Clonal somato-mammotroph GH3/B6 cells and lactotroph MMQ cells express two (ergl, erg2) of the three cloned rat ether-à-go-go-related gene (erg) K channel subunits. To study whether the erg subunits form heteromultimers, dominant-negative mutants of erg and erg2 were constructed by point mutation (erg1G630S, erg2G480S). After co-expression of these mutants with the wild-type erg1, erg2, or erg3 in Chinese hamster ovary (CHO) cells no erg currents could be detected. In contrast, in co-expression experiments with members of the other ether-à-go-go (EAG) subfamilies (eagl, elkl) the mutant erg1G630S had no effect. These results strongly suggest that erg channel subunits are able to form heteromultimers within the erg channel subfamily. Suppression of the endogenous E-4031-sensitive currents in GH3/B6 and MMQ cells by erg1G630S confirms that they are mediated by erg channels despite the differences in gating kinetics in these cells. Reduction of the erg current in GH3/B6 cells by erg2G480S indicates that erg heteromultimers can also be formed in these cells.