Proteolytic processing of osteopontin by PHEX and accumulation of osteopontin fragments in Hyp mouse bone, the murine model of X-linked hypophosphatemia

Autor(es)

Barros, Nilana M. T. ^{[1, 2]} ; Hoac, Betty ^[3] ; Neves, Raquel L. ^{[1, 2]} ; Addison, William N. ^[4] ; Assis, Diego M. ^[1] ; Murshed, Monzur ^{[3, 5]} ; Carmona, Adriana K. ^[1] ; McKee, Marc D. ^{[3, 6]}

Afiliação do(s) autor(es)

^[1] Univ Fed Sao Paulo, Dept Biofis, BR-04039032 Sao Paulo - Brazil

^[2] Univ Fed Sao Paulo, Dept Ciencias Exatas & Terra, Diadema - Brazil

^[3] McGill Univ, Fac Dent, Montreal, PQ - Canada

^[4] Harvard Univ, Sch Dent Med, Dept Oral Med Infect & Immun, Boston, MA 02115 - USA

^[5] McGill Univ, Dept Med, Fac Med, Montreal, PQ - Canada

^[6] McGill Univ, Fac Med, Dept Anat & Cell Biol, Montreal, PQ - Canada

• Tipo de documento: Artigo Científico
• Fonte: Journal of Bone and Mineral Research; v. 28, n. 3, p. 688-699, MAR 2013.
• Citações Web of Science: 51

Resumo

X-linked hypophosphatemia (XLH/HYP)with renal phosphate wasting, hypophosphatemia, osteomalacia, and tooth abscessesis caused by mutations in the zinc-metallopeptidase PHEX gene (phosphate-regulating gene with homologies to endopeptidase on the X chromosome). PHEX is highly expressed by mineralized tissue cells. Inactivating mutations in PHEX lead to distal renal effects (implying accumulation of a secreted, circulating phosphaturic factor) and accumulation in bone and teeth of mineralization-inhibiting, acidic serine- and aspartate-rich motif (ASARM)-containing peptides, which are proteolytically derived from the mineral-binding matrix proteins of the SIBLING family (small, integrin-binding ligand N-linked glycoproteins). Although the latter observation suggests a local, direct matrix effect for PHEX, its physiologically relevant substrate protein(s) have not been identified. Here, we investigated two SIBLING proteins containing the ASARM motifosteopontin (OPN) and bone sialoprotein (BSP)as potential substrates for PHEX. Using cleavage assays, gel electrophoresis, and mass spectrometry, we report that OPN is a full-length protein substrate for PHEX. Degradation of OPN was essentially complete, including hydrolysis of the ASARM motif, resulting in only very small residual fragments. Western blotting of Hyp (the murine homolog of human XLH) mouse bone extracts having no PHEX activity clearly showed accumulation of an approximate to 35kDa OPN fragment that was not present in wild-type mouse bone. Immunohistochemistry and immunogold labeling (electron microscopy) for OPN in Hyp bone likewise showed an accumulation of OPN and/or its fragments compared with normal wild-type bone. Incubation of Hyp mouse bone extracts with PHEX resulted in the complete degradation of these fragments. In conclusion, these results identify full-length OPN and its fragments as novel, physiologically relevant substrates for PHEX, suggesting that accumulation of mineralization-inhibiting OPN fragments may contribute to the mineralization defect seen in the osteomalacic bone characteristic of XLH/HYP. (c) 2013 American Society for Bone and Mineral Research. (AU)

• Processo FAPESP: 11/50495-0 - Estudo do envolvimento da metalo-peptidase PHEX em processos fisiológicos e fisiopatológicos
• Beneficiário: Nilana Meza Tenório de Barros Jaggi
• Modalidade de apoio: Auxílio à Pesquisa - Regular