Panel A: A series of microscopy images showing yeast cells under different genetic conditions and treatments. The images include differential interference contrast (DIC) and fluorescence microscopy images stained with LipiBlue to visualize lipid droplets (LDs). The cells express either Hmg1-GFP or Hmg2-GFP and are observed under glucose-rich (Log) and glucose-starved (GS) conditions. Panel B: A bar graph displaying that the y-axis represents the percentage of cells, and the x-axis lists different yeast strains. Panel C: A box plot showing the percentage of LD area relative to total cell area. The y-axis represents the percentage of LD area, and the x-axis lists different yeast strains. Panel D: Immunoblot images of whole-cell lysates from various yeast strains, with or without a 1-day glucose starvation treatment, analyzed for the presence of Hmg1-GFP, Hmg2-GFP, Nsg1, Nsg2, Ypf1, and Tim23. Panel E shows confocal fluorescence microscopy images of LipiBlue-stained nsg1 deletion nsg2 deletion hmg2 deletion cells expressing empty vector or Nsg2 after glucose starvation.
LDs accumulate in an Hmg1-dependent manner in nsg1Δnsg2Δ cells. (A) LDs in the indicated cells after a 1-day GS treatment were visualized using the LD-staining dye LipiBlue and imaged by confocal fluorescence microscopy. The indicated yeast cells expressing Hmg1-GFP or Hmg2-GFP were observed by confocal fluorescence microscopy under glucose-rich (Log) and glucose-starved (GS) conditions. Yeast cells with strongly and weakly stained LDs were outlined with solid and dotted lines, respectively. LDs are shown as maximum intensity projection images. Fluorescence images of Hmg1-GFP and Hmg2-GFP represent a single optical section. Scale bars, 5 µm. (B) Bar graph shows the populations of cells with weak and strong LD fluorescence intensity based on maximum projection images acquired in A. Values are means ± SE (n = 4). At least 100 cells were analyzed per biological replicate. (C) Box-and-whisker plot shows the percentage of LD area relative to total cell area (n = 40 cells pooled from three independent experiments, N = 3). The boxes indicate the median and interquartile range, and the whiskers extend to 1.5 times the interquartile range (Tukey method). ns: not significant, *P < 0.05, ***P < 0.001, and ****P < 0.0001. P values were obtained using the Mann–Whitney U test. (D) Whole-cell lysates prepared from the indicated yeast cells, with or without a 1-day GS treatment, were analyzed by immunoblotting. HmgX denotes either Hmg1 or Hmg2. Asterisks indicate nonspecific bands. (E) LDs in nsg1Δnsg2Δhmg2Δ cells carrying either an empty vector or a CEN-plasmid expressing Nsg2 were stained with LipiBlue and visualized by confocal fluorescence microscopy after a 1-day GS treatment. Yeast cells showing weak LD staining were outlined with dotted lines. LDs are shown as maximum intensity projection images. Scale bars, 5 µm. Source data are available for this figure: SourceData F5.