Fig. 3.

Genes from the FA elongation cycle and sphingolipid metabolism impact the localization and the activity of Acc1-GFP. a) Deletion of ELO2, ELO3, and TSC3 impacts the localization of Acc1-GFP. Wild-type (YKB 3954), elo2Δ (YKB 4593), elo3Δ (YKB 4594), and tsc3Δ (YKB 4599) cells expressing endogenously tagged Acc1-GFP were grown to early-log phase at 30°C in YPD and assessed for Acc1-GFP localization. Representative brightfield and fluorescent images are shown. Scale bar: 4 µm. b) The average volume of Acc1-GFP structures in each strain was quantified using IMARIS software. Quantification is the average of 3 biological replicates, a minimum of 100 cells per replicate was scored. Error bars denote the standard error of the mean (SEM). n = 3, *P < 0.05 determined using a 1-way ANOVA test with a Tukey’s multiple comparisons test. c) elo2Δ, elo3Δ, and tsc3Δ cells display increased sensitivity to Sor A. Wild-type (YKB 1079), elo2Δ (YKB 3913), elo3Δ (YKB 3914), and tsc3Δ (YKB 3228) cells were grown to early-log phase before being diluted to an OD₆₀₀ of 0.1 in YPD with DMSO control or 0.1 µM or 0.2 µM Sor A in DMSO, and an automated growth curve analysis was performed at 30°C for 48 h. Growth rate was calculated from 3 biological replicates. Error bars denote the standard error of the mean (SEM). n = 3, *P < 0.05 determined using a 2-way ANOVA test with a Tukey’s multiple comparisons test. d) Acc1 activity is reduced in elo3Δ cells. Wild-type (YKB 3954) and elo3Δ (YKB 4594) cells expressing endogenously tagged Acc1-GFP were grown to early-log phase at 30°C in YPD. Acc1-GFP was immunoprecipitated and Acc1 activity measured and normalized to Acc1-GFP protein abundance. Error bars denote the standard error of the mean (SEM). n = 3, *P < 0.05 determined using a 2-tailed t-test.