Abstract
We have performed a proteomic analysis of the inner ear proteins using 2D-GE. In the process of analysis, we have found very unique properties of the LOCH gene product. The COCH gene mutated in DFNA9, an autosomal dominant hereditary sensorineural hearing loss and vestibular disorder, encodes cochlin. DFNA9 patients show symptoms such as episodes of vertigo, tinnitus, aural fullness and hearing loss. Clinically, these symptoms are consistent with the criteria for Meniere's disease. LOCH is the only gene identified so far whose mutation leads to the symptoms of Meniere's disease in a significant portion of the carriers.
We showed that Cochlin constitutes 70% of inner ear proteins, and identified three cochlin isoforms in the inner ear tissue, p63s, p44s and p40s, which exhibit significant molecular heterogeneity. Structure analysis of Cochlin isoforms showed that the mutations influence only the full-length isoform of Cochlin (p63s), and not the processed Cochlin isoforms (p44s and p40s), which do not contain the LCCL domain. What happens to the LCCL domain once it is cleaved from full-length Cochlin was an open issue.
We further characterized the expression and structure of Cochlin isoforms by iso-form-specific antibodies that recognize three distinct domains. Inner ear, as well as perilymph proteins were analyzed by western blot analysis. We have detected Cochlin isoforms in the inner ear tissue and we have identified a novel short 16 kDa isoform in the perilymph, named Cochlin-tomoprotein (CTP), corresponding to the N-terminus of full-length Cochlin (p63s) and the LCCL domain. Notably, CTP contains all of the known mutation sites associated with DFNA9. Our results on the formation and processing of these isoforms in the inner ear will be central to a better understanding of Cochlin function and its role in the pathophysiology of DFNA9.
Furthermore, using above mentioned results, we are now performing a translational research to improve diagnosis and prognosis in patients with sensorineural hear-ing loss and vestibular disorders.
