Plasmalemmal Na+-independent Cl-/HCO3- exchangers regulate intracellular pH, [Cl-], and cell volume. In polarized epithelial cells they contribute also to transepithelial secretion and reabsorption of acid-base equivalents and of Cl-. These transporters are encoded by members of the two evolutionarily unrelated mammalian gene families, SLC4 and SLC26. Human SLC4A1/AE1 mutations cause either the erythroid disorders spherocytic hemolytic anemia or ovalocytosis, or distal renal tubular acidosis. SLC4A2/AE2 knockout mice die at weaning. Human SLC4A3/AE3 polymorphisms have been associated with seizure disorder. Mutations in 4 of the human SLC26 genes cause chondrodysplasias, congenital diarrhea, deafness, and hypothyroid goiter. Although mammalian SLC4/AE polypeptides mediate only electroneutral Cl-/anion exchange, the structural and mechanistic differences between electroneutral and electrogenic anion transport can be subtle. Thus, the electroneutral trout AE1 anion exchanger also promotes or mediates increased anion conductance and uncoupled osmolyte transport. The presence of the electroneutral anion exchanger mouse AE1 in the erythrocyte membrane is required for DIDS-sensitive erythroid Cl- conductance (although definitive evidence for AE1 mediation of Cl- conductance is lacking). A single missense mutation allows mouse AE1 to mediate both electrogenic SO42-/Cl- exchange or electroneutral, H+-independent SO42-/ SO42- exchange. SLC4 anion exchangers appear to be homooligomers, but also function with the assistance of auxiliary subunits. Thus, glycophorin A and protein 4.2 act as subunits of erythroid AE1 affecting trafficking and transport rate. Depending on the expression system, direct binding of carbonic anhydrases bySLC4 anion exchangers is either stimulatory or obligatory for HCO3- transport. In the Xenopus oocyte, the AE1 C-terminal cytoplasmic tail residues thought to bind carbonic anhydrase II are dispensable for Cl-/Cl- exchange, but required for Cl-/HCO3- exchange. Membrane-associated ecto-carbonic anhydrases also bind to SLC4 anion exchangers and modulate their activity. Cellular pH regulation by SLC4 anion exchangers implies that anion exchanger activity is itself pH-sensitive. However, SLC4/AE1 is remarkably insensitive to pH over the physiological range, whereas SLC4A2/AE2 is acutely and independently inhibited by intracellular and extracellular H+. This regulation requires integrity of the most highly conserved sequence of the AE2 N-terminal cytoplasmic domain, along with adjacent residues in which individual missense mutations acid-shift pHo sensitivity of AE2. These regulatory determinants together are modeled to form contiguous surface patches on the AE2 cytoplasmic domain. In contrast, the N-terminal variant AE2c polypeptide exhibits an alkaline-shifted pHo-sensitivity, as do certain transmembrane domain His mutants. A novel, non-conserved region of the AE2 transmembrane domain has been recently shown to be required for pH-sensitivity. AE2-mediated anion exchange is also stimulated by ammonium and hypertonicity via Ca2+-dependent pathways. In contrast to SLC4 anion exchangers, the SLC26 anion transporter polypeptides exhibit remarkable sequence diversity among near-species orthologs, most marked among SLC26A6 polypeptides. This sequence diversity prompted systematic functional comparison in Xenopus oocytes of mouse slc26a6 with human SLC26A6 variants. Whereas these orthologs exhibited similar rates of bidirectional 14C-oxalate flux and of Cl-/HCO3- and Cl-/OH- exchange subject to similar stimulation by CFTR, they differed markedly in rates of Cl- exchange for trans-Cl-, sulphate, and formate. Studies with mouse-human chimera slc26a6 cRNAs showed that high transport rates cosegregated with the transmembrane domain of mouse slc26a6, and that multiple parts of the transmembrane domain contribute to the phenotype. We found that both species orthologs mediated electroneutral Cl-/HCO3- and Cl-/OH- exchange as measured by two electrode voltage clamp. In contrast, Cl-/oxalate exchange by mouse slc26a6 was electrogenic. Isotopic flux studies performed under voltage clamp conditions confimed these observations. Although studies of mouse SLC26 knockout models are revealing physiological roles of these transporters in secretory processes for bicarbonate and other substrate anions, the substantial sequence differences between orthologous mouse and human transporter polypeptides correlate with species-specific differences in transport properties. These intrinsic differences must be considered when extrapolating results from the mouse to human pathophysiology. Delineation of the physical basis for the unusual species-specific differences in anion selectivity of slc26a6 polypeptides should also provide general insight into structure-function relationships of anion binding and translocation.