Gβγ Signaling Promotes Breast Cancer Cell Migration and Invasion

Abstract

Signaling through G protein-coupled receptors (GPCRs) promotes breast cancer metastasis. G proteins convey GPCR signals by dissociating into G␣ and G␤␥ subunits. The aim of the present study was to determine whether blockade of G␤␥ signaling suppresses breast cancer cell migration and invasion, which are critical components of metastasis. Conditioned media (CM) of NIH-3T3 fibroblasts are widely used as chemoattractants in in vitro cancer metastasis studies. Expression of a G␤␥ scavenger peptide attenuated NIH-3T3 CM-induced migration and invasion of both metastatic breast cancer MDA-MB-231 and MDA-MB-436 cells by 40 to 50% without effects on cell viability. Migration and invasion of cells in response to NIH-3T3 CM were also blocked by 8-(4,5,6-trihydroxy-3-oxo-3H-xanthen-9-yl)-1-naph-thalene-carboxylic acid) (M119K), a G␤␥ inhibitor, with maximum inhibition exceeding 80% and half-maximal inhibitory concentration (IC 50 ) values of 1 to 2 M.M119K also attenuated Rac-dependent formation of lamellipodia, a key structure required for metastasis. Constitutively active Rac1 rescued G␤␥ blockade-mediated inhibition of breast cancer cell migration, whereas dominant negative Rac1 inhibited cell migration similar to G␤␥ blockade. Furthermore, M119K suppressed G i protein-coupled CXC chemokine receptor 4 (CXCR4)-dependent MDA-MB-231 cell migration by 80% with an IC 50 value of 1 M, whereas tyrosine kinase receptor-dependent cell migration was significantly less inhibited. However, CXCR4-dependent inhibition of adenylyl cyclase, a G i ␣-mediated response in MDA-MB-231 cells, was not blocked by M119K but was blocked by pertussis toxin, which selectively inactivates G i ␣. This report is the first to directly demonstrate the role of G␤␥ in cancer cell migration and invasion and suggests that targeting G␤␥ signaling pathways may provide a novel strategy for suppressing breast cancer metastasis.Breast cancer is the second largest killer of women by cancer with approximately 40,000 deaths in the United States each year (Jemal et al., 2008). Patients who cannot be cured are primarily those in whom breast cancer has metastasized. Metastasis is a complex pathophysiological process, beginning with the migration and invasion of cancer cells into the surrounding tissues and lymphatics. From these sites, cancer cells can gain access to the circulation and proliferate to form new colonies at metastatic sites in vital organs such as the lung, bone, liver, and brain (StetlerStevenson and Kleiner, 2001). This directed cell migration and invasion is triggered by chemoattractants such as chemokines, growth factors, and matrix metalloproteases, which are sensed by cancer cell surface receptors including integrins, receptor tyrosine kinases, and G protein-coupled receptors (GPCRs). Evidence from preclinical and clinical studies shows excessive activation of GPCRs in breast cancer caused by overexpression of receptors, abnormally elevated ligands for GPCRs, and/or down-regulation of RGS proteins (Dorsam and Gutkind, 2007;Xie et al., 200...

NO triggers RGS4 degradation to coordinate angiogenesis and cardiomyocyte growth

“…Loss of RGS4 in cancer cells results in aberrant Gβγ-induced Rac activation that is rescued by proteasomal inhibitor or Gβγ scavenger peptide (53,54). Although the causes of RGS4 degradation in advanced breast cancer remain unknown, an interesting observation is that increased angiogenesis and NO production occur at about the same time (55).…”

The Expanding Roles of GβγSubunits in G Protein–Coupled Receptor Signaling and Drug Action

“…Alterations in Gbg subunit expression and signaling have also been implicated in the progression of various types of cancer (Kirui et al, 2010;Yajima et al, 2012), and much work has been done to determine the benefits of interfering with Gbg signaling in cancer growth. GRK2-ct-mediated inhibition of Gbg signaling in a human prostate cancer cell line, PC3, reduced cancer cell proliferation, and impeded formation of prostate tumors in a mouse model (Bookout et al, 2003).…”

Inverted Formin 2 Regulates Actin Dynamics by Antagonizing Rho/Diaphanous-related Formin Signaling

“…These INF2-deficient cells also exhibited elongated morphology. We quantified the fraction of cells containing lamellipodia by measuring the relative area of cortical actin meshwork and the relative length of plasma membrane enriched with cortactin 25 (Figure 1), a protein that is highly associated with the branched lamellipodial actin network. We observed a statistically significant decrease in the relative size of the lamellipodia in cells with INF2 depletion.…”

“…25 Following the standard quantification method described previously, 25 images were captured with a Cool-SNAPCF camera attached to a Nikon (Tokyo, Japan) Eclipse 80i microscope. Podocytes immunostained for cortactin and F-actin were used for the quantification of lamellipodia.…”