Abstract
Renal Rhesus type B glycoprotein (Rhbg) is a glycosylated mammalian NH(3)/NH(4)(+) transporter expressed in α-intercalated cells of the collecting duct. Carbonic anhydrase-IV (CA-IV) is also expressed in the mammalian kidney, where it catalyzes the reversible hydration of CO(2). This study aims to demonstrate: 1) whether Rhbg and CA-IV proteins physically interact; and 2) if this interaction functionally affects transport of NH(3)/NH(4)(+) and possibly CO(2). We measured transport of NH(4)(+), NH(3), and CO(2) in four groups of Xenopus oocytes. In the first group, we coexpressed Rhbg with CA-IV and compared the measurements to three groups of oocytes expressing either Rhbg or CA-IV or injected with H(2)O. We used ion-selective microelectrodes to measure surface pH, to monitor NH(3) transport, and intracellular pH to monitor NH(4)(+) and CO(2) transport. We also used a two-electrode voltage clamp to measure current changes caused by electrogenic NH(4)(+) transport. These parameters measured NH(3)/NH(4)(+) and CO(2) transport in oocytes expressing Rhbg and/or CA. Our results indicate that: 1) Rhbg and CA-IV were coimmunoprecipitated, suggesting a physical interaction; and 2) coexpressing CA-IV with Rhbg: i) inhibited electrogenic NH(4)(+) transport by Rhbg in the presence and absence of CO(2); ii) reduced NH(3) transport by Rhbg only in the presence of CO(2); and iii) had no detectable effect on CO(2) transport by Rhbg. We demonstrated for the first time that Rhbg and CA-IV physically interact, and this interaction has inhibitory effects on Rhbg function but not CA-IV. The interaction of Rhbg and CA-IV is important to explain their role in renal acid-base homeostasis.NEW & NOTEWORTHY Our study revealed the complex regulation of NH(3)/NH(4)(+) transport, highlighting the roles of Rhbg, CA-IV, and environmental factors such as CO(2) concentration. These interactions are critical to our understanding of NH(3)/NH(4)(+) transport and regulation. Our findings lay a strong foundation for future investigations into the molecular dynamics among these transport proteins and their physiological significance. These studies are essential to fully understand how these mechanisms influence renal ammonia handling, urinary acidification, and systemic pH balance.