CHARACTERIZATION OF THE NUCLEAR-LOCALIZATION SIGNAL IN THE DNA HELICASE INVOLVED IN WERNERS-SYNDROME

The nuclear localization signal (NLS) of the DNA helicase involved in Werner's syndrome (WS) was studied. Previously, we noted that the C-te rminal region of WS helicase contains the NLS. In this study, we gener ated in HeLa cells various chimeric proteins consisting of the N-termi nal tagged with an enhanced green fluorescent protein and the C-termin al fragments of the WS helicase that were truncated either from N- or C-termini, and we examined the ability of fragments to transfer the fu sion 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 a mino acid substitution experiments showed that a sequence of three pos itively charged amino acids (Lys(1371)-Arg(1372)-Arg(1373)) in this re gion are particularly important. Similar sequence has previously been defined as the nuclear localization signal of SV-40 large T antigen th at also acts as a viral DNA helicase. Conservation of this motif was f ound in the C-terminal regions of the other RecQ type DNA helicases, i ncluding murine WS helicase, yeast sgs1 and rqh+1 and human Bloom synd rome DNA helicases.