Cyclooxygenase-2 gene expression regulation by KSHV G protein coupled receptor vGPCR. Transcriptional and mRNA stability approaches.

M. VICTORIA MEDINA; DAIANA A. SAPOCHNIK; CLARA FONTANA; AGATA M. D'AGOSTINO; JULIAN NAIPAUER; ENRIQUE MESRI; OMAR A. COSO

Bariloche

Simposio; The Fourth South American Symposium in Signal Transduction and Molecular Medicine SISTAM; 2018

Comité organizador de SISTAM 2018

Kaposi?s sarcoma associated herpesvirus (KSHV) vGPCR is a constitutively active G protein-coupled receptor that subverts proliferative and inflammatory signaling pathways to induce cell transformation in Kaposi?s sarcoma. For this reason, identifying pathways leading to vGPCR angiogenesis could have therapeutic significance. Cyclooxygenase-2 (COX-2) is an important inflammatory mediator involved in tumor angiogenesis. We intend to characterize theputative link between signal transduction pathways triggered by vGPCR,COX-2 promoter activation and it´s mRNA stability. To achieve our goal we worked with SVEC cells and SVEC cells that constitutively express the vGPCR (SVEC vGPCR). We observed by Real Time PCR and by Western Blot that SVEC vGPCR cells overexpress COX-2 mRNA and protein respect to control cells. We transfected SVEC cells with a plasmid containing the COX-2 promoter followed by a luciferase open reading frame and co-transfected with plasmids which express several MEK kinases. We observed an increase in luciferase expression when co-expresing MEKEE. We also worked with a plasmid that expresses luciferase fused to the 3?-UTR of COX-2 mRNA together with different AUBPs expression vectors and observed an increase in luciferase expression when coexpressing TTP. We also observed that treatment with the COX-2 selective inhibitor y drug Celecoxib produced a significant retardation in tumor growth when injected cells that express vGPCR in mice. Using an intradermal angiogenesis assay, we found that treatment with NS398 before inoculation with vGPCR-transformed cells abolished the angiogenic response induced by vGPCR expression. Based on these results we conclude that vGPCR upregulates COX-2 levels in endothelial cells due to a dual effect upon it´s promoter region and upon elements in the 3?UTR region and that vGPCR regulates angiogenicity and tumorigenicity via COX-2 activation. These facts pinpoint COX-2 as potential target for KS chemoprevention and therapy.