The glycosylation profile of osteoadherin alters during endochondral bone formation

Endochondral bone formation involves the dynamic interplay between the cells and their extracellular environment to facilitate the deposition of a calcified matrix. Numerous molecules are involved within this process, including collagens and non-collagenous proteins, and their post-translational modifications have been shown to effect their biomolecular interactions. Osteoadherin (OSAD), a keratin sulfate (KS)-substituted small leucine-rich proteoglycan has been isolated from mineralized tissues and is considered to be a mineralized tissue-specific protein. However, to date, information is limited concerning the dynamic expression and role of this proteoglycan during bone formation and the biomineralization process. The current study aimed to examine the dynamic expression of this protein throughout mouse metatarsal long bone development, from the cartilage anlagen (E15) to the fully formed bone (Adult). Using quantitative gene expression analysis we observed that OSAD was produced with the onset of mineralization and the formation of the ossification center. This finding was reflected in the localization studies, using both light and electron microscopy, and showed that initial OSAD localization was restricted to the endosteal surfaces of the diaphysis and forming metaphysis. Furthermore, we analyzed protein extracts, both mineral and non-mineral associated fractions, and showed that OSAD was substituted with varying patterns of glycosylation during bone development. Sequential enzymatic digestions of the non-mineral bound protein extracts demonstrated that OSAD lacked any KS chains throughout all development stages. Whereas, in the mineral bound fractions, with long bone maturation the substitution with KS became more apparent with development. Therefore, it can be concluded that different pools of OSAD are produced during endochondral bone formation and these may have specific roles in directing the mineralization process. (C) 2013 Elsevier Inc. All rights reserved.