Dominant connexin26 mutants associated with human hearing loss have trans-dominant effects on connexin30.

Dominant mutations in GJB2, the gene encoding the human gap junction protein connexin26 (Cx26), cause hearing loss. We investigated whether dominant Cx26 mutants interact directly with Cx30. HeLa cells stably expressing nine dominant Cx26 mutants, six associated with non-syndromic hearing loss (W44C, W44S, R143Q, D179N, R184Q and C202F) and three associated with hearing loss and palmoplantar keratoderma (G59A, R75Q and R75W), individually or together with Cx30, were analyzed by immunocytochemistry, co-immunoprecipitation, and functional assays (scrape-loading and/or fluorescence recovery after photobleaching). When expressed alone, all mutants formed gap junction plaques, but with impaired intercellular dye transfer. When expressed with Cx30, all mutants co-localized and co-immunoprecipitated with Cx30, indicating they likely co-assembled into heteromers. Furthermore, 8/9 Cx26 mutants inhibited the transfer of neurobiotin or calcein, indicating that these Cx26 mutants have trans-dominant effects on Cx30, an effect that may contribute to the pathogenesis of hearing loss.