Sip1, an AP-1 Accessory Protein in Fission Yeast, Is Required for Localization of Rho3 GTPase
Figure 4
Both of Rho3 and Apm1 fail to co-localize at the Golgi/endosomes in sip1-i4 mutant cells.
(A) Subcellular localization of Apm1-GFP in wild-type (wt) and sip1-i4 mutant cells (sip1-i4). Cells expressing Apm1-GFP were cultured in YPD medium at 27°C, were incubated with the dye FM4-64 for 5 min at 27°C to visualize the Golgi/endosomes. The fluorescence of the FM4-64 was examined under the fluorescence microscope. Arrowheads indicated the localization of Apm1-GFP to the Golgi/endosomes. Bar, 10 µm. (B) Subcellular localization of GFP-Rho3 in wild-type (wt) and sip1-i4 mutant cells (sip1-i4). Cells expressing Rho3 were cultured in YPD medium at 27°C, following which they were incubated with FM4-64 dye for 5 min at 27°C to visualize the Golgi/endosomes. FM4-64 fluorescence was examined using a fluorescence microscope. Arrowheads indicate the dot-like structures of GFP-Rho3 and the Golgi/endosomes stained with FM4-64, double arrowheads indicate cytoplasmic accumulation, and arrows indicate the concentrated fluorescence at the cell division site. Bar, 10 µm. (C) Percentage of cells in which Rho3 were localized at the cell division site in wild-type (wt) and sip1-i4 cells. (D) Quantitative analysis for the number of Rho3 dots co-localizing with FM4-64/cells in wt and sip1-i4 cells. Cells in C and D were incubated as those in B. Data are the means ± standard deviations of 3 independent experiments with 150 cells in C and D.