The team also analyzed the amino acid sequences of protocadherin-15 and discovered four distinct forms -- three of which are present in various developmental stages of the mouse inner ear. The researchers refer to the three alternative forms found in the inner ear as CD1, CD2, and CD3 because the sequential variations occur in the protein's "cytoplasmic domain" a stretch of amino acids anchored inside the stereocilium. (The fourth form, referred to as SI, is likely to be secreted.) With the help of two imaging techniques that use antibodies to label a targeted protein, the team found that the distribution of protocadherin-15 along the stereocilium varies by form, with the CD3 form stationed only at the tips of the stereocilia in mature hair cells, while the CD1 form is found along the lengths of the stereocilia in mature cells, but not at the tips. In contrast, the CD2 form is expressed along the lengths of stereocilia during hair cell development, but is not present in mature hair cells.
Finally, the team found that a chemical known to break tip links called BAPTA had no effect on the CD1 and CD2 forms of protocadherin-15 but destroyed the CD3 form. Likewise, just as tip links are known to reappear roughly four hours after the chemical is removed, the CD3 form returned within four to 24 hours upon removal of the chemical.
Based on these findings, the researchers conclude that, not only is protocadherin-15 now identified as the tip-link antigen, but it is
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Contact: Jennifer Wenger
jwenger@mail.nih.gov
301-496-7243
NIH/National Institute on Deafness and Other Communication Disorders
27-Jun-2006