Rescue of Chga knockout phenotype by CHGA-BAC transgene

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DESCRIPTION (provided by applicant): Chromogranin A (CGA) is co-stored with catecholamines in chromaffin granules and co-released in response to sympathoadrenal stimulation. From CGA pro-protein several biologically active peptides including catecholamine release-inhibitory peptide catestatin are derived. Nicotine, the primary component of tobacco, stimulates secretion of catecholamines, augments biosynthesis of CGA, and, as a nicotinic cholinergic agonist, promotes desensitization of catecholamine secretion. In developed countries, tobacco use is the largest single cause of premature death and heart disease the most common cause of death. Studies have demonstrated that smoking, diabetes and hypertension are major risk factors for coronary heart disease. Catestatin level is diminished in hypertensive patients. Chga knockout (Ko) mice have been generated in this laboratory. Their striking phenotypic characteristics include elevated systolic blood pressure (SBP), decreased catecholamines levels, loss of chromaffin granules and partial prenatal lethality. The proposed study specifically aims to: 1. Rescue the Chga-Ko phenotype by introducing the human CHGA gene using bacterial artificial chromosome (BAC) transgenic approach. 2. Sequencing of the human CHGA gene, revealed three non-synonymous SNPs (single nucleotide polymorphism) in the catestatin sequence. The proposed study will analyze the differences in regulation of SBP, catecholamine levels and nicotine cholinergic responses between the wild type and mutant variants of CGA. Variant catestatin peptides will be synthesized and administered into Chga -/- mice. The 3 mutant CHGA-BAC mouse models (Gly364Ser, Pro370Leu, and Arg374GIn) will also be generated to explore the significance of such polymorphisms. 3. Finally, the CGA variants will be assessed for processing by proteases in vitro and in vivo.