Regulatory interactions of the cystic fibrosis transmembrane conductance regulator (CFTR) and the epithelial Na⁺ channel (ENaC) are readily apparent in Xenopus oocytes. However, the mechanism underlying these interactions remains controversial. CFTR's first nucleotide binding fold (NBD-1) may be important in these interactions, as dysfunctional CFTRs containing mutations within NBD-1, such as F508 and G551D, lack such functional interactions with murine ENaC (mENaC). We hypothesized that a dysfunctional CFTR containing a non-NBD-1 mutation would retain regulatory interactions with mENaC, and tested this hypothesis for N1303K-CFTR where the mutation is located in CFTR's second nucleotide binding fold (NBD-2). cRNA for mENaC and N1303K-CFTR was injected separately or together into Xenopus oocytes. ENaC and CFTR functional expression were assessed by two-electrode voltage clamp. Injection of N1303K (class II trafficking mutation) yielded low levels of CFTR function upon activation with forskolin and isobutylmethlxanthine (IBMX). In co-injected oocytes, N1303K did not alter mENaC functional expression or surface expression before activation of N1303K. This is similar to our prior observations with F508. However, unlike our observations with F508, activation of N1303K acutely decreased mENaC functional and surface expression, and N1303K-currents were enhanced by co-injection of mENaC. Furthermore, genistein only mildly enhanced the functional expression of N1303K-CFTR, and did not improve regulation of ENaC by N1303K-CFTR. These data suggest that a structurally and functionally intact CFTR NBD-1 in activated CFTR can regulate mENaC surface expression independent of chloride transport in Xenopus oocytes.