiation, and immune treatment.

Pristimerin Inhibits Breast Cancer Cell Migration by Up-regulating Regulator of G Protein Signaling 4 Expression

“…RGS4 can selectively inhibit the Gi-and Gq-coupled GPCRs signaling (Huang et al, 1997;Xie GX et al, 2007), and modulation of RGS4 levels can regulate cancer cell metastasis (Xie et al, 2009). We found that treatment with pristimerin or the proteasome inhibitor of MG132 increased the levels of intra-plasmatic RGS4.…”

“…The effects of pristimerin on the migration and invasion of MDA-MB-231 cells in vitro were determined by transwell migration assay, as described previously (Xie et al, 2009). Briefly, MDA-MB-231 cells (5 × 10 4 / well) in the upper chambers were treated in triplicate with 0.25-1 μM pristimerin or 10 μM MG132.…”

“…Six week-old female BALB/c nude mice were obtained from the National Rodent Laboratory Animal Resources, Shanghai, China, and housed in a specific pathogen-free facility in a 12-h light-dark cycle in China Pharmaceutical University. The orthotopic mammary fat pad model of breast cancer was established, as previously described (Xie et al, 2009). Individual BALB/c nude mice were inoculated with 8×106 MDA-MB-231 cells in 100 μL Matrigel into a mammary fat pad.…”

“…A previous study has shown that inhibition of proteaosme activity is associated with the inhibition of MDA-MB-231cell migration and invasion (Xie et al, 2009). We tested whether treatment with pristimerin could modulate the migration and invasion of breast cancer cells by transwell assays in vitro.…”

“…Previous study has shown that RGS4 is a critical negative regulator of breast cancer cell migration and invasion (Xie et al, 2009). RGS4 is predominantly degraded by proteasome and the N-end rule pathway (Lee et al, 2005;Bodenstein et al, 2007) and pristimerin inhibits proteasome activity (Tiedemann et al, 2009).…”