Sparc protein is required for normal growth of zebrafish otoliths.
Otoliths and the homologous otoconia in the inner ear are essential for balance. Their morphogenesis is less understood than that of other biominerals, such as bone, and only a small number of their constituent proteins have been characterized. As a novel approach to identify unknown otolith proteins, we employed shotgun proteomics to analyze crude extracts from trout and catfish otoliths. We found three proteins that had not been associated previously with otolith or otoconia formation: 'Secreted acidic cysteine rich glycoprotein' (Sparc), an important bone protein that binds collagen and Ca(2+); precerebellin-like protein, which contains a C1q domain and may associate with the collagenous otolin-1 during its assembly into a framework; and neuroserpin, a serine protease inhibitor that may regulate local protease activity during framework assembly. We then used the zebrafish to investigate whether Sparc plays a role in otolith morphogenesis. Immunodetection demonstrated that Sparc is a true constituent of otoliths. Knockdown of Sparc expression in morphant zebrafish resulted in four principal types of defective otoliths: smaller, extra and ectopic, missing and fused, or completely absent. Smaller size was the predominant phenotype and independent of the severity of otic-vesicle defects. These results suggested that Sparc is directly required for normal otolith growth.
Kang
Young-Jin
YJ
Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, 407 South Goodwin Avenue, MC-114, Urbana, IL 61801, USA.
Stevenson
Amy K
AK
Yau
Peter M
PM
Kollmar
Richard
R
D000367:Age Factors; D000595:Amino Acid Sequence; D000818:Animals; D005455:Fluorescent Antibody Technique; D018507:Gene Expression Regulation, Developmental; D013058:Mass Spectrometry; D008903:Minerals / Q000378:metabolism; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense; D015676:Osteonectin / Q000235:genetics* / Q000378:metabolism; D010037:Otolithic Membrane / Q000196:embryology / Q000254:growth & development* / Q000502:physiology*; D040901:Proteomics; D015027:Zebrafish / Q000196:embryology / Q000254:growth & development*; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
D008903:Minerals; D016376:Oligonucleotides, Antisense; D015676:Osteonectin; D029961:Zebrafish Proteins
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