Figure 8.

Butanol inhibits 2B body movement and butanol-resistant GCN3 mutants lack the 2B body. (A) Images from time-lapse microscopy studies using the eIF2Bγ-GFP bearing strains yMK876 [GCD1-S180-GFP (BUT^S)] and yMK880 [GCD1-P180-GFP (BUT^R)] strains, top three rows and bottom three rows, respectively. The strains were incubated in media (−) or media with 1% (vol/vol) or 2% (vol/vol) butanol for 10 min as indicated. Each row contains three stills from a series of 25 images over a period of 2 min, as well as a merged image of all 25 stills, which serves to depict the total extent of 2B body movement. (B) Bar chart depicting the mean distance moved in μ over a 2-min period from at least 40 time-lapse experiments (as in A) using eIF2α-GFP to mark the 2B body in the strains yMK914 [SUI2-GFP GCD1-S180 (BUT^S)] and yMK883 [SUI2-GFP GCD1-P180 (BUT^R)] Error bars, ±1 SEM; *p < 0.01 from an analysis of variance (ANOVA). (C) As in B, except eIF2Bγ is GFP-tagged to follow the 2B body in the strains yMK876 [GCD1-S180-GFP (BUT^S)] and yMK880 [GCD1-P180-GFP (BUT^R)]. (D) Images showing the absence of the eIF2B body in the yMK1567 [BUT1-1 (GCN3-R148K)] and yMK1595 [(BUT1-2 (GCN3-T41A)] strains relative to wild-type controls yMK1596 (GCD1-P180 BUT^R) and yMK1597 (GCD1-S180 BUT^S) using eIF2Bε-GFP (GCD6-GFP) to mark the eIF2B body.