- Otoferlin is a protein that is essential in the process of hearing. Mutations in otoferlin are associated with profound recessive deafness. The long-term goal of the Johnson lab is to understand how otoferlin contributes to hearing and why certain mutations result in deafness. This project sought to characterize the difference between wildtype and mutant otoferlin in vitro to go alongside in vivo studies on mutant zebrafish presenting with deafness and vestibular effects. The wildtype otoferlin was cloned from zebrafish DNA, and site directed mutagenesis introduced an early stop codon in the transmembrane domain to create the mutant otoferlin. Preliminary immunofluorescence results indicate that wildtype otoferlin appears to associate with the membrane to a greater extent than mutant, the mutant otoferlin traffics differently in the cell, and is possibly degraded faster than wild type. These results and future studies will further our understanding of the specific role of otoferlin in the process of hearing and why this particular nonsense mutation results in deafness.
- Key Words: otoferlin, ferlins, zebrafish, HEK293, vesicle trafficking