Fig. 3

rips-1 mutants are resistant to thiol-reducing agents, independent of METR-1 and enhanced by loss of MCE-1 (a, b). Plotted percentage of adults of wild-type (N2) and DTT-resistant rips-1(ij109) mutant. Embryos were placed on soy peptone NGM agar plates with OP50-1 as food source, supplemented with 0 mM DTT (n = N2: 292, rips-1: 265), 2.5 mM DTT (n = N2: 305, rips-1: 268), or 5 mM DTT (n = N2: 292, rips-1: 261) (a), or with 0 mM β-mercaptoethanol (2-ME) (n = N2: 164, rips: 220) or 2.5 mM 2-ME (n = N2: 200, rips-1: 227) (b). c rips-1(ij109) mutant embryos were assessed for development to adults on 0 mM DTT (n = 205), 0.5 mM DTT (n = 190), 1 mM DTT (n = 244), 2 mM DTT (n = 176), or 5 mM DTT (n = 214) and compared to metr-1;rips-1 double mutant [TP391, metr-1(ok521);rips-1(ij109)] on 0 mM DTT (n = 280), 0.5 mM DTT (n = 272), 1 mM DTT (n = 260), 2 mM DTT (n = 270), or 5 mM DTT (n = 210). d Development of wild-type (N2), rips-1(ij109), metr-1(ok521);rips-1(ij109), and metr-1(ok521) worms on 5 mM DTT OP50-1 plates supplemented with 0, 1, or 10 mM methionine. The number of N2, rips-1 mutant, metr-1;rips-1 mutant, and metr-1 mutant animals used in this experiment in this order are as follows: no methionine (n = 166, 244, 303, 217), 1 mM methionine (n = 259, 279, 259, 292), 10 mM methionine (n = 213, 176, 125, 140). e RNAi by feeding was performed in a rips-1(ij109) mutant with control (L4440) or mce-1 RNAi followed by selection on 5 mM DTT plates. Enhanced resistance was confirmed for a double mutant on DTT, as rips-1(ij109) single mutant could not survive on 6 mM DTT (bottom left) while mce-1(ok243);rips-1(ij109) double mutant survived up to 7 mM DTT (bottom right). Scale bars denote 1 mm. f Percentages of adults of N2 wild type and various mutants on 0, 5, 6, and 7 mM DTT. The numbers of animals used in this experiment in each treatment group (0, 5, 6, and 7 mM, respectively) are as follows: N2—n = 245, 206, 129, and 206; rips-1—n = 202, 243, 173, and 267; mce-1;rips-1—n = 305, 253, 305, and 259; mce-1—n = 225, 206, 195, and 214; metr-1—n = 252, 308, 257, and 261; and mmcm-1—n = 184, 191, 148, and 123. Percentage survival to adulthood was determined after 4 days of treatment. In a, b, and f, red points with lines denote the mean and SEM. For a and f, p-values were determined from two-way ANOVA, followed by Dunnett’s test. For b, p-values were determined with the two-tailed t-test. For c and d, p-values were determined from Fisher’s exact test. NS, not significant, *p < 0.05, **p < 0.01, ***p < 0.001. For all panels, purple significance marks indicate the comparison of mutant worm strains to N2 wild type for each treatment group, blue significance marks indicate the comparison of treatment groups (e.g., 1 mM DTT) to no treatment groups (e.g., 0 mM DTT) for each worm strain, and red significance marks indicate comparison of metr-1;rips-1 mutant strain to metr-1 mutant strain for each treatment group