Molecular physiology and genetics of Na+-independent SLC4 anion exchangers

Abstract

Plasmalemmal Cl(-)/HCO(3)(-) exchangers are encoded by the SLC4 and SLC26 gene superfamilies, and function to regulate intracellular pH, [Cl(-)] and cell volume. The Cl(-)/HCO(3)(-) exchangers of polarized epithelial cells also contribute to transepithelial secretion and reabsorption of acid-base equivalents and Cl(-). This review focuses on Na(+)-independent electroneutral Cl(-)/HCO(3)(-) exchangers of the SLC4 family. Human SLC4A1/AE1 mutations cause the familial erythroid disorders of spherocytic anemia, stomatocytic anemia and ovalocytosis. A largely discrete set of AE1 mutations causes familial distal renal tubular acidosis. The Slc4a2/Ae2(-/-) mouse dies before weaning with achlorhydria and osteopetrosis. A hypomorphic Ae2(-/-) mouse survives to exhibit male infertility with defective spermatogenesis and a syndrome resembling primary biliary cirrhosis. A human SLC4A3/AE3 polymorphism is associated with seizure disorder, and the Ae3(-/-) mouse has increased seizure susceptibility. The transport mechanism of mammalian SLC4/AE polypeptides is that of electroneutral Cl(-)/anion exchange, but trout erythroid Ae1 also mediates Cl(-) conductance. Erythroid Ae1 may mediate the DIDS-sensitive Cl(-) conductance of mammalian erythrocytes, and, with a single missense mutation, can mediate electrogenic SO(4)(2-)/Cl(-) exchange. AE1 trafficking in polarized cells is regulated by phosphorylation and by interaction with other proteins. AE2 exhibits isoform-specific patterns of acute inhibition by acidic intracellular pH and independently by acidic extracellular pH. In contrast, AE2 is activated by hypertonicity and, in a pH-independent manner, by ammonium and by hypertonicity. A growing body of structure-function and interaction data, together with emerging information about physiological function and structure, is advancing our understanding of SLC4 anion exchangers.

Fig. 1.

Schematic diagram of polypeptide variants expressed by the genes encoding the SLC4 Na⁺-independent anion exchangers, AE1, AE2 and AE3. Predicted transmembrane (TM) domains are blue. Total polypeptide lengths are on the right. Lengths of variant N-terminal sequences are indicated within the leftmost boxes, and lengths of variant C-terminal domains (for the AE3-14nt variants) in the rightmost boxes. Modified from Stewart et al. (Stewart et al., 2007).

Fig. 2.

Proposed topographical model for the human SLC4A1/AE1 Cl^–/HCO₃^– exchanger polypeptide, after Zhu et al. (Zhu et al., 2003). Met66 (arrow) marks the start of kidney AE1. Polymorphisms encoding blood group antigens are blue. The mutations associated with hereditary spherocytic anemia and ovalocytosis are orange, and include missense, nonsense, splicing and deletion mutations. Missense mutations associated with hereditary stomatocytosis and xerocytosis are red. Mutations associated with dominant and recessive distal renal tubular acidosis are green. Terminal deletions are in lighter orange and green. Upper left: scanning electron micrographs of wild-type erythrocytes and AE1^–/– bovine spherocytes (HS) (Inaba et al., 1996). Upper right: consecutive semithin sections from rat kidney cortex immunostained with antibodies recognizing vH⁺-ATPase (left) and kAE1 (right). Only the Type A intercalated cell with apical vH⁺-ATPase expresses basolateral kAE1 (Alper et al., 1989). HS, hereditary spherocytic anemia; HSt, hereditary stomatocytosis; dRTA, distal renal tubular acidosis. Scale bars 10 μm at top left; 7 μm, top right. Modified from Shayakul and Alper, and Stewart (Shayakul and Alper, 2004; Stewart et al., 2007).

Fig. 3.

Model of mouse AE2a NH₂-terminal cytoplasmic domain amino acids 317–623, highlighting conserved residues required for normal regulation of Cl^–/anion exchange by pH_o and pH_i. (A) Ribbon diagram structure of AE2 amino acids 317–623 based on the crystal structure of the corresponding region of human AE1 (Zhang et al., 2000). The structural model (B) and the linear schematic diagram (C) each indicate residues which when mutated alter AE2 regulation by pH_i (yellow), by pH_o (red) or by both pH_i and pH_o (orange). (B) Space-filling structure of AE2 amino acids 317–610, with surface amino acid residues indicated by the same colors. P610 (blue) is the most C-terminal surface residue in this view. Mutation en bloc of AE2 amino acids 403–408, at the bottom in pink, altered sensitivity only to pH_o. AE2 amino acids 397–402 are located out of view at the bottom right, adjacent to amino acids 403–408. L323 is modeled to be not at the domain surface. (Modified from Stewart et al., 2004.)

Fig. 4.

Re-entrant loop 1 (RL1) of the mouse AE2 transmembrane domain plays a critical role in the acute regulation of anion exchange by pH. Summary of transmembrane subdomains (shaded boxes) and individual amino acid residues identified from mutagenesis studies as contributing importantly to regulation of AE2 activity by pH, NH₄⁺ and calmidazolium. Residues of RL1 interact with as yet unidentified amino acids within the TM1–6 region to mediate `pH sensor' functions in the AE2 transmembrane domain. Residues involved in regulation by pH_o are gray, those involved in regulation by pH_i are white, and those involved in regulation by both pH_o and pH_i are black. White boxes marked with an X are residues that when mutated yielded functional activity too low for study (modified from Stewart et al., 2008).