Human Cytomegalovirus Infection of ARPE–19 Cells Dampens Cellular Transcription Factor AP–1 Activity

Abstract

Abstract: : Purpose:We have shown previously that the cellular transcription factor NF–ΚB is downregulated in human retinal pigment epithelial (RPE) cells during human cytomegalovirus (HCMV) replication. Since additional transcription factors such as AP–1 and Sp1 may also be influenced by HCMV gene expression, we performed studies to test the hypothesis that HCMV infection of RPE cells results in downregulation of AP–1 in a fashion similar to that observed for NF–ΚB. Methods:Monolayers of ARPE–19 cells were transfected with AP–1 luciferase DNA, inoculated with HCMV [Towne] (moi = 1) at 24 hrs post–transfection, harvested at 4 hr postinfection by lysis with a reporter buffer, and quantified for luciferase activity as a measure of cellular AP–1 activity. Uninfected ARPE–19 cells transfected with AP–1 luciferase DNA served as controls. Immunoblot assays were performed on HCMV–infected and uninfected ARPE–19 cell extracts to detect AP–1 directly. In addition, cellular expression of c–Fos and c–Jun was determined by chemiluminescence and autoradiography. Results:HCMV–infected ARPE–19 cells transfected with AP–1 luciferase DNA exhibited a 2–fold decrease in promoter activity when compared with transfected uninfected ARPE–19 cells. Both c–Fos and c–Jun were detected in HCMV–infected and uninfected cells, but no significant difference in their expression was apparent. Conclusions: HCMV infection of ARPE–19 cells results in dampening of AP–1 activity, although the dampening was not as great as for NF–ΚB. This finding supports our suspicion that HCMV infection of RPE cells causes a global downregulation of all cellular transcription factors. This effect might explain the unusual virus/host interaction between HCMV and RPE. Work is in progress to assess the fate of Sp1 during HCMV infection.

Keywords: cytomegalovirus • retinal pigment epithelium • gene/expression