This five-year project will primarily focus on the molecular and cell biology studies of human class 1 heparin-binding growth factor (HBGF-1). HBGF's are mitogens for a variety of mesoderm-derived cells in vitro and can induce neovascularization in vivo. They are characterized by an unusual affinity for heparin and probably play a role in blood vessel homeostasis. Two overlapping cDNA clones encoding human HBGF were isolated from a human brain cDNA library. Southern blot analysis suggested that there is a single copy of the HBGF-1 gene and that it maps to human chromosome 5 at bands 5q31.3 to 33.2. The complete amino acid sequence of human HBGF has been deduced from the nucleic acid sequence of these clones and is identical to the recently published amino acid sequence of human HBGF-1. With the availability of these cDNA probes, it is now possible to screen the human genomic library to isolate the genes coding for HBGF-1. The positive clones will be characterized and sequenced to deduce the exon-intron structure. The identification of the 5' end of the gene will rely on S1 nuclease analysis and primer extension studies. The promoter region and other cis-acting sequences will be identified by constructing a series of recombinant molecules which will direct the expression of the easily assayed gene chloramphenicol acetyl transferase in mammalian cells. Better understanding of the gene organization including its promoter region will help us understand how this gene is regulated during cellular differentiation and/or malignant transformation. This last point is important in the light of a chromosomal abnormality in patients of refectory anemia or acute non-lymphocytic leukemia. The abnormality resided at the region where HBGF gene is located. Thus, cloning of HBGF gene will likely lead us to identify the breakpoint in the chromosomes of these patients at the molecular level. We will also study the spatial and temporal expression of the HBGF-1 gene in developing human placenta. Furthermore, we propose to study the transforming potential and the functional domains of HBGF in both fibroblasts and endothelial cells. These studies will further our understanding of the regulatory mechanism of cell growth in general. In specific, we may be able to find ways to prognose and eventually to treat patients with acute non-lymphocytic leukemia.
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