Trans-dominant inhibition of connexin-43 by mutant connexin-26: implications for dominant connexin disorders affecting epidermal differentiation

Dominant mutations of GJB2-encoding connexin-26 (Cx26) have pleiotropic eff ects, causing either hearing impairment (HI) alone or in association with p almoplantar keratoderma (PPK/HI). We examined a British family with the lat ter phenotype and identified a new dominant GJB2 mutation predicted to elim inate the amino acid residue E42 (Delta E42) in Cx26. To dissect the pathom echanisms that result in diverse phenotypes of dominant GJB2 mutations, we studied the effect of three Cx26 mutants (Delta E42, D66H and R75W) identif ied in individuals with PPK/HI, and another (W44C) present in individuals w ith non-syndromic HI on gap junctional intercellular communication. We expr essed mutant Cx26 alone and together with the epidermal connexins Cx26, Cx3 7 and Cx43 in paired Xenopus oocytes, and measured the intercellular coupli ng by dual voltage clamping. Homotypic expression of each connexin as well as co-expression of wild-type (wt) Cx26/wtCx43 and wtCx26/wtCx37 yielded va riable, yet robust, levels of channel activity. However, all four Cx26 muta nts were functionally impaired and failed to induce intercellular coupling. When co-expressed with wtCx26, all four mutants suppressed the wtCx26 chan nel activity consistent with a dominant inhibitory effect. However, only th ose Cx26 mutants associated with a skin phenotype also significantly (P<0.0 5) inhibited intercellular conductance of co-expressed wtCx43, indicating a direct interaction of mutant Cx26 units with wtCx43. These results demonst rate, for the first time, a trans-dominant negative effect of Cx26 mutants in vitro. Furthermore, they support a novel concept suggesting that the pri ncipal mechanism for manifestation of dominant GJB2 mutations in the skin i s their dominant interference with the function of wtCx43. This assumption is further corroborated by our finding that Cx26 and Cx43 focally colocaliz e at gap junctional plaques in affected skin tissue of two carriers of <Del ta>E42.