Chemically induced murine erythroleukemia cell differentiation is severely impaired when cAMP-dependent protein kinase activity is repressed by transfected genes.

Abstract

During chemically induced differentiation of murine erythroleukemia (MEL) cells, cAMP-dependent protein kinase activity increases, and the enzyme's isozyme pattern changes. To examine the enzyme's role during MEL cell differentiation, we stably transfected MEL cells with recombinant plasmids in which the mouse metallothionein I promoter controlled expression of either a mutant form of the type I regulatory subunit of cAMP-dependent protein kinase (RI) or the enzyme's specific peptide inhibitor (PKI); expressing either sequence rendered cells cAMP-dependent protein kinase-deficient. Chemically induced differentiation of MEL cells as assessed by beta-globin mRNA and hemoglobin accumulation was inhibited in RI mutant and PKI transfectants; adding zinc further inhibited differentiation in the transfectants but had no effect on parental MEL cells. The inhibition of differentiation correlated with the amount of RI mutant mRNA and protein in the RI mutant transfectants and with the cells' degree of cAMP-dependent protein kinase deficiency in both the RI mutant and PKI transfectants. Overexpression of wild type RI did not interfere with differentiation or enzyme activity. We conclude that cAMP-dependent protein kinase activity is important for chemically induced differentiation of MEL cells and that the down-regulation of RI protein which occurs during MEL cell differentiation is not essential for differentiation to proceed.