PKC-β and MARK are required for the synthesis of PAI-1 during the phorbol estermediated adhesion of the human cell line HL-60

PAI-I is recognized to regulate cell adhesion. The identification of the signal transduction pathway triggering PAI-1 synthesis should help to elucidate its role during adhesion process. Phorbol ester-stimulated HL-60 promyelocytic cell line were used as a model of cells developing the adherent phenotype. The PMA-activated synthesis of PAI1 was shown to be dependent of protein kinase C (PKC), tyrosine kinase (TK) and mitogen activated protein kinase (MARK) p42/p44 pathway, since Ro 31 8220, herbimycin and PD 098059, respectively, abolished PAI-1 synthesis in a synchronous manner and also prevented adhesion to normally develop. SB 203580, a specific inhibitor of the MAPK p38/HOG has no effect on PMAinduced PAI-1 synthesis and cell adhesion. Taking advantage of the PKC-β-deficient HL-525 cell line, a clone issued from HL-60 rendered resistant to PMA, we were able, either by a pretreatment with retinoic acid that reinduces PKC-β or transfection with PKC-β gene, to identify PKC-β as the isoform being involved in PMA-induced PAI-1 synthesis. This study was alsoextented to uPAR and uPA. In HL-60, uPAR surface expression was drastically enhanced by PM A, but was partly inhibited (50 to 60 %) by either Ro 31 -8220 or PD098059, and strongly inhibited by combination of both. In HL-525, uPAR surface expression was significantly enhanced after PMA treatment and partially inhibited by Ro 31 -8220 suggesting the involvement of a PKC isoform different to PKC-β. PMA did not activate uPA synthesis in HL-60. These results allow us to propose the colinear pathway PKC-β-MAPKKMAPK p42/p44 as a potential route for PAI-1 synthesis during PMA-induced adherence of HL-60, whereas uPAR synthesis appears mediated by two additive transduction pathways one involving a PKC and the other the MAPK p42/p44. The parallelism we observed between adherence and PAI-1 synthesis in uPAR-expressing cells underlines the role of this inhibitor in uPARmediated cell adhesion.