Factor IX is a vitamin K dependent plasma protein synthesized in the liver and required for normal coagulation. It occupies a central position in the clotting cascade, where it undergoes critical interactions with Factors XI, VII, VIII and X. A deficiency of Factor IX results in the clinical syndrome hemophilia B. The major thrust of this project is to make use of the cloned Factor IX gene to study the structure, function and expression of Factor IX. Sources of material for study will include the large numbers of patients with hemophilia B followed at this institution, and the colony of dogs with hemophilia B here. An initial goal will be to isolate and characterize at the nucleotide level selected human Factor IX mutations. The patients selected for study include: 1) patients with a variant designated hemophilia BM. These variants are known to have a defect in interaction with Factors VII and/or X. 2) Patients with a subtype of CRM-reduced hemophilia B (those with equal reductions in antigen and activity). Through examination of these variants, regions of the Factor IX DNA sequence which control or are necessary for expression will be identified. Methods to be used include construction and screening of genomic libraries, and M13/dideoxy sequence analysis. The goal of the second group of experiments is to determine the level at which changes in Factor IX protein levels are regulated. Utilizing an animal model, it will be possible to examine rates of transcription, transcript stability, transcript size, translational efficiency, post translational modifications and protein secretion; and to determine at which of these levels changes in Factor IX expression are regulated. Finally, taking advantage of Factor IX's location on the X chromosome, experiments will be carried out to determine whether differences in the chromatin configuration can be detected between the active and inactive X chromosomes in the case of Factor IX. Specifically, use will be made of liver tissue from dogs who are carriers of hemophilia B (and in whom, therefore, the two X alleles can be differentiated with the use of appropriate probes), to determine whether differences in DNAse hypersensitive sites and cytosine methylation exist between the expressed and the non-expressed allele.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Clinical Investigator Award (CIA) (K08)
Project #
1K08HL001922-01
Application #
3082332
Study Section
Research Manpower Review Committee (MR)
Project Start
1987-04-01
Project End
1992-03-31
Budget Start
1987-04-01
Budget End
1988-03-31
Support Year
1
Fiscal Year
1987
Total Cost
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Type
Schools of Medicine
DUNS #
078861598
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Chaing, S; Clarke, B; Sridhara, S et al. (1994) Severe factor VII deficiency caused by mutations abolishing the cleavage site for activation and altering binding to tissue factor. Blood 83:3524-35
Rose, V L; Dermott, S C; Murray, B F et al. (1993) Decentralized testing for prothrombin time and activated partial thromboplastin time using a dry chemistry portable analyzer. Arch Pathol Lab Med 117:611-7
Larson, P J; High, K A (1992) Biology of inherited coagulopathies: factor IX. Hematol Oncol Clin North Am 6:999-1009
Holycross, B J; Saye, J; Harrison, J K et al. (1992) Polymerase chain reaction analysis of renin in rat aortic smooth muscle. Hypertension 19:697-701
Huang, M N; Hung, H L; Stanfield-Oakley, S A et al. (1992) Characterization of the human blood coagulation factor X promoter. J Biol Chem 267:15440-6
Watzke, H H; Wallmark, A; Hamaguchi, N et al. (1991) Factor XSanto Domingo. Evidence that the severe clinical phenotype arises from a mutation blocking secretion. J Clin Invest 88:1685-9
Wallmark, A; Rose, V L; Ho, C et al. (1991) A NlaIV polymorphism within the human factor X gene. Nucleic Acids Res 19:4022
Lozier, J N; High, K A (1990) Molecular basis of hemophilia. Hematol Pathol 4:1-26
Lozier, J N; Monroe, D M; Stanfield-Oakley, S et al. (1990) Factor IX New London: substitution of proline for glutamine at position 50 causes severe hemophilia B. Blood 75:1097-104
Watzke, H H; Lechner, K; Roberts, H R et al. (1990) Molecular defect (Gla+14----Lys) and its functional consequences in a hereditary factor X deficiency (factor X ""Vorarlberg""). J Biol Chem 265:11982-9

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