The objective of this application is to clone and characterize a putative new cell attachment protein that has been isolated from both cementum and human cementum tumor.This cementum attachment protein (CAP) promotes the attachment of fibroblasts but not epithelial cells, and its activity is inhibited by peptides. CAP binds to fibronectin and hydroxyapatite, but not collagen and it may use both the alpha 1- and alpha 5-integrin subunits as its cell surface receptor. Polyclonal and monoclonal antibodies to CAP inhibit cell attachment and demonstrate that CAP is found exclusively in cementum and in a human cementum tumor adapted to culture by the applicant and his colleagues.
In Specific Aim 1, a cultured human cementum tumor library already prepared will be screened with existing antibodies and the CAP cDNA will be cloned, characterized and expressed in vitro.
In Specific Aim 2, the biosynthesis of CAP will be studied. The applicant has found that CAP is synthesized as a 43 kDa precursor that is processed to 56, 49, 39,and 26 kDa forms based on preliminary studies that employ Western analysis of metabolically labeled cementum tumor cells. Potential glycosylation, phosphorylation, and sulfation sites of CAP will be studied by standard methods. Biosynthetic labeling and immunoprecipitation experiments, and Northern analysis when the CAP cDNA become available, will be used to explore the regulation of CAP expression in cementum tumor cells by growth factors known to regulate other extracellular matrix molecules (TGF-beta, PDGF, vitamin D, PTH and interferon-gamma). The binding of CAP to cell surface receptors in gingival and periodontal ligament fibroblasts and bone cells will be assayed by the use of biotin-conjugated CAP and radioiodine-labeled streptavidin.
In Specific Aims 3 and 4, immunohistochemistry will be used to determine the distribution of CAP in oral and other tissues under both normal and pathological conditions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE010491-02
Application #
2131379
Study Section
Oral Biology and Medicine Subcommittee 1 (OBM)
Project Start
1994-02-01
Project End
1998-01-31
Budget Start
1995-02-01
Budget End
1996-01-31
Support Year
2
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Washington
Department
Pathology
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Grzesik, Wojciech J; Narayanan, A S (2002) Cementum and periodontal wound healing and regeneration. Crit Rev Oral Biol Med 13:474-84
Saito, M; Iwase, M; Maslan, S et al. (2001) Expression of cementum-derived attachment protein in bovine tooth germ during cementogenesis. Bone 29:242-8
Yokokoji, T; Narayanan, A S (2001) Role of D1 and E cyclins in cell cycle progression of human fibroblasts adhering to cementum attachment protein. J Bone Miner Res 16:1062-7
BarKana, I; Narayanan, A S; Grosskop, A et al. (2000) Cementum attachment protein enriches putative cementoblastic populations on root surfaces in vitro. J Dent Res 79:1482-8
Komaki, M; Kang, M; Narayanan, A S (2000) Role of MAP kinases p42erk-2/p44erk-1 in cementum-derived attachment-protein-mediated cell attachment. J Dent Res 79:1789-93
Ivanovski, S; Komaki, M; Bartold, P M et al. (1999) Periodontal-derived cells attach to cementum attachment protein via alpha 5 beta 1 integrin. J Periodontal Res 34:154-9
Saito, M; Narayana, A S (1999) Signaling reactions induced in human fibroblasts during adhesion to cementum-derived attachment protein. J Bone Miner Res 14:65-72
Ikezawa, K; Ohtsubo, M; Norwood, T H et al. (1998) Role of cyclin E and cyclin E-dependent kinase in mitogenic stimulation by cementum-derived growth factor in human fibroblasts. FASEB J 12:1233-9
Liu, H W; Yacobi, R; Savion, N et al. (1997) A collagenous cementum-derived attachment protein is a marker for progenitors of the mineralized tissue-forming cell lineage of the periodontal ligament. J Bone Miner Res 12:1691-9
Narayanan, A S; Bartold, P M (1996) Biochemistry of periodontal connective tissues and their regeneration: a current perspective. Connect Tissue Res 34:191-201

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