title: Identification of the hair cell soma-1 antigen, HCS-1, as otoferlin.
abstract: Hair cells, the mechanosensitive receptor cells of the inner ear, are critical for our senses of hearing and balance. The small number of these receptor cells in the inner ear has impeded the identification and characterization of proteins important for hair cell function. The binding specificity of monoclonal antibodies provides a means for identifying hair cell-specific proteins and isolating them for further study. We have generated a monoclonal antibody, termed hair cell soma-1 (HCS-1), which specifically immunolabels hair cells in at least five vertebrate classes, including sharks and rays, bony fish, amphibians, birds, and mammals. We used HCS-1 to immunoprecipitate the cognate antigen and identified it as otoferlin, a member of the ferlin protein family. Mutations in otoferlin underlie DFNB9, a recessive, nonsyndromic form of prelingual deafness characterized as an auditory neuropathy. Using immunocytochemistry, we find that otoferlin is associated with the entire basolateral membrane of the hair cells and with vesicular structures distributed throughout most of the hair cell cytoplasm. Biochemical assays indicate that otoferlin is tightly associated with membranes, as it is not solubilized by alterations in calcium or salt concentrations. HCS-1 immunolabeling does not co-localize with ribeye, a constituent of synaptic ribbons, suggesting that otoferlin may, in addition to its proposed function in synaptic vesicle release, play additional roles in hair cells.
authors.affiliation: School of Life Sciences, University of Sussex, Falmer, Brighton, BN1 9QG, UK. r.j.goodyear@sussex.ac.uk
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Goodyear
authors.forename: P Kevin
authors.initials: PK
authors.lastname: Legan
authors.forename: Jeffrey R
authors.initials: JR
authors.lastname: Christiansen
authors.forename: Bei
authors.initials: B
authors.lastname: Xia
authors.forename: Julia
authors.initials: J
authors.lastname: Korchagina
authors.forename: Jonathan E
authors.initials: JE
authors.lastname: Gale
authors.forename: Mark E
authors.initials: ME
authors.lastname: Warchol
authors.forename: Jeffrey T
authors.initials: JT
authors.lastname: Corwin
authors.forename: Guy P
authors.initials: GP
authors.lastname: Richardson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D002645:Chickens; D003638:Deafness / Q000235:genetics / Q000378:metabolism; D004284:Dogfish; D005849:Gerbillinae; D006168:Guinea Pigs; D006199:Hair Cells, Auditory, Inner / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D023421:Models, Animal; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011892:Rana catesbeiana; D013572:Synaptic Vesicles / Q000378:metabolism; D015027:Zebrafish
chemical_list: D000911:Antibodies, Monoclonal; D008565:Membrane Proteins
references.citation: J Med Genet. 2002 Jul;39(7):502-6
references.pmid: 12114484
references.citation: Hum Mol Genet. 2009 Aug 1;18(15):2779-90
references.pmid: 19417007
references.citation: Genes Dev. 2007 Sep 1;21(17):2118-30
references.pmid: 17785522
references.citation: J Neurosci. 1999 Jan 1;19(1):119-32
references.pmid: 9870944
references.citation: Cell. 2006 Oct 20;127(2):277-89
references.pmid: 17055430
references.citation: Dev Biol. 2006 Jun 1;294(1):148-60
references.pmid: 16566913
references.citation: Proc Natl Acad Sci U S A. 1995 Oct 24;92(22):10297-301
references.pmid: 7479771
references.citation: Hum Mutat. 2007 Jun;28(6):571-7
references.pmid: 17301963
references.citation: Hum Mutat. 2003 Dec;22(6):451-6
references.pmid: 14635104
references.citation: Int J Dev Neurosci. 1997 Jul;15(4-5):417-32
references.pmid: 9263023
references.citation: Proc Natl Acad Sci U S A. 2005 Feb 1;102(5):1496-501
references.pmid: 15677337
references.citation: Nat Rev Mol Cell Biol. 2008 Feb;9(2):99-111
