The ability of KCNQ (Kv7) channels to form hetero-oligomers is of high physiological importance. We have recently identified a carboxy-terminal subunit interaction (si) domain that determines their subunit-specific assembly. Within this si domain, there are two motifs which comprise about 30 amino acids residues each and which exhibit a high probability for coiled-coil formation. Transfer of the first or the second coiled-coil (TCC) domain from KCNQ3 into the KCNQ1 scaffold resulted in chimeras KCNQ1(TCC1)Q3 and KCNQ1(TCC2)Q3, both of which co-immunoprecipitated with KCNQ2. However, only KCNQ1(TCC2)Q3 enhanced KCNQ2 currents and surface expression or exerted a strong dominant negative effect on KCNQ2. Deletion of TCC2 within KCNQ2 yielded functional homomeric channels, but prevented the current augmentation measured upon co-expression of KCNQ2 and KCNQ3. In contrast, deleting TCC1 within KCNQ2 did not give functional homomeric KCNQ2 or heteromeric KCNQ2/KCNQ3 channels. Our data suggest that the TCC1 domains of KCNQ2 and KCNQ3 are required to form functional homo- as well as heteromeric channels, whereas both TCC2 domains facilitate an efficient transport of heteromeric KCNQ2/KCNQ3 channels to the plasma membrane.