Panel A shows immunoblots and line graphs comparing glucose starvation-dependent protein accumulation in wild-type and elo3 mutant cells. Panel B shows confocal fluorescence microscopy images, bar graphs, and box plots analyzing Ypf1-GFP NVJ partitioning dynamics. Panel C shows immunoblots examining protein accumulation changes following cycloheximide treatment during prolonged glucose starvation conditions. Panel D shows immunoblots and line graphs comparing glucose starvation-dependent protein accumulation in wild-type and elo2 cells. Panel E shows immunoblots and line graphs analyzing Elo2-FLAG and Elo3-FLAG protein abundance changes during starvation.
Fatty acid biogenesis is involved in NVJ remodeling. (A) Immunoblotting of WT and elo3Δ cell extracts after GS treatments for the indicated durations. Line graphs show relative protein levels normalized to the protein abundance of the indicated protein in WT cells at 24 h, which was defined as 100%. For Ypf1, values indicate the proportion of the doubly glycosylated form. Data are means ± SEM (n = 3). (B) The progression of Ypf1-GFP NVJ partitioning was assessed in cells expressing Vph1-mCherry after GS treatment for the indicated time periods. Single focal plane images obtained by confocal fluorescence microscopy are shown. Bar graph shows the percentage of cells displaying NVJ-localized Ypf1-GFP. Values are means ± SE (n = 3). Box-and-whisker plots show the fluorescence intensity and area of Ypf1-GFP localized at the NVJ. The boxes indicate the median and interquartile range, and the whiskers extend to 1.5 times the interquartile range (Tukey method). A total of 25 cells pooled from three independent experiments were analyzed. ns: not significant, ∗∗∗: P < 0.001 ∗∗∗∗: P < 0.0001. P values were obtained using the Mann–Whitney U test. (C) Immunoblotting was performed using whole-cell extracts prepared from cells shifted to glucose-starvation medium containing either DMSO or 100 µg/ml cycloheximide and incubated for the indicated times. (D) Immunoblotting of WT and elo2Δ cell extracts after GS treatments for the indicated durations. Line graphs were generated as in A. (E) Immunoblotting of whole-cell extracts prepared from WT or elo2Δ cells expressing Elo3-FLAG and from WT or elo3Δ cells expressing Elo2-FLAG after GS treatments for the indicated durations. Line graphs show relative protein levels normalized to the abundance of Elo2-FLAG or Elo3-FLAG in WT cells before GS, which was defined as 100%. Data are means ± SEM (n = 3). Asterisks indicate nonspecific bands. Source data are available for this figure: SourceData F4.