Role of regulatory subunits and protein kinase inhibitor (PKI) in determining nuclear localization and activity of the catalytic subunit of protein kinase A

Regulation of protein kinase A by subcellular localization may be critical to target catalytic subunits to specific substrates. We employed epitope-ta gged catalytic subunit to correlate subcellular localization and gene-induc ing activity in the presence of regulatory subunit or protein kinase inhibi tor (PKI). Transiently expressed catalytic subunit distributed throughout t he cell and induced gene expression. Go-expression of regulatory subunit or PKI blocked gene induction and prevented nuclear accumulation. A mutant PK I lacking the nuclear export signal blocked gene induction but not nuclear accumulation, demonstrating that nuclear export is not essential to inhibit gene induction. When the catalytic subunit was targeted to the nucleus wit h a nuclear localization signal, it was not sequestered in the cytoplasm by regulatory subunit, although its activity was completely inhibited. PKI re distributed the nuclear catalytic subunit to the cytoplasm and blocked gene induction, demonstrating that the nuclear export signal of PKI can overrid e a strong nuclear localization signal. With increasing PKI, the export pro cess appeared to saturate, resulting in the return of catalytic subunit to the nucleus. These results demonstrate that both the regulatory subunit and PKI are able to completely inhibit the gene-inducing activity of the catal ytic subunit even when the catalytic subunit is forced to concentrate in th e nuclear compartment.