Regulation of EPC-1/PEDF in normal human fibroblasts is posttranscriptional.

Abstract

The EPC-1 (early population doubling level cDNA-1) gene, also known as pigment epithelium-derived factor, encodes a protein belonging to the serine protease inhibitor (serpin) superfamily that has been reported to inhibit angiogenesis and proliferation of several cell types. We have previously reported that the EPC-1 mRNA and the secreted EPC-1 protein are expressed at levels more than 100-fold higher in early passage, G(0), WI-38 cells compared to either proliferating or senescent WI-38 fibroblasts. To examine the molecular mechanisms that regulate changes in EPC-1 gene expression in WI-38 cells, we isolated and characterized the human EPC-1 gene and determined the mRNA cap site. Transcriptional assays showed no change in the transcription rates of EPC-1 between young proliferating, quiescent, and senescent WI-38 cells. These results suggest posttranscriptional regulation of the EPC-1 gene. Reverse transcriptase polymerase chain reaction measurements (of hnRNA) indicate regulation at the hnRNA level. The regulation of the EPC-1 gene at the level of hnRNA can explain the observed slow increase in the steady-state EPC-1 mRNA levels when cells become quiescent. The reduction of EPC-1 mRNA levels that occurs when cells exit G(0) and are induced to proliferate can be accounted for by a reduction of the EPC-1 mRNA stability in stimulated cells as compared to quiescent cells.