title: Posttranslational processing of FGF23 in osteocytes during the osteoblast to osteocyte transition.
abstract: FGF23 is an O-glycosylated circulating peptide hormone with a critical role in phosphate homeostasis; it is inactivated by cellular proprotein convertases in a pre-release degradative pathway. We have here examined the metabolism of FGF23 in a model bone cell line, IDG-SW3, prior to and following differentiation, as well as in regulated secretory cells. Labeling experiments showed that the majority of (35)S-labeled FGF23 was cleaved to smaller fragments which were constitutively secreted by all cell types. Intact FGF23 was much more efficiently stored in differentiated than in undifferentiated IDG-SW3 cells. The prohormone convertase PC2 has recently been implicated in FGF23 degradation; however, FGF23 was not targeted to forskolin-stimulatable secretory vesicles in a regulated cell line, suggesting that it lacks a targeting signal to PC2-containing compartments. In vitro, PC1/3 and PC2, but not furin, efficiently cleaved glycosylated FGF23; surprisingly, PC5/6 accomplished a small amount of conversion. FGF23 has recently been shown to be phosphorylated by the kinase FAM20C, a process which was shown to reduce FGF23 glycosylation and promote its cleavage; our in vitro data, however, show that phosphorylation does not directly impact cleavage, as both PC5/6 and furin were able to efficiently cleave unglycosylated, phosphorylated FGF23. Using qPCR, we found that the expression of FGF23 and PC5/6, but not PC2 or furin, increased substantially following osteoblast to osteocyte differentiation. Western blotting confirmed the large increase in PC5/6 expression upon differentiation. FGF23 has been linked to a variety of bone disorders ranging from autosomal dominant hypophosphatemic rickets to chronic kidney disease. A better understanding of the biosynthetic pathway of this hormone may lead to new treatments for these diseases.
authors.affiliation: Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yamamoto
authors.affiliation: Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors.forename: Bruno
authors.initials: B
authors.lastname: Ramos-Molina
authors.affiliation: Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors.forename: Adam N
authors.initials: AN
authors.lastname: Lick
authors.affiliation: School of Dentistry, Department of Oral and Craniofacial Sciences, University of Missouri-Kansas City, Kansas City, MO 64108, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Prideaux
authors.affiliation: Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
authors.forename: Valeria
authors.initials: V
authors.lastname: Albornoz
authors.affiliation: School of Dentistry, Department of Oral and Craniofacial Sciences, University of Missouri-Kansas City, Kansas City, MO 64108, USA.
authors.forename: Lynda
authors.initials: L
authors.lastname: Bonewald
authors.affiliation: Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA. Electronic address: ilindberg@som.umaryland.edu.
authors.forename: Iris
authors.initials: I
authors.lastname: Lindberg
mesh_terms: D000818:Animals; D016466:CHO Cells; D002454:Cell Differentiation; D006224:Cricetinae; D003412:Cricetulus; D018076:DNA, Complementary / Q000235:genetics; D000089703:Fibroblast Growth Factor-23; D005346:Fibroblast Growth Factors / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D017350:N-Acetylgalactosaminyltransferases / Q000378:metabolism; D051859:Neuroendocrine Secretory Protein 7B2 / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism*; D010011:Osteocytes / Q000166:cytology* / Q000378:metabolism*; D010766:Phosphorylation; D043484:Proprotein Convertases / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D000097763:Polypeptide N-acetylgalactosaminyltransferase
chemical_list: D018076:DNA, Complementary; D000089703:Fibroblast Growth Factor-23; D005346:Fibroblast Growth Factors; D017350:N-Acetylgalactosaminyltransferases; D051859:Neuroendocrine Secretory Protein 7B2; D043484:Proprotein Convertases; D011994:Recombinant Proteins; D000097763:Polypeptide N-acetylgalactosaminyltransferase; C000717427:FGF23 protein, human; C000717428:Fgf23 protein, mouse
keywords: 7B2; FGF23; Furin; Osteocyte; PC5/6; Posttranslational processing; Proprotein convertase