title: Osteocrin, a novel bone-specific secreted protein that modulates the osteoblast phenotype.
abstract: Although a number of secreted factors have been demonstrated to be bone regulators, none of these are unique to bone. Using a viral-based signal-trap strategy we have identified a novel gene we have termed "osteocrin." A 1280-bp mRNA encodes osteocrin producing a mature protein of 103 amino acids with a molecular mass of 11.4 kDa. Osteocrin shows no homology with any known gene except for two conserved sequence motifs reminiscent of dibasic cleavage sites found in peptide hormone precursors. Immunofluorescence and Western blot analysis confirmed the secretory nature of osteocrin. Two protein species were identified in the medium of cells overexpressing osteocrin, a full-length 11.4 kDa species and a processed approximately 5 kDa species. Mutation of the 76KKKR79 dibasic cleavage site abolished the appearance of this smaller osteocrin fragment. By in situ hybridization in mouse embryos, osteocrin was expressed specifically in Cbfa-1-positive, osteocalcin-negative osteoblasts. Immunohistochemistry on adult mouse bone showed osteocrin localization in osteoblasts and young osteocytes. By Northern blot analysis, osteocrin expression was only detected in bone, expression peaking just after birth and decreasing markedly with age. In primary osteoblastic cell cultures osteocrin expression coincided with matrix formation then decreased in very mature cultures. Treatment of cultures with 1,25-dihydroxyvitamin D3 resulted in a rapid dose-dependent down-regulation of osteocrin expression, suggesting direct regulation. Chronic treatment of primary cultures with osteocrin-conditioned media inhibited mineralization and reduced osteocalcin and alkaline phosphatase expression. These results suggest that osteocrin represents a novel, unique vitamin D-regulated bone-specific protein that appears to act as a soluble osteoblast regulator.
authors.affiliation: Phenogene Therapeutics Inc., 416 de Maisonneuve West, Suite 1020, Montreal, Quebec H3A 1L2, Canada. gthomas@phenogene.com
authors.forename: Gethin
authors.initials: G
authors.lastname: Thomas
authors.forename: Pierre
authors.initials: P
authors.lastname: Moffatt
authors.forename: Patrick
authors.initials: P
authors.lastname: Salois
authors.forename: Marie-Hélène
authors.initials: MH
authors.lastname: Gaumond
authors.forename: Rock
authors.initials: R
authors.lastname: Gingras
authors.forename: Eric
authors.initials: E
authors.lastname: Godin
authors.forename: Dengshun
authors.initials: D
authors.lastname: Miao
authors.forename: David
authors.initials: D
authors.lastname: Goltzman
authors.forename: Christian
authors.initials: C
authors.lastname: Lanctôt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015152:Blotting, Northern; D015153:Blotting, Western; D002117:Calcitriol / Q000494:pharmacology; D003001:Cloning, Molecular; D017077:Culture Media, Conditioned / Q000494:pharmacology; D017931:DNA Primers / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D051379:Mice; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009124:Muscle Proteins; D009154:Mutation; D010006:Osteoblasts / Q000378:metabolism*; D010641:Phenotype; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014157:Transcription Factors; D014162:Transfection
chemical_list: D017077:Culture Media, Conditioned; D017931:DNA Primers; D018076:DNA, Complementary; D009124:Muscle Proteins; C480236:OSTN protein, human; D011506:Proteins; D014157:Transcription Factors; D002117:Calcitriol