The specific aims of this proposal are 1) to analyze in detail the chemistry and biosynthesis of bone proteoglycans, 2) to use antibodies in immunolocalization experiments, and 3) to evaluate the molecular mechanism by which 1,25-dihydroxyvitamin D3 appears to regulate osteopontin biosynthesis. The first two aims depend on classical biochemical and immunological methods while the last aim relies, to some extent, on the techniques of molecular biology. The applicant has some experience using these techniques, however, the funding of this Research Career Development Award would greatly enhance his research career development by allowing more extensive training in this area. The major hypothesis upon which this research is based is that non-collagenous components of mineralized connective tissues are intimately involved in the formation of the tissues. Furthermore, an understanding of the chemial composition, localization and regulatory mechanisms of biosynthesis are required before specific, functional roles can be elucidated. In this proposal we will address the aforementioned areas as they relate to proteoglycans and osteopontin. The approach taken is to isolate chemical amounts of these components, verify their purity and use them as antigens. The antibodies will be used in immunolocalization studies and as reagents in immunoassays. We will explore the kinetics of proteoglycan biosynthesis in rat bone, in ROS 17/2.8 cells and in cultures of rat calvarial osteoblasts. We will evaluate the role and mechanism that 1,25- dihydroxyvitamin D3 may have in the regulation of osteopontin biosynthesis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Modified Research Career Development Award (K04)
Project #
5K04DE000247-04
Application #
3072206
Study Section
Oral Biology and Medicine Subcommittee 1 (OBM)
Project Start
1988-07-01
Project End
1993-06-30
Budget Start
1991-07-01
Budget End
1992-06-30
Support Year
4
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Alabama Birmingham
Department
Type
Schools of Allied Health Profes
DUNS #
004514360
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Chang, P L; Yang, W C; Prince, C W (1995) Effects of okadaic acid on calcitriol- and phorbol ester-induced expression and phosphorylation of osteopontin in mouse JB6 epidermal cells. Ann N Y Acad Sci 760:24-34
Chang, P L; Ridall, A L; Prince, C W (1994) Calcitriol regulation of osteopontin expression in mouse epidermal cells. Endocrinology 135:863-9
Chambers, A F; Hota, C; Prince, C W (1993) Adhesion of metastatic, ras-transformed NIH 3T3 cells to osteopontin, fibronectin, and laminin. Cancer Res 53:701-6
Boskey, A L; Maresca, M; Ullrich, W et al. (1993) Osteopontin-hydroxyapatite interactions in vitro: inhibition of hydroxyapatite formation and growth in a gelatin-gel. Bone Miner 22:147-59
Singh, K; Deonarine, D; Shanmugam, V et al. (1993) Calcium-binding properties of osteopontin derived from non-osteogenic sources. J Biochem 114:702-7
Chang, P L; Prince, C W (1993) 1 alpha,25-Dihydroxyvitamin D3 enhances 12-O-tetradecanoylphorbol-13-acetate- induced tumorigenic transformation and osteopontin expression in mouse JB6 epidermal cells. Cancer Res 53:2217-20
Chang, P L; Prince, C W (1991) 1 alpha,25-dihydroxyvitamin D3 stimulates synthesis and secretion of nonphosphorylated osteopontin (secreted phosphoprotein 1) in mouse JB6 epidermal cells. Cancer Res 51:2144-50
Prince, C W; Dickie, D; Krumdieck, C L (1991) Osteopontin, a substrate for transglutaminase and factor XIII activity. Biochem Biophys Res Commun 177:1205-10
Prince, C W (1989) Secondary structure predictions for rat osteopontin. Connect Tissue Res 21:15-20