EAST/SeSAME syndrome mutations disrupt function of heteromeric Kir2.3/Kir4.1 channels at the basolateral membrane of the distal nephron

Abstract

Basolateral inwardly rectifying K channels (Kir) in the distal nephron are essential for salt and mineral reabsorption. Mutations in Kir 4.1 cause profound renal salt wasting (EAST/SeSAME syndrome), suggesting that Kir 4.1 plays a dominant role through assembly with the other Kir subunits, like Kir 5.1. Here, we report that Kir2.3 and Kir4.1 co-immunoprecipitate when co-expressed in Cos cells, and co-localize in the mouse distal nephron. As studied in Xenopus oocytes, co-expression of Kir 2.3 with Kir4.1 potentiated K current density and increased cell surface expression. A dominant negative mutation in the Kir4.1 blocked Kir2.3 activity, and a comparable mutation in Kir2.3 inhibited Kir4.1, indicating heteromeric assembly. At the single channel level, Kir2.3/4.1 channels exhibit properties different than Kir2.3 or Kir4.1 alone, more similar to the native channel in the CCD basolateral membrane. Interestingly, of the 6 studied EAST/SeSAME syndrome mutations, 4 of them (C140R, T164I, R297C, R199X) reduced Kir4.1/5.1 function, and 3 of them (R65P, C140R, T164I) exerted a similar effect on Kir2.3. In conclusion: 1) basolateral K channels of the distal nephron can be formed through heteromeric assembly of Kir4.1/5.1 or Kir4.1/2.3 subunits; 2) Consequently, the inhibitory effects of EAST/SeSAME syndrome mutations can be broadly transmitted throughout the basolateral membrane conductance. Funds from NIH & AHA.