Editors' ChoiceCell Biology

Turned off by chloride

Sci. Signal.  27 Jan 2015:
Vol. 8, Issue 361, pp. ec22
DOI: 10.1126/scisignal.aaa7526

Ions control membrane potential and function as signaling molecules in cells. Additionally, the electrical and chemical differences in ionic concentrations serve as driving forces for various types of transporters, including the Na+/HCO3- transporters. Na+/HCO3- transporters of the NBC family are essential for the proper functioning of epithelia in the kidney. Shcheynikov et al. showed that NBCe1-A, NBCe1-B, and NBCe2-C contained a GXXXP motif that mediated inhibition of the activity of these transporters by intracellular Cl-. These transporters are electrogenic, moving two negative charges with one positive charge, and their activity can be monitored electrophysiologically. However, when expressed in HeLa cells, only increasing the concentration of intracellular Cl- and not other negatively charged ions reduced the activity of NBCe1-B, and altering the concentration of extracellular Cl- did not inhibit transporter activity. GXXXP motifs are involved in Cl- binding of other proteins, and one or more of these motifs are present in these three transporters. NBCe1-B is activated by binding to IRBIT, a protein released from the inositol trisphosphate (IP3) receptor when IP3 binds. Mutating any residue in the first or third GXXXP motif of NBCe1-B, which did not alter IRBIT binding, resulted in a fully activated transporter in the presence or absence of IRBIT, and the mutated transporter was only inhibited at high concentrations of intracellular Cl-. Mutating residues in both of these motifs eliminated the inhibitory effect of Cl- at any concentration (up to 140 mM). Coexpression of IRBIT prevented high concentrations of Cl- from inhibiting NBCe1-B with mutations in the third GXXXP motif, suggesting that IRBIT binding may alter the conformation of a region of the transporter containing this third GXXXP motif. Coexpression of IRBIT did not affect NBCe2-C activity, but mutations of residues in the first GXXXP motif eliminated inhibition by Cl- at concentrations up to 140 mM. NBCe1-A lacks the first GXXXP motif, and mutation of this motif at the conserved second site trapped the protein in the endoplasmic reticulum. NBCe1-A was not inhibited by Cl-; instead, 140 mM intracellular Cl- increased its activity. However, truncation and deletion analysis revealed that a mutant lacking a 13 amino-acid sequence was inhibited by Cl-, suggesting that this transporter may also have a cryptic Cl- regulatory motif that can be revealed by an unknown factor. These studies suggest that Cl- functions as a direct regulator of transporter activity.

N. Shcheynikov, A. Son, J. H. Hong, O. Yamazaki, E. Ohana, I. Kurtz, D. M. Shin, S. Muallem, Intracellular Cl as a signaling ion that potently regulates Na+/HCO3 transporters. Proc. Natl. Acad. Sci. U.S.A. 112, E329–E337 (2015). [Abstract] [Full Text]