Southeast Asian AE1 associated renal tubular acidosis: Cation leak is a class effect

Abstract

Anion Exchanger 1 (AE1) is present in the erythrocyte and also in the α-intercalated cell; different mutations can cause either red cell disease or distal renal tubular acidosis (dRTA). Recently, we described a cation leak property in four dRTA-causing AE1 mutants, three autosomal dominant (AD) European mutants, one autosomal recessive (AR) from Southeast Asia, G701D. G701D had a very large leak property and is unusually common in SE Asia. We hypothesized that this property might confer a survival advantage. We characterized three other AR dRTA-associated AE1 mutants found in SE Asia, S773P, Δ850 and A858D via transport experiments in AE1-expressing Xenopus oocytes. These three SE Asian mutants also had cation leaks of similar magnitude to that seen in G701D, a property that distinguishes them as a discrete group. The clustering of these cation-leaky AE1 mutations to malarious areas of SE Asia suggests that they may confer malaria resistance.

Introduction

The anion exchanger, AE1 (or Band 3), encoded by the gene SLC4A1, is a membrane bound glycoprotein that is predicted to span the cell membrane 12–14 times [1]. It mediates the electroneutral exchange of chloride for bicarbonate across the cell membrane, and is present chiefly in the erythrocyte, (where it is highly expressed, at approximately 1 × 10⁶ copies per cell), and in the α-intercalated cell of the kidney.

Its function in the red cell is to reclaim bicarbonate from the red cell in the peripheral circulation after CO₂ absorption by the erythrocyte. An isoform, truncated by 65 amino acids at the N-terminal end of the protein is present in the basolateral membrane of the acid-secreting α-intercalated cell [2], to reclaim bicarbonate back into the systemic circulation following the hydration of CO₂ catalyzed by carbonic anhydrase in the intercalated cell interior.

Distinct mutations of the SLC4A1 gene may cause disease of either the red cell or the kidney. Certain mutations of SLC4A1 can cause red cell diseases such as hereditary spherocytosis (HS) [3], hereditary stomatocytosis (HSt) [4] and Southeast Asian ovalocytosis (SAO) [5]. In these last two diseases, the defect is often functional-the AE1 protein having reduced efficiency at anion exchange when expressed in Xenopus oocytes [4], [5].

Other mutations can cause kidney disease, namely distal renal tubular acidosis (dRTA), a failure of the α-intercalated cell resulting in faulty urinary acidification, resulting in varying degrees of systemic acidosis, renal stone formation, bone demineralisation and hypokalemia. These mutations usually result in proteins which have normal anion exchange activity when expressed in Xenopus oocytes, but are mistargetted in polarized mammalian cell models [6].

There are two phenotypes of AE1 associated dRTA; the rare European autosomal dominant (AD) dRTA, where one mutant allele will generate a mutant protein that is able to ‘capture’ a wild-type protein in the cytosol when heterodimerized with it, when expressed in a polarized cell model [7]. The other is comprised of the commoner Southeast Asian autosomal recessive (AR) dRTA, in which one wild-type allele is capable of ‘rescuing’ a mutant protein when heterodimerized with it, resulting in cell membrane expression of the heterodimer [7], [8].

More recent work by our group has shown that when expressed in Xenopus laevis oocytes, dRTA-associated AE1 mutants are ‘cation leaky’. This effect is demonstrable in the three European AD mutants that we characterized (R589H, G609R and S613F), but much greater in G701D, the only SE Asian AR mutant that we examined [9].

As this disparity between the European and Asian mutant seemed so large, we decided to characterize three other described SE Asian mutants (S773P, Δ850 and A858D) [10], [11], [12]., alongside the G701D mutant, using the X. laevis expression system. Our results show that these three other AE1 mutations also induce a cation leak in a still functional anion exchanger. The effect of these mutations on the anion exchange structure of the protein is then discussed as the consequences of this cation leak in cells expressing AE1.