COMPLEX REGULATION OF PLASMINOGEN-ACTIVATOR INHIBITOR TYPE-1 (PAI-1) GENE-EXPRESSION BY SERUM AND SUBSTRATE ADHESION

Expression of plasminogen activator inhibitor type-1 (PAI-1), a member of the serine protease inhibitor (SERPIN) superfamily that functions to negatively regulate the plasmin-based pericellular proteolytic casc ade, was induced early after exposure of growth-arrested normal rat ki dney (NRK) cells to serum-containing medium. Increased PAI-1 transcrip tion was rapid (evident within 10 min of serum addition) and involved immediate-early response kinetics. [H-3]Thymidine autoradiography was used to map the time frame of PAI-1 expression during a synchronous gr owth cycle. PAI-1 transcript accumulation peaked in mid-G(1) phase (ap prox. 4-6 h post-stimulation) and declined prior to, or concomitant wi th, the onset of DNA synthetic phase. Serum increased PAI-1 expression in NRK cells in agarose suspension, as well as monolayer, culture; in duction in suspended cells (similar to monolayer culture conditions) a lso occurred in the presence of cyclohexamide or puromycin. The serum- inductive pathway leading to PAI-1 gene activation is thus functional regardless of adhesive conditions or capacity for de novo protein synt hesis. The amplitude of induction and maintenance of expression in lat er stages of G(1), however, were subject to adhesive influences. PAI-1 transcript accumulation at 4 and 8 h poststimulation in newly adheren t cells, moreover, was blocked by puromycin, indicating that both imme diate-early and secondary mechanisms regulate PAI-1 mRNA levels during progression of NRK cells through an 'activated' G(1) growth phase.