Posttranscriptional regulation of expression of plasminogen activator inhibitor type-1 by sphingosine 1-phosphate in HepG2 liver cells

Abstract

Altered expression of plasminogen activator inhibitor type-1 (PAI-1), a major physiologic inhibitor of fibrinolysis, is implicated in the progression of atherosclerosis. Sphingosine 1-phosphate (S1P) regulates expression of diverse genes and alters expression of PAI-1 in several types of cells. However, the nature of posttranscriptional regulation of expression of PAI-1 by S1P has not yet been thoroughly elucidated. The present study was undertaken to determine whether S1P has important effects on the posttranscriptional regulation of PAI-1 expression. To evaluate this possibility, we determined promoter activity, mRNA levels, 3′-untranslated region (UTR) activity, and protein levels of PAI-1 in HepG2 cells. S1P increased PAI-1 promoter activity and the expression of PAI-1 mRNA within 4h of exposure. It decreased the expression of PAI-1 mRNA and the accumulation of PAI-1 protein into the media in 24h. Human PAI-1 mRNA exists in two subspecies (3.2 and 2.2kb). S1P decreased the baseline luciferase activity of the 1kb fragment of the 3′ terminus (+2177 to 3176nt) of the 3′-UTR of the 3.2kb PAI-1 mRNA [3′-UTR (+2177–3176)]. S1P decreased expression of PAI-1 protein, presumably by regulating PAI-1 expression at the posttranscriptional level thereby affecting mRNA stability. SERPINE1 mRNA binding protein (SERBP1) and ARE3 in the 3′-UTR were involved in the posttranscriptional regulation by S1P. Our data suggest that S1P can destabilize 3.2kb PAI-1 mRNA through specific effects on the 3′-UTR. These effects appear to involve SERBP1 leading to decreased expression of PAI-1 protein.