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. 2002 Apr 1;157(1):63–78. doi: 10.1083/jcb.200201037

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Copyright © 2002, The Rockefeller University Press

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Figure 7. — Kes1p targeting to yeast Golgi membranes. (A) Kes1p localization domains. YCp(GFP), YCp(KES1-GFP) and mutant YCp(kes1 ^3E-GFP), YCp(kes1 ^E312K-GFP), YCp(kes1 ^W317A-GFP), and YCp(kes1 ^K109A-GFP) plasmids were introduced into a kes1 yeast strain. Fluorescence images were captured at 26°C. The kes1p^3E-GFP and chimeras exhibit a diffuse cytosolic distribution that mimics that of GFP alone. In these three cases, signal is excluded from vacuoles (the dark circular structures). The wild-type (WT) control Kes1p-GFP and the mutant Kes1p^E312K-GFP and Kes1p^W317A-GFP chimeras exhibit typical punctate staining patterns. (B) PI(4)P synthesis and Kes1p localization. A YCp(KES1::GFP) plasmid was introduced into wild-type, pik1-101 ^ts, stt4 ^ts, sec14-1 ^ts, and mss4-2 ^ts strains as indicated at right. Fluorescence images were captured at 26°C and after shift to 37°C for 2 h (indicated at top) at an excitation wavelength of 495nm. Pik1p defects exert major defects in Kes1p localization to Golgi membranes, whereas inactivation of Stt4p and Sec14p has more modest effects.

Figure 7. — Kes1p targeting to yeast Golgi membranes. (A) Kes1p localization domains. YCp(GFP), YCp(KES1-GFP) and mutant YCp(kes1 ^3E-GFP), YCp(kes1 ^E312K-GFP), YCp(kes1 ^W317A-GFP), and YCp(kes1 ^K109A-GFP) plasmids were introduced into a kes1 yeast strain. Fluorescence images were captured at 26°C. The kes1p^3E-GFP and chimeras exhibit a diffuse cytosolic distribution that mimics that of GFP alone. In these three cases, signal is excluded from vacuoles (the dark circular structures). The wild-type (WT) control Kes1p-GFP and the mutant Kes1p^E312K-GFP and Kes1p^W317A-GFP chimeras exhibit typical punctate staining patterns. (B) PI(4)P synthesis and Kes1p localization. A YCp(KES1::GFP) plasmid was introduced into wild-type, pik1-101 ^ts, stt4 ^ts, sec14-1 ^ts, and mss4-2 ^ts strains as indicated at right. Fluorescence images were captured at 26°C and after shift to 37°C for 2 h (indicated at top) at an excitation wavelength of 495nm. Pik1p defects exert major defects in Kes1p localization to Golgi membranes, whereas inactivation of Stt4p and Sec14p has more modest effects.