In Na-reabsorbing tight epithelia, the rate-limiting step for Na transport is the highly selective low-conductance amiloride-sensitive epithelial Na channel (type 1 ENaC). In rat distal colon, type 1 ENaC is made of three homologous subunits. The aim of this study was to identify the corresponding genes of the renal channel from the kidney-derived A6 cell line of Xenopus laevis. Three homologous subunits were identified and coexpressed in the Xenopus oocyte system. The reconstituted channel had all the characteristics of the native type 1 ENaC described in A6 cells: 1) high selectivity, 2) low single-channel conductance, 3) slow gating kinetics, and 4) high affinity for amiloride. Transcripts for alpha-, beta-, and gamma-subunits of the Xenopus epithelial Na channel (xENaC) were detected in A6 kidney cells, Xenopus kidney, lung, and to a lesser extent in stomach and skin. Each subunit of the xENaC shares approximately 60% overall identity with the corresponding rat homologue (alpha, beta, and gamma rENaC). Our data suggest that the triplication of the ENaC subunits occurred before the divergence between mammalian and amphibian lineages.