Regulation of NBCe1-B by IRBIT, PIP2 and the WNK/SPAK Kinases

Abstract

The electrogenic Na+-HCO3− co-transporter NBCe1-B plays a crucial role in pHin regulation and epithelial HCO3− secretion by mediating basolateral HCO3− entry, when the luminal HCO3− exit mediated by CFTR and members of the SLC26 transporters.The regulation of NBCe1-B is poorly understood. IRBIT, (inositol 1,4,5-trisphosphate receptor binding protein released with IP3), binds to N terminus of NBCe1-B to markedly increase its activity, while the WNK/SPAK pathway setting the resting state by reducing surface expression of NBCe1-B. NBCe1-B may also be regulated by PIP2 interacting with highly charge module in the N terminus. The site and mechanisms for the regulation by IRBIT, PIP2, and SPAK and the relationship between them are not known.We identified a positive clustered module in NBCe1-B and three arginines within required for regulation by IRBIT and PIP2. The effect of IRBIT and PIP2 are not additive but complementary. The module contains two critical phosphorylation sites. The constitutively cAMP phosphorylated Thr49 is required for all form of NBCe1-B regulations, while Ser65 mediates the function of SPAK. The module is conserved in most NBC superfamily and have the same roles in NBCn1-A. These findings suggest the sites of regulation on NBCe1-B by IRBIT/PIP2 and SPAK and provide a molecular mechanism by which these regulatory factors converge to coordinate epithelia HCO3− secretion.