ACTIVATION OF CAMP-DEPENDENT PROTEIN-KINASE ALTERS THE CHROMATIN STRUCTURE OF THE UROKINASE-TYPE PLASMINOGEN-ACTIVATOR GENE PROMOTER

In LLC-PK1 cells, the urokinase-type plasminogen activator (uPA) gene is induced by two of the major signal transduction pathways, the prote in kinase C (PKC) and the cAMP-dependent protein kinase (PKA) pathways . We have analyzed the chromatin structure of 26 kb of the uPA gene lo cus and have shown that PKA activation but not PKC activation induces major chromatin structural alterations in the uPA gene promoter. In un induced cells, several DNase I hypersensitive (HS) sites were detected in the 5' and 3' flanking regions but not in the transcribed region, Two of the sites correspond to previously characterized regulatory sit es: a cAMP responsive site at nucleotide position -3500 with respect t o the initiation site, and the PEA3/AP1 site at -2100 that mediates PK C activation. After the activation of PKA but not PKC, a strong HS sit e was induced at -2600. Functional analysis of this region revealed cA MP responsive activity. Chromatin structural alterations again brought about specifically by PKA but not by PKC were were also detected in t he upstream of the promoter by topoisomerase I cleavage site analysis, with two prominent sites appearing at -2800 and -3300. These results suggest that the strong cAMP induction of the uPA gene requires struct ural alterations that permit cooperative interactions between the mult iple cAMP responsive sites.