Inducible expression of a constitutively active mutant of mitogen-activated protein kinase kinase 7 specifically activates c-JUN NH2-terminal protein kinase, alters expression of at least nine genes, and inhibits cell proliferation

MKK7 is a recently discovered mitogen-activated protein kinase (MAPK) kinase that is unique in that it specifically activates only the c-JUN NH2-terminal protein kinase (JNK) family of enzymes. Very little is known about the biological role of MKK7. We generated inducible cell lines from the human embryonal kidney carcinoma cell line, HEK293, by stable transfection with a constitutively active mutant of MKK7, MKK7(3E), fused to green fluorescent protein (GFP), under the control of an ecdysone-inducible promoter. Treatment of cells with the synthetic ecdysone analog ponasterone A induced expression of GFP-MKK7(3E) and resulted in sustained activation of endogenous JNK, but neither of the other endogenous MAPKs, ERK or p38. Red and green fluorescing cDNA copies of mRNA extracted from cells obtained before and after induction of GFP-MKK7(3E) were hybridized to microarrays containing more than 6,000 cDNAs in eight independent experiments. By selection criteria, 23 genes were differentially regulated after 24 h of induction of GFP-MKK7(3E) and 16 after 48 h. The expression of 9 genes was consistently changed after both 24 and 48 h of induction. These changes included down-regulation of three genes, c-myc, angiopoietin-2, and glucose-regulated protein 58, and up-regulation of 6 genes, tissue factor pathway inhibitor-2, GRP78, autotaxin, PPP1R7, the DKFZ cDNA p434D0818, and I unknown gene. Consistent with previously described roles of several of the altered genes, MKK7(3E) inhibited cell proliferation. These data implicate active MKK7 in the negative regulation of cell proliferation and provide evidence for a new role for this kinase in the regulation of a distinct, hitherto unrecognized set of genes.