The tumor microenvironment has a characteristic low pH due to inadequate blood perfusion, hypoxia, altered cellular metabolism, and inflammation. The effects of acidic pH on cancer progression are complex. It has been shown that low pH increases the metastatic spread of cancer by assisting in the degradation of extracellular matrix. Other studies show that acidic microenvironments decrease metastatic potential in B16F10 melanoma cells upon activation of the GPR4 receptor and also inhibit Akt activity in breast cancer cells. In this study we have investigated the attachment, spreading, and migration of B16F10 cells that have been genetically modified to express the GPR4 receptor at a high level (named B16F10/GPR4 cells). The results showed that upon stimulation of GPR4 in B16F10/GPR4 cells with low pH, cell spreading, membrane ruffling, and migration were significantly decreased. The phosphorylation levels of paxillin Y118 and focal adhesion kinase (FAK) Y397 following one-hour cell attachment were also decreased, and the spatial localization of the phosphorylated paxillin and FAK was substantially altered. Further tests were performed to identify the G-protein signaling pathway responsible for this effect. By using a G13 dominant negative construct or inhibiting Rho activation with C3 transferase (CT04) in B16F10/GPR4 cells, the cell spreading and membrane ruffling abilities were almost completely restored back to the level of B16F10/vector control cells, whereas the inhibition of the Gs and Gq pathways had little effect. These results suggest that activation of the GPR4 receptor and subsequently the G13/Rho downstream pathway are responsible for decreased cell attachment, spreading and membrane ruffling, which may cause decreased metastatic potential of B16F10/GPR4 cells found previously.

Citation Format: Calvin R. Justus, Li V. Yang. Regulation of tumor cell attachment, spreading and migration by the GPR4 receptor and related G protein pathways. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2799. doi:10.1158/1538-7445.AM2013-2799