It has been reported that Lysine-305 is needed for the nuclear import of the p53 protein (Liang et al., 1998). In the present study, further mutagenesis analyses were carried out between Lys-305 and the major nuclear localization signal (NLS I) of p53. It was found that a single mutation of Arg-306 resulted in the defect of p53 nuclear import. This effect is the same as that of Lys-305 mutation. Other mutations between Arg-306 and NLS I have no effect on the nuclear import of p53. However, deletions of more than two amino acids between this region abolished the transport of p53 into the nucleus. These results indicate that a basic domain other than the well defined NLS is required for the nuclear import of p53. A spacer between this basic domain and NLS I is necessary for the entrance of p53 into the cell nucleus.