Figure 2.
Loss of Matriptase inhibition activates Par2b-EGFR-PLD-mTORC1 pathway. (A–F) Representative bright field images of the caudal fin fold at 48 hpf. hai1ahi2217 embryos were treated with vehicle (B), a morpholino targeting par2b (C), or inhibitors of EGFR (PD168393, D), PLD (FIPI, E), or mTORC1 (rapamycin, F). White, gray, and black boxes correspond with the phenotypic classes No phenotype, Moderate, and Severe, respectively, as shown in G. See also Fig. S2. (A′–F′) Representative images of anti-pEGFR staining (brown) in the caudal fin fold. (A′′–F′′) Representative maximum intensity projection images of BrdU labeling (red) in the caudal fin fold. (G) Distributions of the morphological phenotypes in treated embryos, scored according to severity of the phenotype. n = 32–50 embryos per condition in two independent experiments. (H) Quantification of the numbers of pEGFR-positive cells in A′–F′, in relation to the total area of the fin fold. n = 5–9 embryos. (I) Representative immunoblots of pRPS6 levels in par2b morpholino-injected or inhibitor-treated embryos. n = 20 pooled tails per condition. Quantification below each blot indicates the average ratios of each pRPS6 band, normalized against tubulin and relative to the sibling control, calculated from at least three independent experiments. (J) Quantification of numbers of BrdU-positive cells in A′′–F′′ relative to the total number of nuclei. n = 7–15 embryos. (K–N)krt19:EGFP-PASS transient expression to visualize PA localization in basal keratinocytes of control siblings and hai1ahi2217mutants. Maximal intensity projections and single planes of boxed regions (insets, same magnification), as used for the determination of relative membrane-cytoplasm values shown in O. See also Fig. S2 J. (O) Quantification of EGFP-PASS localization to lateral membranes. Ratios of fluorescence in the proximity of the cell membranes versus the cytoplasm were determined as demonstrated in Fig. S2 J. n = 5–8 cells. For quantifications and statistical significances, see legend of Fig. 1. Scale bars in A–F, 200 µm; in A′–F′′, 50 µm; and in K–N, 10 µm.

Loss of Matriptase inhibition activates Par2b-EGFR-PLD-mTORC1 pathway. (A–F) Representative bright field images of the caudal fin fold at 48 hpf. hai1ahi2217 embryos were treated with vehicle (B), a morpholino targeting par2b (C), or inhibitors of EGFR (PD168393, D), PLD (FIPI, E), or mTORC1 (rapamycin, F). White, gray, and black boxes correspond with the phenotypic classes No phenotype, Moderate, and Severe, respectively, as shown in G. See also Fig. S2. (A′–F′) Representative images of anti-pEGFR staining (brown) in the caudal fin fold. (A′′–F′′) Representative maximum intensity projection images of BrdU labeling (red) in the caudal fin fold. (G) Distributions of the morphological phenotypes in treated embryos, scored according to severity of the phenotype. n = 32–50 embryos per condition in two independent experiments. (H) Quantification of the numbers of pEGFR-positive cells in A′–F′, in relation to the total area of the fin fold. n = 5–9 embryos. (I) Representative immunoblots of pRPS6 levels in par2b morpholino-injected or inhibitor-treated embryos. n = 20 pooled tails per condition. Quantification below each blot indicates the average ratios of each pRPS6 band, normalized against tubulin and relative to the sibling control, calculated from at least three independent experiments. (J) Quantification of numbers of BrdU-positive cells in A′′–F′′ relative to the total number of nuclei. n = 7–15 embryos. (K–N)krt19:EGFP-PASS transient expression to visualize PA localization in basal keratinocytes of control siblings and hai1ahi2217mutants. Maximal intensity projections and single planes of boxed regions (insets, same magnification), as used for the determination of relative membrane-cytoplasm values shown in O. See also Fig. S2 J. (O) Quantification of EGFP-PASS localization to lateral membranes. Ratios of fluorescence in the proximity of the cell membranes versus the cytoplasm were determined as demonstrated in Fig. S2 J. n = 5–8 cells. For quantifications and statistical significances, see legend of Fig. 1. Scale bars in A–F, 200 µm; in A′–F′′, 50 µm; and in K–N, 10 µm.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal