Figure 1

Loss of Grk4 disrupts normal pronephros patterning. (A) Cartoon illustrating the segments of the developing zebrafish pronephros and marker genes used to visualize the segments. (B) Loss of Grk4 increased the area of the glomeruli, as shown by wt1a expression. n=6 experiments with 150-192 embryos in total. ^****P<0.0001 (data were analyzed using a two-tailed Welch's test). n=64 WT and 52 homozygous mutant embryos, respectively. ^**P=0.0012 (data were analyzed using a two-tailed Welch's test). (C) Grk4 depletion limited the abundance of cells positive for the PCT marker slc20a1a. n=7 experiments with 118-143 embryos in total. ^**P=0.0029 (data were analyzed using a two-tailed Welch's test). (D) Grk4 knockdown resulted in the elongation of the PST as shown by trpm7 expression. n=3 experiments with 73-74 embryos in total. ^****P<0.0001 (data were analyzed using a two-tailed Welch's test). (E) The DE segment, as detected by rfx2 expression, was enlarged in Grk4 knockdown and mutant embryos compared to control embryos. n=3 experiments with 51-128 embryos injected with MOs in total. ^****P<0.0001 (data were analyzed using a two-tailed Welch's test). n=13 WT and 15 homozygous mutant embryos, respectively. ^*P=0.0451 (data were analyzed using a two-tailed Welch's test). (F) The DL segment was shortened upon Grk4 loss-of-function. n=3 experiments with 56-77 embryos in total. ^****P<0.0001 (data were analyzed using a two-tailed Welch's test). All images: Whole mount in situ hybridization at 20-22 somite stage (ss). Arrows and brackets indicate the expression domain. Scale bars, 200 μm. Glom, glomerulus; PCT, proximal convoluted tubule; PST, proximal straight tubule; DE, distal early tubule; DL, distal straight tubule; Grk4, G protein-coupled receptor kinase 4; CTRL, control; MO, morpholino oligonucleotide; WT, wild-type.