This is a study of structural and metabolic defects in procollagen in genetic diseases of connective tissues. Each simple inherited defect in a procollagen gene is a unique opportunity to study one or more steps in the metabolism of procollagen. Abnormalities in the structure and metabolism of procollagen will be identified and characterized in ascorbate stimulated post-confluent cultures of skin and tendon fibroblasts, and in collagens and procollagens extracted from affected tissues. We will study the effects of each mutation on collagen synthesis, post-translational modification, secretion, extracellular processing of procollagen to collagen, and covalent cross-linking of collagen within fibrils. The initial studies will utilize pulse chase experiments with cultured fibroblasts from humans and domestic animals with various forms of the Ehlers-Danlos syndrome (EDS), Marfans syndrome, and osteogenesis imperfect (OI) (J. Biol> Chem. 261:10006, 1986). The radiolabeled and extracted proteins will be separated and characterized with liquid chromatography, SDS-PAGE, peptide mapping, amino acid analysis, CD analysis and by partial enzymatic degradation at different temperatures. These studies will serve to identify primary and secondary effects of each mutation of different types of procollagen, and identify those mutations that result in regulatory defects that affect the transcription of procollagen genes. The date on both the primary structural and metabolic defects in procollagen chains will be used by laboratories of molecular genetics to study the mutation in procollagen genes. At the same time these data will be correlated with analyses of the effects of the structural and metabolic abnormalities on the thermal stability, secretion, and processing of procollagens, and on the deposition and cross-linking of collagens within fibrils.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR020793-11
Application #
3155144
Study Section
Pathobiochemistry Study Section (PBC)
Project Start
1979-04-01
Project End
1993-06-30
Budget Start
1991-07-20
Budget End
1993-06-30
Support Year
11
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Cornell University
Department
Type
Schools of Veterinary Medicine
DUNS #
City
Ithaca
State
NY
Country
United States
Zip Code
14850
Campbell, B G; Wootton, J A; Macleod, J N et al. (2001) Canine COL1A2 mutation resulting in C-terminal truncation of pro-alpha2(I) and severe osteogenesis imperfecta. J Bone Miner Res 16:1147-53
Campbell, B G; Wootton, J A; MacLeod, J N et al. (2000) Sequence of normal canine COL1A1 cDNA and identification of a heterozygous alpha1(I) collagen Gly208Ala mutation in a severe case of canine osteogenesis imperfecta. Arch Biochem Biophys 384:37-46
Campbell, B G; Wootton, J A; MacLeod, J N et al. (1998) Sequence of canine COL1A2 cDNA: nucleotide substitutions affecting the cyanogen bromide peptide map of the alpha 2(I) chain. Arch Biochem Biophys 357:67-75
Campbell, B G; Wootton, J A; Krook, L et al. (1997) Clinical signs and diagnosis of osteogenesis imperfecta in three dogs. J Am Vet Med Assoc 211:183-7
Sams, A E; Minor, R R; Wootton, J A et al. (1995) Local and remote matrix responses to chondrocyte-laden collagen scaffold implantation in extensive articular cartilage defects. Osteoarthritis Cartilage 3:61-70
Hendrickson, D A; Nixon, A J; Grande, D A et al. (1994) Chondrocyte-fibrin matrix transplants for resurfacing extensive articular cartilage defects. J Orthop Res 12:485-97
Todhunter, R J; Wootton, J A; Altman, N et al. (1994) Cross-validation of cyanogen bromide-peptide ratios to measure the proportion of type II collagen in pepsin digests of equine articular cartilage, meniscus, and cartilage repair tissue. Anal Biochem 216:195-204
Todhunter, R J; Wootton, J A; Lust, G et al. (1994) Structure of equine type I and type II collagens. Am J Vet Res 55:425-31
Hendrickson, D A; Nixon, A J; Erb, H N et al. (1994) Phenotype and biological activity of neonatal equine chondrocytes cultured in a three-dimensional fibrin matrix. Am J Vet Res 55:410-4
Todhunter, R J; Minor, R R; Wootton, J A et al. (1993) Effects of exercise and polysulfated glycosaminoglycan on repair of articular cartilage defects in the equine carpus. J Orthop Res 11:782-95

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