Alterations in the synthesis of extracellular matrix components including type I collagen occur in a number of fibrotic diseases such as scleroderma, ostearithritis as well as osteoporosis. Studies of the regulatory mechanisms which control the expression of the two type I collagen genes are critical to understand these alterations. In most physiological and pathological conditions, the expression of the alpha1(I) and alpha2(I) collagen genes are coordinately expressed. This coordinate regulation can be explained by the interaction of DNA-binding proteins with common cis-acting elements in the promoter region of both genes. One sequence specific DNA-binding protein that binds similar cis-acting elements located at approximately the same distance from the transcriptional start site in the promoter regions of the alpha1(I) and alpha2(I) mouse collagen genes, has been purified to homogeneity. This factor acts as a transcriptional inhibitor and is called Inhibitory Factor 2 (IF-2). IF-2, a metallo protein that requires zinc ions to bind to DNA, is present in a large variety of cells and presumably participates in the control of expression of several genes. We propose to purify IF-2 to homogeneity, to obtain amino-acid sequences for this protein and to clone cDNAs corresponding to the factor. We will use the purified bacterially expressed protein to generate polyclonal antibodies against IF-2. Using the purified protein and the cDNA clone we plan to perform, at a later time, a structure-function analysis of the molecule, to study the molecular mechanism of transcriptional repression by this transcription factor. We expect that the characterization of transcription factors like IF-2 and the molecular studies of its function win increase understanding of the mechanisms which control the expression of the type I collagen genes in physiological situations and in pathological situations and probably other genes and will bring new information about the regulatory mechanisms used by eukaryotic cells to synthesize specialized proteins.
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