POSITIVE REGULATION OF INSULIN GENE TRANSCRIPTION

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The ultimate goal of our research program is to understand, in molecular terms, all of the cis- and trans-acting elements of the rat insulin II gene which are involved in its tissue-specific expression within the pancreatic beta-cell. In particular we are attempting to understand the positive- acting components of this system. We have recently characterized the rat insulin III gene 5'-flanking sequences which encompass its enhancer. We have also shown that there are multiple, discrete elements which comprise this enhancer is an -10bp sequence located t positions -100 to -91 within the rat insulin gene II transcription unit. We have detected a DNA-binding protein, which is uniquely present (active) in insulin producing cells, that specifically interacts with this important regulatory sequence. Moreover, recently we have identified and isolated two lamda-gt11-cDNA clones which express a fusion protein which specifically binds to the oligomerized -100 to -91 element. We propose to further characterize these cis- and trans-acting components by purifying the factor which binds to the -100 to -91 element, cloning the gene which encodes it and studying both its mechanism of action regarding the promotion of insulin gene transcription and the regulation of expression of this insulin gene transcriptional regulator itself. We will also search for additional trans-acting factors which positively affect insulin gene transcription in beta-cells. Once this factors(s) has been identified it will be subjected to the analyses outlined above for the factor operating through the -100 to -91 element. Understanding the mechanism of regulation of insulin gene transcription is fundamental to gaining a thorough, basic understanding of glucose homeostasis and pathological states thereof.