Effects of variants of the Epstein-Barr Virus LMP1 protein in the regulation Epithelial-Mesenchymal transition in human epitelial cells in vitro

Abstract

It is estimated that 15% all cases cancer diagnosticated are related to the infection with infectious agents viruses, including viruses. The Epstein Barr Virus - EBV is the herpevirus gama discovered in 1964, being the first carcinogenic virus describe for human, The contagious occurs by saliva and 90% of adult worldwide presents of latent infection, that is etiologic related with the development of Infectious Mononucleosis, Endemic Burkitt Lymphoma and undifferentiated carcinoma associated of nasopharyngeal, among others diseases. The main EBV oncogenic product is the latent protein of membrane 1 (LMP1), that interfere in several intracellular signaling pathway, favoring the immortalization and cell proliferation, which occasionally can culminates in cancer. The present study aims to analyze the properties of the LMP1 variants derivate of the viruses genotype B95.8 and M81 into regulation Epithelial-Mesenchymal program (EMT), crucial phenomenon in the progression carcinoma. For this, will be generated model in vitro of epithelial cells expressing the variants B95.8 and M81 of LMP1 with finally of discriminate the LMP1 effects into regulation EMT and in intracellular signaling pathways relevant to cell migration and invasion. Will be employed recombinant DNA techniques to the production of vectors for expression of B95.8 and M81 of LMP1, which will be used to transfect SW480 (human colorectal adenocarcinoma) and NP69SV40T (Nasopharyngeal epithelial cell) cells. The cells expressing LMP1 will submitted the total RNA extraction, what will be converted in cDNA and analyzed, through polymerase chain reaction quantitative in time real - qPCR, as the expression in transcriptional level of selected genes that encodes transcription inductors factors of EMT( Snail, Slug, Twist and ZEB1), markers of epithelial phenotype (E-Cadherin, ZO-1) and mesenchymal (N-Cadherin, vimentin, ±-SMA, fibronectin, metalloproteinases 2 and 9) and intracellular proteins signaling pathway involved into EMT regulation (PI3K p85a, AKT, PTEN, FAK, Src, IºB±, NFºB p65, e STAT3). To depend on results of the differential expression obtained, the expression. Of the selected genes will be in protein level for the specifics targets using western blotting. In the experiments final will wait to find data's about possible different in the effects of the B95.8 and M81 variants involved in the EMT and, in components the intracellular signaling pathway related this phenomenon that has an important role in the biology aggressiveness carcinoma.