Regulation of the expression of the regulatory subunit of cAMP-dependent protein kinase IIβ in Friend erythroleukemic cells. Evidence for posttranscriptional control and a central role for the C subunit

Friend erythroleukemic cells provide a system for studying the regulation of the expression of regulatory (R) and catalytic (C) subunit isoforms of cAMP-dependent protein kinases. Friend cells contain RIα, two RII subunits previously designated RII-52 and RII-54, and Cα. When the cells are treated with 0.2 mM methylisobutylxanthine (MIX) and either 20 μM forskolin or 0.5 mM 8-Br-cAMP, RIα content declines 50-75% because of a large decrease in the t( 1/2 ) value for the dissociated RIα subunit; RII-54 expression is invariant, but the amount and rate of synthesis of RII-52 increases 10-15-fold (Schwartz, D.A., and Rubin, C.S. (1985) J. Biol. Chem. 260, 6296-6303). We now demonstrate that RII-52 and RII-54 correspond to RIIβ and RIIα, respectively. When cAMP levels are elevated in Friend cells the abundance of the 3.3-kilobase RIIβ mRNA increases 25-30-fold in parallel with the rate of RIIβ subunit synthesis indicating that pretranslational control is operative. Other R and C mRNAs are not markedly induced. Surprisingly, the rate of transcriptional initiation of the RIIβ gene and the stability of RIIβ mRNA not altered during RIIβ induction. Rather, the induction of RIIβ mRNA is associated with the accumulation of major (3.4 kilobases) and minor (4 kilobases) RIIβ pre-mRNAs in the nucleus. It appears that the cAMP signal-transduction system alters a nuclear protein(s) such that either the proportion of RIIβ pre-mRNAs that are processed to mature mRNAs and are exported to the cytoplasm is greatly increased or the nuclear precursors are stabilized. Thus, regulation is exerted at a posttranscriptional level. In order to establish directly a causal role for C in RIIβ induction and to rule out artifacts introduced by the use of drugs such as forskolin, MIX, and cAMP analogs we stably transfected Friend cells with a vector containing Vα cDNA under the regulation of the zinc-activated metallothionein I promoter. The addition of 0.15 mM ZnSO₄ caused the accumulation of dissociated C subunits and the selective induction of RIIβ.