IRBIT governs epithelial secretion in mice by antagonizing the WNK/SPAK kinase pathway
Dongki Yang, … , Philip J. Thomas, Shmuel Muallem
Dongki Yang, … , Philip J. Thomas, Shmuel Muallem
Published February 7, 2011
Citation Information: J Clin Invest. 2011;121(3):956-965. https://doi.org/10.1172/JCI43475.
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Research Article

IRBIT governs epithelial secretion in mice by antagonizing the WNK/SPAK kinase pathway

Abstract

Fluid and HCO3– secretion are fundamental functions of epithelia and determine bodily fluid volume and ionic composition, among other things. Secretion of ductal fluid and HCO3– in secretory glands is fueled by Na+/HCO3– cotransport mediated by basolateral solute carrier family 4 member 4 (NBCe1-B) and by Cl–/HCO3– exchange mediated by luminal solute carrier family 26, member 6 (Slc26a6) and CFTR. However, the mechanisms governing ductal secretion are not known. Here, we have shown that pancreatic ductal secretion in mice is suppressed by silencing of the NBCe1-B/CFTR activator inositol-1,4,5-trisphosphate (IP3) receptor–binding protein released with IP3 (IRBIT) and by inhibition of protein phosphatase 1 (PP1). In contrast, silencing the with-no-lysine (WNK) kinases and Ste20-related proline/alanine-rich kinase (SPAK) increased secretion. Molecular analysis revealed that the WNK kinases acted as scaffolds to recruit SPAK, which phosphorylated CFTR and NBCe1-B, reducing their cell surface expression. IRBIT opposed the effects of WNKs and SPAK by recruiting PP1 to the complex to dephosphorylate CFTR and NBCe1-B, restoring their cell surface expression, in addition to stimulating their activities. Silencing of SPAK and IRBIT in the same ducts rescued ductal secretion due to silencing of IRBIT alone. These findings stress the pivotal role of IRBIT in epithelial fluid and HCO3– secretion and provide a molecular mechanism by which IRBIT coordinates these processes. They also have implications for WNK/SPAK kinase–regulated processes involved in systemic fluid homeostasis, hypertension, and cystic fibrosis.

Authors

Dongki Yang, Qin Li, Insuk So, Chou-Long Huang, Hideaki Ando, Akihiro Mizutani, George Seki, Katsuhiko Mikoshiba, Philip J. Thomas, Shmuel Muallem

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Figure 1

The WNK/SPAK and IRBIT/PP1 pathways in ductal fluid secretion.

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The WNK/SPAK and IRBIT/PP1 pathways in ductal fluid secretion. (A) Seale...
(A) Sealed pancreatic ducts in primary culture were treated with the indicated siRNA for 48–60 hours before measurement of stimulated ductal fluid secretion by video microscopy. Ducts treated with scrambled siRNA were also treated with the specific PP1 inhibitor tautomycin (3 nM). Similar inhibition of fluid secretion by tautomycin was observed with ducts not treated with scrambled siRNA. Secretion was initiated by incubation of the ducts in HCO3–-buffered medium and stimulation with 10 μM forskolin. The traces are the mean ± SEM of 4–10 experiments. (B) Model of the main transporters determining ductal fluid secretion and how they may be regulated by the WNK/SPAK pathway.