Figure S4.
unc-73/ced-10 pathway negatively regulates klp-7 for the extension of the anterior process of the PLM, related to Fig. 7. (A and B) Confocal images of worms expressing Ppak-1:PAK-1::GFP (shrEx439) and Pmec-4:mCherry in WT (A) and lin-17(0; B). (C and D) Confocal images of worms expressing Pmec-7-GFP (muIs32). Worms were immunostained using anti–phospho-PAK-1(Thr423; magenta) and anti-GFP (green) antibodies in WT (C) and lin-17(0; D). The processed image obtained to visualize the PLM-specific localization of PAK-1 is also presented separately, which is labeled as “α-Phospho-PAK-1(Thr423) (Only PLM).” (E) Quantification of fluorescence intensity corresponding to the anti–PAK-1 immunohistochemistry shown in C and D. Ant, anterior; Pst, posterior. (F) Confocal images of worms expressing Pmec-7-GFP (muIs32) reporter show that an activated version of CED-10 (CED-10G12V) leads to ectopic extensions from ALM and PLM cell bodies (orange arrows). Pmec-4-CED-10WT (shrEx262) and Pmec-4-CED-10G12V (shrEx256) transgenes were used in this experiment. (G and H) The percentage of worms showing ectopic extension phenotype in ALM (G) and PLM (H) neurons. N = 3–4 independent replicates; n (number of neurons) = 23–65. (I and J) Normalized lengths of anterior (I) and posterior (J) processes of the PLM due to the expression of Pmec-4-CED-10G12V (shrEx256) at various concentrations in the WT background. N = 3–4 independent replicates; n (number of neurons) = 20–62. (K) The change in the percentage of PLM neurons with polarity reversal phenotype due to expression of either an inactive (B-type) version of KLP-7 (KLP-7S546E S555E) or an activated version of CED-10 (CED-10G12V) in lin-17(0) background. N = 3–4 independent replicates; n (number of neurons) = 32–115. Error bars represent SEM. Statistical comparisons were made using the χ2 (Fisher’s exact) test (G, H, and K) and ANOVA with Tukey’s multiple comparison test (E, I, and J). *, P < 0.05; **, P < 0.01; ***, P < 0.001.

unc-73/ced-10 pathway negatively regulates klp-7 for the extension of the anterior process of the PLM, related to Fig. 7. (A and B) Confocal images of worms expressing Ppak-1:PAK-1::GFP (shrEx439) and Pmec-4:mCherry in WT (A) and lin-17(0; B). (C and D) Confocal images of worms expressing Pmec-7-GFP (muIs32). Worms were immunostained using anti–phospho-PAK-1(Thr423; magenta) and anti-GFP (green) antibodies in WT (C) and lin-17(0; D). The processed image obtained to visualize the PLM-specific localization of PAK-1 is also presented separately, which is labeled as “α-Phospho-PAK-1(Thr423) (Only PLM).” (E) Quantification of fluorescence intensity corresponding to the anti–PAK-1 immunohistochemistry shown in C and D. Ant, anterior; Pst, posterior. (F) Confocal images of worms expressing Pmec-7-GFP (muIs32) reporter show that an activated version of CED-10 (CED-10G12V) leads to ectopic extensions from ALM and PLM cell bodies (orange arrows). Pmec-4-CED-10WT (shrEx262) and Pmec-4-CED-10G12V (shrEx256) transgenes were used in this experiment. (G and H) The percentage of worms showing ectopic extension phenotype in ALM (G) and PLM (H) neurons. N = 3–4 independent replicates; n (number of neurons) = 23–65. (I and J) Normalized lengths of anterior (I) and posterior (J) processes of the PLM due to the expression of Pmec-4-CED-10G12V (shrEx256) at various concentrations in the WT background. N = 3–4 independent replicates; n (number of neurons) = 20–62. (K) The change in the percentage of PLM neurons with polarity reversal phenotype due to expression of either an inactive (B-type) version of KLP-7 (KLP-7S546E S555E) or an activated version of CED-10 (CED-10G12V) in lin-17(0) background. N = 3–4 independent replicates; n (number of neurons) = 32–115. Error bars represent SEM. Statistical comparisons were made using the χ2 (Fisher’s exact) test (G, H, and K) and ANOVA with Tukey’s multiple comparison test (E, I, and J). *, P < 0.05; **, P < 0.01; ***, P < 0.001.

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