CB-17CHARACTERIZATION OF HCMV-ECODED CHEMOKINE RECEPTOR US28 TRANSDUCED MSU1.1 HUMAN FIBROBLAST CELLS

The Human Cytomegalovirus (HCMV) encodes a G-protein coupled chemokine receptor called US28. Using a mouse model system, US28 was previously found to be oncomodulatory, increasing proliferation, inducing anchorage independent growth and loss of contact inhibition. Similarly, in the human glioblastoma cell line U373, US28 activated VEGF gene expression. In order to determine if US28 is oncomodulatory in a normal human cell, we engineered the human fibroblast cell line MSU1.1 to constitutively express US28 via lentivirus infection. MSU1.1 cells were transduced with pHAGE-US28-ZsGreen. Fluorescent confocal microscopy was utilized to detect the ZsGreen protein tag and confirmed the successful transduction of MSU1.1 cells with the US28 vector or empty pHAGE vector. MSU1.1 cells expressing US28 did not exhibit loss of contact inhibition nor anchorage independent growth in focus formation and soft agar assays, respectively. Using the AlamarBlue assay, MSU1.1 cells expressing US28 showed a 2.303 fold increase in proliferation compared to wild type MSU1.1 cells and a 2.201 fold increase in proliferation compared to mock transduced cells. US28 expression in the immortalized human fibroblast cell line MSU1.1 increased proliferation but did not trigger a transformed phenotype. This study reveals potential cell-type and species-specific differences from previous findings assessing oncogenic and oncomodulatory properties of US28 expression in mammalian cell lines such as NIH-3T3 (mouse fibroblast), and U373 (malignant glioma). Although ectopic expression of US28 does not appear to drive transformation, the reported prevalence in GBM and confirmation of oncomodulation via increased proliferation in MSU1.1 cells makes US28 a potential target warranting further understanding of the virus' role in GBM.