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Fig. 7 kctd15a/b-tfap2a autoregulatory feedback loop balances DE pronephric differentiation. (A) Whole-mount in situ hybridization of slc12a1 (DE, purple) and cdh17 (tubule, red) in hs:tfap2a transgenic background in combination with kctd15b MO treatment at 24 hpf. HS+ indicates application of heat-shock treatment at the 12 ss. Blue arrowheads annotate proximal and distal edges of the slc12a1 expression domain. Scale bar: 35 μm. (B) Quantification of cdh17 tubule expression domain length in control and treatment groups. (C) Bar graph depicting percentage DE occupancy of the tubule. HS−, no heat-shock treatment; HS+, heat-shock treatment. (D) Whole-mount in situ hybridization of slc12a1 in wild-type siblings and trm−/− uninjected controls or trm−/− injected with kctd15a/b MO. Blue arrowheads indicate single DE cells. Scale bar: 35 μm. (E) Quantification of slc12a1 expression domain absolute length per nephron. (F) Penetrance of reduced slc12a1 expression in trm+/− incrosses, uninjected controls versus kctd15a/b MO treatment. (G) Graph depicting the number of DE cells per nephron in trm−/− uninjected controls versus trm−/−+kctd15a/b MO. n≥6 embryos quantified for each control and experimental group. *P<0.05; **P<0.01; ***P<0.001; N.S., not significant. Data are mean±s.d. Absolute lengths and DE:tubule ratios were compared using ANOVA. Penetrances were compared using Fischer's exact test. Cell counts were compared using unpaired t-tests.