TY - JOUR
T1 - Intracellular chloride and scaffold protein mo25 cooperatively regulate transepithelial ion transport through WNK signaling in the malpighian tubule
AU - Sun, Qifei
AU - Wu, Yipin
AU - Jonusaite, Sima
AU - Pleinis, John M.
AU - Humphreys, John M.
AU - He, Haixia
AU - Schellinger, Jeffrey N.
AU - Akella, Radha
AU - Stenesen, Drew
AU - Krämer, Helmut
AU - Goldsmith, Elizabeth J.
AU - Rodan, Aylin R.
N1 - Publisher Copyright:
© 2018 by the American Society of Nephrology.
PY - 2018/5
Y1 - 2018/5
N2 - BackgroundWith No Lysine kinase (WNK) signaling regulates mammalian renal epithelial ion transport to maintain electrolyte and BP homeostasis. Our previous studies showed a conserved role for WNK in the regulation of transepithelial ion transport in the Drosophila Malpighian tubule. Methods Using in vitro assays and transgenic Drosophila lines, we examined two potential WNK regulators, chloride ion and the scaffold protein mouse protein 25 (Mo25), in the stimulation of transepithelial ion flux. Results In vitro, autophosphorylation of purified Drosophila WNK decreased as chloride concentration increased. In conditions inwhich tubule intracellular chloride concentration decreased from30 to 15mMas measured using a transgenic sensor, DrosophilaWNK activity acutely increased. DrosophilaWNK activity in tubules also increased or decreased when bath potassium concentration decreased or increased, respectively. However, a mutation that reduces chloride sensitivity of Drosophila WNK failed to alter transepithelial ion transport in 30 mM chloride. We, therefore, examined a role for Mo25. In in vitro kinase assays, Drosophila Mo25 enhanced the activity of the Drosophila WNK downstream kinase Fray, the fly homolog of mammalian Ste20-related proline/alanine-rich kinase (SPAK), and oxidative stress-responsive 1 protein (OSR1). Knockdown of Drosophila Mo25 in the Malpighian tubule decreased transepithelial ion flux under stimulated but not basal conditions. Finally, whereas overexpression of wild-type Drosophila WNK, with or without Drosophila Mo25, did not affect transepithelial ion transport, Drosophila Mo25 overexpressed with chloride-insensitive Drosophila WNK increased ion flux. Conclusions Cooperative interactions between chloride andMo25 regulateWNK signaling in a transporting renal epithelium.
AB - BackgroundWith No Lysine kinase (WNK) signaling regulates mammalian renal epithelial ion transport to maintain electrolyte and BP homeostasis. Our previous studies showed a conserved role for WNK in the regulation of transepithelial ion transport in the Drosophila Malpighian tubule. Methods Using in vitro assays and transgenic Drosophila lines, we examined two potential WNK regulators, chloride ion and the scaffold protein mouse protein 25 (Mo25), in the stimulation of transepithelial ion flux. Results In vitro, autophosphorylation of purified Drosophila WNK decreased as chloride concentration increased. In conditions inwhich tubule intracellular chloride concentration decreased from30 to 15mMas measured using a transgenic sensor, DrosophilaWNK activity acutely increased. DrosophilaWNK activity in tubules also increased or decreased when bath potassium concentration decreased or increased, respectively. However, a mutation that reduces chloride sensitivity of Drosophila WNK failed to alter transepithelial ion transport in 30 mM chloride. We, therefore, examined a role for Mo25. In in vitro kinase assays, Drosophila Mo25 enhanced the activity of the Drosophila WNK downstream kinase Fray, the fly homolog of mammalian Ste20-related proline/alanine-rich kinase (SPAK), and oxidative stress-responsive 1 protein (OSR1). Knockdown of Drosophila Mo25 in the Malpighian tubule decreased transepithelial ion flux under stimulated but not basal conditions. Finally, whereas overexpression of wild-type Drosophila WNK, with or without Drosophila Mo25, did not affect transepithelial ion transport, Drosophila Mo25 overexpressed with chloride-insensitive Drosophila WNK increased ion flux. Conclusions Cooperative interactions between chloride andMo25 regulateWNK signaling in a transporting renal epithelium.
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U2 - 10.1681/ASN.2017101091
DO - 10.1681/ASN.2017101091
M3 - Article
C2 - 29602832
AN - SCOPUS:85046371439
SN - 1046-6673
VL - 29
SP - 1449
EP - 1461
JO - Journal of the American Society of Nephrology
JF - Journal of the American Society of Nephrology
IS - 5
ER -