The long-term objective of this revised proposal is to determine the role of the extracellular matrix (ECM) in the mineralization of Type I collagen-rich connective tissues such as bone and dentin. Numerous components of the extracellular matrix have been suggested to function as promoters or inhibitors of hydroxyapatite formation. It is proposed that the role of these non-collagenous components of the ECM can be better understood by using a simpler system such as the mineralizing turkey tendon. This system has a simpler tissue architecture and a linear mineralization sequence, thus facilitating the comparison of mineralized and non-mineralized matrices. The amended proposal will focus on the study of two proteins: 1) decorin, a collagen-binding proteoglycan which has been proposed to act as inhibitor of soft tissue mineralization, and 2)a 67 kDa protein which appears to be an avian analog of mammalian bone silo protein, which has the ability to nucleate hydroxyapatite crystals. The following specific studies will be done: 1) characterization of the 67 kDa avian protein and investigation of its effect on hydroxyapatite formation in vitro; 2) characterization of the proteoglycans in mineralized and non-mineralized tendon, and analysis of decorin degradation products at the mineralization front; and 3) investigation of matrix synthesis by tendon organ culture, in particular synthesis of the 67 kDa protein. The studies proposed will provide insights to understand the mechanisms involved in bone and dentin mineralization.
