The Ankle-link Antigen: an Epitope Sensitive to Calcium Chelation Associated with the Hair-cell Surface and the Calycal Processes of Photoreceptors

A monoclonal antibody, mAb E40, that specifically recognizes hair cells and photoreceptors was derived from a mouse immunized with a membrane fraction prepared from the sensory maculae of the chick inner ear. In the mature chick inner ear, punctate labeling is observed along each stereocilium, but staining is mostly concentrated around the basal end of the sensory hair bundles, where it is closely associated with surface specializations known as ankle links. The epitope recognized by mAb E40 is therefore referred to as the ankle-link antigen (ALA). During early embryogenesis, the ALA is initially distributed evenly over the surface of the hair bundle. As development proceeds, it becomes more restricted to the base of the hair bundle, although a spot of the ALA remains associated with the bundle tip until just before hatching. In the eye, mAb E40 stains the calycal processes of photoreceptors. When maculae and retinae are treated with the calcium chelator BAPTA at room temperature, the ALA disappears. BAPTA-induced loss of the ALA from the hair-bundle surface is substantially reduced by lowering the temperature to 2 degrees C. The ALA and ankle links reappear on the hair-bundle surface when cells are cultured for 20 hr after BAPTA treatment. BAPTA sensitivity and recovery after BAPTA-induced loss are properties similar to those described for the tip link, a surface structure thought to gate the mechanotransducer channel. However, unlike the tip link, the ALA and ankle links are sensitive to subtilisin treatment. The results define a new component of the hair-bundle surface, with properties both common to and distinct from those of the tip link.