Figure 4
From: Yeast karyopherin Kap95 is required for cell cycle progression at Start
Control of the subcellular localization of Swi4. (A) A putative NLS sequence in the Swi4 protein is indicated (white box). Amino acids substitutions introduced to inactivate the NLS are shown in grey. Grey boxes represent the DNA binding (DBD), the ankyrin (ANK) and the Swi6 binding (SBD) domains. (B) Exponentially growing cells of the wild type strain (W303-1a) transformed with plasmids pSWI4NLS-GFP4 and pSWI4NLSi-GFP4 were analyzed by fluorescence microscopy. GFP signal images are shown. (C) Exponentially growing cells of the wild type (W303-1a) and tetO 7 :KAP95 (JCY635) strains transformed with plasmid pSWI4NLS-GFP4 were incubated in the presence of 5 μg/mL doxycycline for 6 hours and analyzed by fluorescence microscopy. GFP signal images are shown. (D) Exponentially growing cells of the wild type strain (W303-1a) transformed with plasmids pGAL1:HA-SWI4 and pGAL1:HA-SWI4NLSi were incubated in the presence of 5 μg/mL doxycycline for 6 hours and assayed by indirect immunofluorescence. The HA indirect-fluorescence signals, the DAPI staining of DNA and DIC images are shown. (E) Exponentially growing cells of the the wild type (W303-1a) and tetO 7 :KAP95 (JCY635) strains transformed with plasmid pGAL1:HA-SWI4 were incubated in the presence of 5 μg/mL doxycycline for 6 hours and assayed by indirect immunofluorescence. The HA indirect-fluorescence signals, the DAPI staining of DNA and DIC images are shown. (F) Exponentially growing cells of the the wild type (W303-1a) and the srp1 strains transformed with the plasmid pGAL1:HA-SWI4 were incubated at 37° for 2 hours and and assayed by indirect immunofluorescence. The wild type cells showed a nuclear localization of Swi4 similar to that showed in (E) and are not shown.