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ABI Research Summaries - Lawrence E. Cornett, Ph.D. Dr. Cornett's research interests are focused on regulation of adrenergic receptor gene expression and the homeostatic roles of the neurohypophysial hormone vasotocin in the domestic chicken. Adrenergic receptors mediate the effects of epinephrine and norepinephrine. My laboratory is investigating the molecular events involved in the regulation of 2-adrenergic receptor gene expression by glucocorticoids, an action that is clinically useful in the treatment of asthma. We have identified a glucocorticoid response element (GRE) and other regulatory elements upstream of the promoter that play key roles in regulating 2-adrenergic receptor gene transcription in airway epithelial cells. Regarding the actions of vasotocin, immunohistochemical techniques, radioimmunoassays and in situ hybridization histochemistry are being used to investigate regulation of vasotocin gene expression in the chicken hypothalamus under conditions of osmotic stress and during oviposition. We have cloned and pharmacologically characterized receptors that bind vasotocin and transduce its physiological effects in the shell gland and anterior pituitary gland. ß2-Adrenergic Receptor Gene Therapy in Asthma. Asthma is a disease that affects approximately 16 million Americans and causes over 5,000 deaths annually. Major pathophysiological components of asthma are allergic airway inflammation and subsequent bronchospasm. Current treatment options include medications that control the inflammatory component of asthma (e.g., corticosteroids, cromolyn sodium, methylxanthines, leukotriene modifiers) and quick relief medications that counteract bronchospasms (e.g., -adrenergic agonists). Relatively few other treatment options are available, particularly for patients with severe forms of the disease. We are developing an innovative approach to develop a gene therapy treatment for asthma. A recombinant 2-adrenergic receptor gene is targeted to airway epithelial cells via an adeno-associated virus vector. Overexpression of the recombinant 2-adrenergic receptor decreases overall airway resistance in a mouse model of airway hypersensitivity by increasing airway sensitivity to the relaxing effects of 2-adrenergic agonists. Glucorticoid Regulation of ß2-Adrenergic Receptor Gene Expression. Regulation of 2-adrenergic receptor levels by glucocorticoids is a physiologically important mechanism for altering 2-adrenergic receptor responsiveness. Glucocorticoids increase 2-adrenergic receptor density by increasing the rate of 2-adrenergic receptor gene transcription. We have demonstrated that one of six potential glucocorticoid response elements (GREs) in the 5'-flanking region of the rat 2-adrenergic receptor gene is necessary for glucocorticoid-dependent stimulation of receptor gene expression. Using a nested set of deletion fragments of the rat 2-adrenergic receptor gene 5'-flanking region fused to a luciferase reporter gene, glucocorticoid-dependent induction of reporter gene expression in HepG2 cells was localized to a region between positions -643 and -152, relative to the transcription initiation site.. In electrophoretic mobility shift assays, a double-stranded oligonucleotide incorporating a near consensus GRE from this region formed complexes with the human recombinant glucocorticoid receptor, as well as with nuclear protein from dexamethasone-treated HepG2 cells. Regulation of Oviposition in Chickens by Vasotocin.
In chickens, oviposition is correlated with increased plasma levels of the neurohypophysial hormone vasotocin, and vasotocin stimulates contraction of uterine strips in vitro. Vasotocin is the avian homolog of the mammalian hormone oxytocin that stimulates uterine contractility during parturition. We have cloned a vasotocin receptor from the domestic chicken. The open reading frame encodes a 370 amino acid polypeptide that displays seven segments of hydrophobic amino acids, typical of guanine nucleotide protein coupled receptors. Expressed VT1 receptors in COS7 cells bind neurohypophysial hormones with a rank order of potency expected of a vasotocin receptor. In the chicken, expression of VT1 receptors is limited to the shell gland (uterus) and the brain. This receptor likely mediates contractions of the shell gland during an egg lay and may activate reproductive behaviors known to be stimulated by vasotocin in birds. Regulation of Stress Responses in Chickens by the Neurohypophysial Hormone Vasotocin. The neurohypophysial hormone vasotocin stimulates adrenocorticotrophin hormone (ACTH) secretion from the avian anterior pituitary gland resulting in increased adrenal secretion corticosterone in response to stress. We have cloned and functional characterized a gene encoding a vasotocin receptor that appears to mediate the stimulatory effect of vasotocin on ACTH secretion in birds. Expressed VT2 receptors in COS7 cells mediate vasotocin-induced phosphatidylinositol turnover and calcium mobilization. In the domestic chicken, expression of the VT2 receptor gene is limited to the pituitary gland. This receptor is likely the avian homolog of the mammalian V1B-vasopressin receptor and therefore may play an important role in the avian stress response. Selected Publications: Cornett, L.E., F.C. Hiller, S.E. Jacobi, W. Cao, and D.W. McGraw. Identification of a glucocorticoid response element in the rat 2-adrenergic receptor gene. Molecular Pharmacology 54:1016-1023, 1998. Cao, W., D.W. McGraw, T.T. Lee, A. Dicker-Brown, F.C. Hiller, L.E. Cornett, and S.M. Jones. Expression of functional 2-adrenergic receptors in a rat airway epithelial cell line (SPOC1) and cell density-dependent induction by glucocorticoids. Experimental Lung Research 26:421-435, 2000. Tan, F.-l., S.M. Lolait, M.J. Brownstein, N. Saito, V. MacLeod, D.A. Baeyens, P.R. Mayeux, S.M. Jones, and L.E. Cornett. Molecular cloning and functional characterization of a vasotocin receptor subtype that is expressed in the shell gland and brain of the domestic chicken. Biology of Reproduction 62:8-15, 2000. Cornett, L.E., J.D. Kirby, J.A. Vizcarra, J.C. Ellison, J. Thrash, P.R. Mayeux, M.D. Crew, S.M. Jones, N. Ali, and D.A. Baeyens. Molecular cloning and functional characterization of a vasotocin receptor subtype expressed in the pituitary gland of the domestic chicken (Gallus domesticus): Avian homolog of the mammalian V1B-vasopressin receptor. Regulatory Peptides 110:231-239, 2003. Contact Information Lawrence E. Cornett, Ph.D. University of Arkansas for Medical Sciences Phone: (501) 686-5441 |