Characterization of the nuclear localization signal in the DNA helicase involved in Werner's syndrome

The nuclear localization signal (NLS) of the DNA helicase involved in Werner's syndrome (WS) was studied. Previously, we noted that the C-terminal region of WS helicase contains the NLS. In this study, we generated in HeLa cells various chimeric proteins consisting of the N-terminal tagged with an enhanced green fluorescent protein and the C-terminal fragments of the WS helicase that were truncated either from N- or C-termini, and we examined the ability of fragments to transfer the fusion proteins to the nucleoplasm by fluorescence microscopy. A small C-proximal region containing 34 amino acid residues (residues 1369-1402) was found to contain full nuclear migration activities. Subsequent amino acid substitution experiments showed that a sequence of three positively charged amino acids (Lys(1371)-Arg(1372)-Arg(1373)) in this region are particularly important. Similar sequence has previously been defined as the nuclear localization signal of SV-40 large T antigen that also acts as a viral DNA helicase. Conservation of this motif was found in the C-terminal regions of the other RecQ type DNA helicases, including murine WS helicase, yeast sgs1 and rqh+1 and human Bloom syndrome DNA helicases.