Figure 4.

Relieving chloride inhibition of With No Lysine kinase (WNK) is not sufficient to increase transepithelial ion transport. Human WNK1 with a leucine to phenylalanine mutation in the chloride binding amino acid, Leu 369, is resistant to chloride inhibition.⁷ The homologous mutation, L421F, was introduced into D. melanogaster With No Lysine kinase (DmWNK) and transgenically expressed in adult tubule principal cells. Tubules were bathed in isotonic standard bathing medium, and fluid secretion and transepithelial potassium flux were measured using the Ramsay assay and ion-specific electrodes (Table 2). Genotypes were control: w; tub-GAL80^ts20/+; c42-GAL4/+ and experimental: w; tub-GAL80^ts20/UAS-DmWNK^L421F; c42-GAL4/+. Flies were reared at 18°C throughout development and shifted to 28°C for 2 days before testing to allow induction of the DmWNK^L421F transgene in adult tubule principal cells; n=17–20 tubules per genotype. P=0.69, unpaired t test.