The human Rhesus-associated RhAG protein and a kidney homologue promote ammonium transport in yeast

Abstract

The Rhesus blood-group antigens are defined by a complex association of membrane polypeptides that includes the non-glycosylated Rh proteins (RhD and RhCE) and the RHag glycoprotein, which is strictly required for cell surface expression of these antigens. RhAG and the Rh polypeptides are erythroid-specific transmembrane proteins belonging to the same family (36% identity). Despite their importance in transfusion medicine, the function of RhAG and Rh proteins remains unknown, except that their absence in Rh(null) individuals leads to morphological and functional abnormalities of erythrocytes, known as the Rh-deficiency syndrome. We recently found significant sequence similarity between the Rh family proteins, especially RhAG, and Mep/Amt ammonium transporters. We show here that RhAG and also RhGK, a new human homologue expressed in kidney cells only, function as ammonium transport proteins when expressed in yeast. Both specifically complement the growth defect of a yeast mutant deficient in ammonium uptake. Moreover, ammonium efflux assays and growth tests in the presence of toxic concentrations of the analogue methylammonium indicate that RhAG and RhGK also promote ammonium export. Our results provide the first experimental evidence for a direct role of RhAG and RhGK in ammonium transport. These findings are of high interest, because no specific ammonium transport system has been characterized so far in human.

Human Rhesus B and Rhesus C glycoproteins: properties of facilitated ammonium transport in recombinant kidney cells

“…A recent study also identified RhBG and RhCG as having a cell specific, axially heterogeneous and polarized expression in the gastrointestinal tract [13]. As is the case for RhAG, ammonium transport activity of RhCG was first shown by complementation studies in yeast [5] and then confirmed by functional studies in Xenopus oocytes [14]. RhBG-mediated ammonium transport was also recently demonstrated in the Xenopus oocyte expression system [15,16].…”

“…In RBCs (red blood cells) RhCE, RhD and RhAG constitute the core of the Rh-membrane complex, the carrier of the Rh bloodgroup antigens [3,4]. Genetic complementation studies in Mepdeficient yeast provided the first experimental evidence for a direct role of RhAG in ammonium transport [5]. RhAG-dependent ammonium transport in a heterologous system was subsequently demonstrated in injected Xenopus oocytes [6].…”

“…Since RhAG expression is erythroid-restricted, this glycoprotein could not account for ammonium transport in the two organs which play a key role in ammonium metabolism and excretion: liver and kidney. However, two non-erythroid RhAG homologues, RhBG and RhCG, have been characterized and found to be expressed in various tissues including the kidney and the liver [5,7,8].…”

The Structure and Function of the Rh Antigen Complex

“…[5][6][7] The erythrocyte Rh blood group proteins are well known because of their importance in blood transfusion, but recent functional studies and structural modeling reveal that the Rh blood group proteins are members of an ancient family of proteins involved in ammonia transport. [8][9][10][11] Non-erythroid Rh proteins have now been found in other tissues including the kidney, liver, brain, and skin 12-15 , in locations where ammonia production and elimination occurs. The family of Rh proteins has now been expanded significantly through comparative genomics and structure-function studies that reveal the presence of Rh homologs in all domains of life.…”

Amt/MEP/Rh proteins conduct ammonia

“…The RhD, RhCE, and RhAG proteins form a heteromeric, most likely heterotrimeric, complex but only RhAG has been shown to function in ammonium transport [27,48]. RhD and RhCE differ in some otherwise highly conserved pore residues.…”