Trafficking abnormality and ER stress underlie functional deficiency of hearing impairmentassociated connexin-31 mutants

Hearing impairment (HI) affects 1/1000 children and over 2% of the aged population. We have previously reported that mutations in the gene encoding gap junction protein connexin-31 (Cx31) are associated with HI. The pathological mechanism of the disease mutations remains unknown. Here, we show that expression of Cx31 in the mouse inner ear is developmentally regulated with a high level in adult inner hair cells and spiral ganglion neurons that are critical for the hearing process. In transfected cells, wild type Cx31 protein (Cx31wt) forms functional gap junction at cell-cell-contacts. In contrast, two HIassociated Cx31 mutants, Cx31R180X and Cx31E183K resided primarily in the ER and Golgi-like intracellular punctate structures, respectively, and failed to mediate lucifer yellow transfer. Expression of Cx31 mutants but not Cx31wt leads to upregulation of and increased association with the ER chaperone BiP indicating ER stress induction. Together, the HI-associated Cx31 mutants are impaired in trafficking, promote ER stress, and hence lose the ability to assemble functional gap junctions. The study reveals a potential pathological mechanism of HI-associated Cx31 mutations.