references.pmid: 18216767
references.citation: Neurobiol Dis. 2002 Jul;10(2):157-64
references.pmid: 12127154
references.citation: J Comp Neurol. 2007 Feb 1;500(4):646-57
references.pmid: 17154269
references.citation: J Neurosci. 1997 Aug 15;17(16):6289-301
references.pmid: 9236239
references.citation: J Med Genet. 2006 Jul;43(7):576-81
references.pmid: 16371502
references.citation: J Neurosci. 2006 Oct 11;26(41):10438-51
references.pmid: 17035528
references.citation: J Comp Neurol. 2000 Jan 31;417(1):1-16
references.pmid: 10660884
references.citation: J Neurosci. 2006 Jun 28;26(26):7022-34
references.pmid: 16807332
references.citation: Brain Res Brain Res Protoc. 2002 Feb;9(1):65-76
references.pmid: 11852272
references.citation: J Biol Chem. 2009 Jan 16;284(3):1364-72
references.pmid: 19004828
references.citation: J Neurosci. 2006 Jun 14;26(24):6543-53
references.pmid: 16775142
references.citation: Development. 2005 Feb;132(3):541-51
references.pmid: 15634704
references.citation: Hear Res. 2007 Dec;234(1-2):21-8
references.pmid: 17967520
references.citation: J Neurosci. 2003 Jun 15;23(12):4878-87
references.pmid: 12832510
references.citation: Biochim Biophys Acta. 2006 Aug;1761(8):838-49
references.pmid: 16945584
references.citation: J Neurosci. 2008 Feb 27;28(9):2261-73
references.pmid: 18305259
references.citation: Am J Med Genet A. 2005 Sep 15;138(1):6-10
references.pmid: 16097006
references.citation: J Neurosci. 2007 Feb 28;27(9):2163-75
references.pmid: 17329413
references.citation: J Neurobiol. 2002 Feb 5;50(2):81-92
references.pmid: 11793356
references.citation: J Cell Biol. 1990 Apr;110(4):1055-66
references.pmid: 2182645
references.citation: J Med Genet. 2003 Jan;40(1):45-50
references.pmid: 12525542
references.citation: J Comp Neurol. 1999 Apr 5;406(2):183-98
references.pmid: 10096605
references.citation: Cell. 1988 Feb 12;52(3):329-41
references.pmid: 2964276
references.citation: J Neurosci. 2003 Oct 8;23(27):9208-19
references.pmid: 14534255
references.citation: J Neurosci. 2007 Mar 21;27(12):3174-86
references.pmid: 17376979
references.citation: Proc Natl Acad Sci U S A. 2007 Oct 16;104(42):16675-80
references.pmid: 17895386
references.citation: Trends Cell Biol. 2004 Apr;14(4):206-13
references.pmid: 15066638
references.citation: J Neurosci. 2008 Feb 20;28(8):1798-803
references.pmid: 18287496
references.citation: Am J Hum Genet. 2000 Sep;67(3):591-600
references.pmid: 10903124
references.citation: Clin Genet. 2009 Mar;75(3):237-43
references.pmid: 19250381
references.citation: Neuron. 2005 Nov 3;48(3):403-15
references.pmid: 16269359
references.citation: J Comp Neurol. 2005 Feb 28;483(1):91-113
references.pmid: 15672394
references.citation: Hear Res. 1989 Nov;42(2-3):175-94
references.pmid: 2606802
references.citation: J Neurosci. 2010 Sep 15;30(37):12545-56
references.pmid: 20844149
references.citation: Nat Genet. 1999 Apr;21(4):363-9
references.pmid: 10192385
references.citation: J Neurosci. 2001 Jul 15;21(14):5066-78
references.pmid: 11438582
references.citation: J Assoc Res Otolaryngol. 2000 Sep;1(2):172-82
references.pmid: 11545144
references.citation: Eur J Neurosci. 2006 Dec;24(12):3372-80
references.pmid: 17229086
references.citation: J Neurocytol. 1996 Apr;25(4):257-66
references.pmid: 8793731
references.citation: Neuron. 2000 Dec;28(3):857-72
references.pmid: 11163272
references.citation: Proc Natl Acad Sci U S A. 2006 Dec 5;103(49):18615-20
references.pmid: 17124170
references.citation: Nature. 1989 Oct 5;341(6241):397-404
references.pmid: 2677742
references.citation: J Neurosci. 1999 May 15;19(10):3761-72
references.pmid: 10234008
references.citation: J Comp Neurol. 1992 Nov 8;325(2):243-56
references.pmid: 1281174
references.citation: J Comp Neurol. 1991 Dec 22;314(4):777-88
references.pmid: 1816275
references.citation: J Comp Neurol. 2005 Mar 28;484(1):105-20
references.pmid: 15717301