Abstract LB-364: G(alpha)12 is required for thromboxane A2 to regulate tumor cell motility

Abstract

Abstract Introduction: Thromboxane (TX) A2 is a prostaglandin produced by metabolism of arachidonic acid through cyclooxygenase and thromboxane synthase. TXA2 is biologically active, mainly through activating its cognate, seven transmembrane, G protein coupled receptor. We previously reported that thromboxane A2 receptor (TP) was expressed in prostate cancer, and further activation of this receptor elicited cell contraction and modulated tumor cell motility through regulating small GTPase RhoA (Nie et al., Cancer Res 68: 115–121, 2008). This study aims to identify G alpha protein(s) involved in thromboxane A2 signaling in tumor cell motility. Methods: The expression of G alpha proteins in the PC3MM cells was studied by real-time PCR. Cell contraction assay was performed by staining cells with TRITC phalliodin and evaluated under an epifluorescence microscope. Tumor cell motility and invasion were evaluated using wound healing assay and transwell Boyden Chamber assay. To determine the roles of G alpha protein in thromboxane A2 signaling, we expressed different alleles of G alpha proteins (wild type, constitutive active) in PC3MM cells and then the subsequent effects on cell contractions was determined. We also depleted G alpha protein expression by short hairpin RNA and examined subsequent changes in cell contraction and migration after activation of TP. Results: PC3 MM cell line had the endogenous level of all the G alpha protein (G alpha 12, G alpha i1, G alpha 11, G alpha 13), but not G alpha q. Overexpressionof G alpha 12/13 increased cell contraction after activation of thromboxane A2 receptor with U46619. Expression of constitutively active G alpha 12 by itself was sufficient to cause cell contraction, regardless whether TP is activated or not. Depletion of G alpha 12 via shRNAs reduced cell contraction after TP activation. Tumor cells with G alpha 12 depleted had tumor cell motility and invasiveness. Conclusion: The findings indicate that G alpha 12 is required for TP to induce cell contraction and to regulate tumor cell motility. The studies suggest G alpha 12 may be a promising target of intervention to reduce tumor cell motility and metastasis in prostate cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-364. doi:10.1158/1538-7445.AM2011-LB-364