Figure 4

PAT-9 contains a lysine/arginine-rich NLS. A) The amino acid sequence predicted from the pat-9 cDNA cloned from N2 worms is shown. Each of the three putative zinc fingers is underlined. The additional two amino acids not present in the WormBase model are shown in red. The deduced NLS is highlighted in yellow. B-H) Plasmids expressing truncations of PAT-9 fused to GFP under control of the myo-3 promoter were used to identify the PAT-9 NLS. GFP expression from transgenic animals carrying extrachromosomal arrays of B) P _myo-3 ::gfp (pPD118.20) expressing GFP alone, C) P _myo-3 ::pat-9zf1::gfp, containing the amino terminal sequence and the first zinc finger of PAT-9, D) P _myo-3 ::pat-9 C::gfp containing only the carboxyl terminal sequence, E) P _myo-3 ::pat-9CNLS::gfp containing the carboxyl terminal sequence with the putative NLS, F) P _myo-3 ::pat-9zf123NLS::gfp containing three zinc fingers and the predicted NLS. The exclusively nuclear localization of P _myo-3 ::pat-9zf123NLS::gfp was confirmed by G) DAPI staining and H) merging of GFP and DAPI images. Bars = 10 μm.