Kir5.1 Is Essential to Formation of Kir4.2/Kir5.1 Heterotetramer in the Basolateral Membrane of Mouse Proximal Tubules

Background: Kir5.1 is encoded by Kcnj16 in the mouse kidney and it is expressed in the basolateral membrane of the proximal tubule (PT), distal-convoluted-tubule (DCT) and cortical- collecting-duct. In DCT, Kir5.1 interacts with Kir4.1 to form a 40-pS K+ channel, a main type of K+ channel in the basolateral membrane. Moreover, Kir4.1 can also form a homotetramer in the absence of Kir5.1 and provides the basolateral K+ conductance of DCT. In PT, Kir5.1 interacts with Kir4.2 to form Kir4.2/Kir5.1 heterotetramer, a main type basolateral K+ channel. However, it is not clear whether Kir4.2 along is sufficient for forming a Kir4.2 homotetramer without Kir5.1 in the basolateral membrane and maintains PT membrane conductancel. Methods: We now conduct immunofluorescence-staining and patch-clamp-experiments in Kir5.1-knockout (KO) mice and corresponding wild-type (WT) mice to examine the expression of Kir4.2 and to determine the basolateral K+ channel activity and PT membrane potential. Results: The single-channel-recording identified a 50-pS K+-channel in the basolateral membrane of PT of WT mice. This 50-pS K+-channel is a main type basolateral K+ channel and is most likely Kir4.2/Kir5.1 heterotetramer. However, this 50-pS inwardly-rectifying-K+-channel was completely absent in the PT of Kir5.1-KO mice. Moreover, patch-clamp-experiments failed to detect additional basolateral K+ channels in the PT of Kir5.1-KO mice. Immunofluorescence-staining shows that Kir4.2 is exclusively expressed in the PT. While the basolateral membrane staining of Kir4.2 was clearly visible in WT mice, it was largely absent in Kir5.1-KO mice. This is in sharp contrast to Kir4.1, which has a clear basolateral membrane staining in both WT and Kir5.1-KO mice. Also, the membrane potential of PT was more positive in Kir5.1-KO mice than WT mice, suggesting that deletion of Kir5.1 impairs Kir4.2 membrane expression. Immunoblotting also showed that the expression of Kir4.2 and Na/H exchanger-3 was lower in Kir5.1-KO mice than WT mice. Conclusion: We conclude that Kir5.1 is essential for forming the basolateral 50-pS K+-channel in the PT and plays a role in determining Kir4.2 basolateral membrane abundance. Also, Kir4.2 could not form a homeotetrammer in the absence of Kir5.1 and the membrane potential of PT is depolarized in the Kir5.1-KO mice. Funding: NIDDK Support