Up-regulation of nuclear protein import by nuclear localization signal sequences in Living cells

Using an in vivo assay system, nuclear import ability in individual cells w as determined by examining the nuclear import rate. It was found that when a small (not excess) amount of SV40 T-NLS peptides was co-injected, the nuc lear import rate of SV40 T-NLS-containing substrates apparently increased. This up-regulation was reproduced by the co-injection of peptides containin g bipartite type NLS of CBP80, but not mutated nonfunctional NLS peptides, which suggests that these phenomena are specific for functional NLSs, It wa s further shown that although, in growth-arrested cells, the nuclear import rate was down-regulated compared to growing cells, the elevation of the fu nctional import rate by co-injected NLS peptides reached the same level as in proliferating cells. This up-regulation was abolished by the addition of a protein kinase inhibitor, staurosporine. These results suggest that alth ough potential nuclear import ability does not vary in each cell, the rate of nuclear import may be controlled by the amount of karyophilic proteins, which need to be carried into the nucleus from the cytoplasm, possibly via an NLS-dependent phosphorylation reaction. (C) 1999 Federation of European Biochemical Societies.