Figure 2.
Codepletion of GSP-1 and GSP-2 interferes with PAR-2 cortical posterior localization. (A) Left: representative midsection frames of time-lapse imaging of gfp::par-2 zygotes at pronuclear meeting, comparing ctrl(RNAi) and gsp-2(RNAi). N = 4. Right: quantification of the gfp::par-2 size domain in live zygotes at pronuclear meeting. Numbers inside the bars indicate sample size. N = 4. Mean is shown and error bars indicate SD. The P values were determined using two-tailed unpaired Student’s t test. (B) Embryonic lethality of gfp::par-2 and pkc-3(ne4246); gfp::par-2 embryos after ctrl(RNAi) and gsp-1(RNAi). The reported values correspond to the percentage of unhatched embryos over the total progeny (larvae and unhatched embryo). gfp::par-2; ctrl(RNAi), n = 2,040, gfp::par-2; gsp-1(RNAi), n = 1,586, pkc-3(ne4246); gfp::par-2; ctrl(RNAi), n = 751, and pkc-3(ne4246); gfp::par-2; gsp-1(RNAi), n = 382. N = 3. Mean is shown and error bars indicate SEM. The P values were determined using two-way ANOVA Tukey’s multiple comparisons test. (C) Representative midsection frames of time-lapse imaging of gfp::par-2 zygotes at pronuclear meeting, comparing ctrl(RNAi), n = 10, ctrl(RNAi); gsp-1(RNAi), n = 9, ctrl(RNAi); gsp-2(RNAi), n = 11, and gsp-1/-2(RNAi) class I, n = 11 and class II, n = 3. N = 3. Zygotes codepleted of both GSP-1 and GSP-2 show PAR-2 uniformly in the cytoplasm in most of the embryos (class I, 78.6%); class II embryos (21.4%) show a weak PAR-2 localization at the cortex compared to ctrl(RNAi). (D) Quantification of the gfp::par-2 size domain in live zygotes at pronuclear meeting. Numbers inside the bars indicate sample size. N = 3. Mean is shown and error bars indicate SD. The P values were determined using two-tailed unpaired Student’s t test. (E) PAR-2 line profile of live zygotes at pronuclear meeting comparing ctrl(RNAi), n = 10, ctrl(RNAi); gsp-1 (RNAi), n = 9 ctrl(RNAi); gsp-2(RNAi) n = 11, and gsp-1/-2(RNAi) class I, n = 11 and class II, n = 3. Mean is shown and error bars indicate SD. N = 3. ns, P > 0.05; ***, P < 0.001; ****, P < 0.0001. The percentage on the top right of each image in A and C indicates the size of the PAR-2 domain compared to the entire embryo length in the embryo shown. RNAi was performed by feeding in the experiments shown in A and B and by injection in C–E. n = number of embryos analyzed; N = number of independent experiments.

Codepletion of GSP-1 and GSP-2 interferes with PAR-2 cortical posterior localization. (A) Left: representative midsection frames of time-lapse imaging of gfp::par-2 zygotes at pronuclear meeting, comparing ctrl(RNAi) and gsp-2(RNAi). N = 4. Right: quantification of the gfp::par-2 size domain in live zygotes at pronuclear meeting. Numbers inside the bars indicate sample size. N = 4. Mean is shown and error bars indicate SD. The P values were determined using two-tailed unpaired Student’s t test. (B) Embryonic lethality of gfp::par-2 and pkc-3(ne4246); gfp::par-2 embryos after ctrl(RNAi) and gsp-1(RNAi). The reported values correspond to the percentage of unhatched embryos over the total progeny (larvae and unhatched embryo). gfp::par-2; ctrl(RNAi), n = 2,040, gfp::par-2; gsp-1(RNAi), n = 1,586, pkc-3(ne4246); gfp::par-2; ctrl(RNAi), n = 751, and pkc-3(ne4246); gfp::par-2; gsp-1(RNAi), n = 382. N = 3. Mean is shown and error bars indicate SEM. The P values were determined using two-way ANOVA Tukey’s multiple comparisons test. (C) Representative midsection frames of time-lapse imaging of gfp::par-2 zygotes at pronuclear meeting, comparing ctrl(RNAi), n = 10, ctrl(RNAi); gsp-1(RNAi), n = 9, ctrl(RNAi); gsp-2(RNAi), n = 11, and gsp-1/-2(RNAi) class I, n = 11 and class II, n = 3. N = 3. Zygotes codepleted of both GSP-1 and GSP-2 show PAR-2 uniformly in the cytoplasm in most of the embryos (class I, 78.6%); class II embryos (21.4%) show a weak PAR-2 localization at the cortex compared to ctrl(RNAi). (D) Quantification of the gfp::par-2 size domain in live zygotes at pronuclear meeting. Numbers inside the bars indicate sample size. N = 3. Mean is shown and error bars indicate SD. The P values were determined using two-tailed unpaired Student’s t test. (E) PAR-2 line profile of live zygotes at pronuclear meeting comparing ctrl(RNAi), n = 10, ctrl(RNAi); gsp-1 (RNAi), n = 9 ctrl(RNAi); gsp-2(RNAi) n = 11, and gsp-1/-2(RNAi) class I, n = 11 and class II, n = 3. Mean is shown and error bars indicate SD. N = 3. ns, P > 0.05; ***, P < 0.001; ****, P < 0.0001. The percentage on the top right of each image in A and C indicates the size of the PAR-2 domain compared to the entire embryo length in the embryo shown. RNAi was performed by feeding in the experiments shown in A and B and by injection in C–E. n = number of embryos analyzed; N = number of independent experiments.

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