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An engineered site for protein kinase C flanking the SV40 large T-antigen NLS confers phorbol ester-inducible nuclear
1Division of Biochemistry and Molecular Biology, John Curtin School of Medical Research, Australian National University, Canberra, ACT.
Abstract:
Nuclear import of simian virus SV40 large tumour antigen (T-ag) is enhanced by the protein kinase CK2 (CK2) site flanking the nuclear localisation sequence (NLS). We report here that replacement of this site with a consensus site for protein kinase C (PK-C) can alter the regulation of T-ag nuclear import and render it inducible by phorbol ester. Measurement of nuclear import kinetics using fluorescently labelled proteins and confocal laser scanning microscopy show that the introduced PK-C site is functional in enhancing T-ag nuclear import compared to a protein lacking the CK2 site. Treatment with the PK-C activator phorbol 12-myristate 13-acetate (PMA) further increases the level of maximal nuclear accumulation and the initial nuclear import rate. This engineered PMA-responsive NLS may have application in targeting of molecules of interest to the nucleus in response to agents stimulating PK-C.