RT Journal Article
SR Electronic
T1 Otoferlin Couples to Clathrin-Mediated Endocytosis in Mature Cochlear Inner Hair Cells
JF The Journal of Neuroscience
JO J. Neurosci.
FD Society for Neuroscience
SP 9508
OP 9519
DO 10.1523/JNEUROSCI.5689-12.2013
VO 33
IS 22
A1 Susanne V. Duncker
A1 Christoph Franz
A1 Stephanie Kuhn
A1 Uwe Schulte
A1 Dario Campanelli
A1 Niels Brandt
A1 Bernhard Hirt
A1 Bernd Fakler
A1 Nikolaus Blin
A1 Peter Ruth
A1 Jutta Engel
A1 Walter Marcotti
A1 Ulrike Zimmermann
A1 Marlies Knipper
YR 2013
UL http://www.jneurosci.org/content/33/22/9508.abstract
AB The encoding of auditory information with indefatigable precision requires efficient resupply of vesicles at inner hair cell (IHC) ribbon synapses. Otoferlin, a transmembrane protein responsible for deafness in DFNB9 families, has been postulated to act as a calcium sensor for exocytosis as well as to be involved in rapid vesicle replenishment of IHCs. However, the molecular basis of vesicle recycling in IHCs is largely unknown. In the present study, we used high-resolution liquid chromatography coupled with mass spectrometry to copurify otoferlin interaction partners in the mammalian cochlea. We identified multiple subunits of the adaptor protein complex AP-2 (CLAP), an essential component of clathrin-mediated endocytosis, as binding partners of otoferlin in rats and mice. The interaction between otoferlin and AP-2 was confirmed by coimmunoprecipitation. We also found that AP-2 interacts with myosin VI, another otoferlin binding partner important for clathrin-mediated endocytosis (CME). The expression of AP-2 in IHCs was verified by reverse transcription PCR. Confocal microscopy experiments revealed that the expression of AP-2 and its colocalization with otoferlin is confined to mature IHCs. When CME was inhibited by blocking dynamin action, real-time changes in membrane capacitance showed impaired synaptic vesicle replenishment in mature but not immature IHCs. We suggest that an otoferlin-AP-2 interaction drives Ca2+- and stimulus-dependent compensating CME in mature IHCs.