MOLECULAR DISTRIBUTION OF HUMAN ADRENERGIC RECEPTORS

人肾上腺素能受体的分子分布

• 批准号: 6638338
• 负责人: Debra Anne Schwinn
• 金额: $ 39.3万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-09-01 至 2005-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6638338
• 关键词: adrenergic receptor; affinity labeling; cell line; chimeric proteins; deoxyribonuclease I; gel mobility shift assay; gene deletion mutation; genetic promoter element; genetic regulatory element; hormone regulation /control mechanism; human tissue; in situ hybridization; laboratory rat; liver cells; nucleic acid sequence; phosphorylation; receptor expression; receptor sensitivity; second messengers; species difference; transcription factor; transfection

The overall long term objective of this research is to determine mechanisms underlying regulation of human adrenergic receptors (ARs). Within this context, this competitive renewal seeks to investigate two specific aspects of AR regulation using the alpha1aAR as a model--1) mechanisms underlying species specific tissue distribution of AR subtypes, and 2) mechanisms underlying agonist-induced regulation of the human alpha1aAR. During the last grant period we demonstrated AR subtype specific and species-dependent tissue expression. Although 5'- regulatory sequences have been shown to bind tissue specific transcription factors, this information is limited to detailed analysis of only a few tissues. Since comparative information regarding human and rat tissue distribution of alpha1AR subtypes was obtained during the last grant period, and other studies in our laboratory (not directly related to this grant) resulted in cloning and characterization of 6.2kb 5'UTR of the human alpha1aAR gene, we are in a unique position to test the hypothesis that species specific tissue expression (rat versus human) of ARs results from differences in 5'-regulatory gene sequences. This will be accomplished by cloning the rat alpha1aAR 5'UTR and comparing rat versus human reporter construct expression in tissues of interest (rat neonatal myocytes, hepatoma cell lines, SK-N-MC cells); DNase I assays using nuclei isolated from fresh human and rat tissues of interest (heart, mammary artery, liver) will also be utilized for these studies. During the previous grant period we also examined the influence of disease and age on human cardiovascular AR subtype expression. Hypertension, congestive heart failure, and aging all increase circulating catecholamine levels, hence a natural progression from our previous studies is to examine the response of ARs to agonist exposure. The alpha1aAR is unique in that agonist exposure in a rat neonatal myocyte model results in upregulation and continued signaling, while alpha1b and alpha1dARs undergo concurrent downregulation and desensitization; this pathway has been shown to be important in alpha1AR-mediated myocardial hypertrophy and raises the question of whether alpha1aARs are capable of undergoing desensitization. In preliminary studies we find that rapid (less than 5 min) acute partial desensitization of human alpha1aARs occurs (signaling remains 50 percent of naive receptor after 30 min of agonist exposure). Therefore we plan to test the hypothesis that alpha1aAR acute partial desensitization occurs via mechanisms distinct from other ARs, enabling persistent signaling in the presence of agonist. These studies will utilize a hemagglutinin (HA)-tagged human alpha1aAR and COOH-terminal deletion mutant to characterize agonist-dependent (norepinephrine [NE]) and agonist-independent (phorbol ester [PMA]) pathways, as well as study the role of receptor phosphorylation (using immunoprecipitation of HA tagged receptors) and sequestration (whole cell binding assays). Mutation of putative phosphorylation sites and chimeric alpha1a/alpha1bARs will then be used to further define mechanisms underlying this process. Extensive training in molecular pharmacology of ARs, experience in protein biochemistry of desensitization, experience in cloning 5'UTRs and characterizing the human alpha1aAR gene, as well as availability of several human alpha1aAR reporter constructs already in the laboratory, places us in a unique position to elucidate mechanisms underlying regulation of ARs using the alpha1aAR as a model. Results from these studies should facilitate therapeutic interventions (e.g. enhanced human tissue targeting for gene therapy) as well as understanding of mechanisms underlying cardiovascular diseases.

这项研究的总体长期目标是确定

项目成果

Genetics and patient outcome after cardiac surgery: unravelling translational findings.

心脏手术后的遗传学和患者结果：揭示转化结果。

• DOI: 10.1093/bja/aer347
• 发表时间: 2011
• 期刊: British journal of anaesthesia
• 影响因子: 9.8
• 作者: Schwinn,DA

Alpha1a-Adrenoceptor Genetic Variant Triggers Vascular Smooth Muscle Cell Hyperproliferation and Agonist Induced Hypertrophy via EGFR Transactivation Pathway.

• DOI: 10.1371/journal.pone.0142787
• 发表时间: 2015
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Gradinaru I;Babaeva E;Schwinn DA;Oganesian A
• 通讯作者: Oganesian A

A critical period for the role of thyroid hormone in development of renal alpha-adrenergic receptors.

甲状腺激素在肾α-肾上腺素能受体发育中发挥作用的关键时期。

• DOI: 10.1203/00006450-199707000-00015
• 发表时间: 1997
• 期刊: Pediatric research.
• 作者: Tan,JP;Seidler,FJ;Schwinn,DA;Page,SO;Slotkin,TA
• 通讯作者: Slotkin,TA

Acute agonist-mediated desensitization of the human alpha 1a-adrenergic receptor is primarily independent of carboxyl terminus regulation: implications for regulation of alpha 1aAR splice variants.

人α1a-肾上腺素能受体的急性激动剂介导的脱敏主要独立于羧基末端调节：对α1aAR剪接变体调节的影响。

• DOI: 10.1074/jbc.m111762200
• 发表时间: 2002
• 期刊: The Journal of biological chemistry
• 作者: Price,RReyn;Morris,DanielP;Biswas,Gopa;Smith,MichaelP;Schwinn,DebraA
• 通讯作者: Schwinn,DebraA

Cloning and pharmacological characterization of human alpha-1 adrenergic receptors: sequence corrections and direct comparison with other species homologues.

• 发表时间: 1995
• 期刊: The Journal of pharmacology and experimental therapeutics
• 作者: D. Schwinn;G. Johnston;S. O. Page;M. J. Mosley;K. Wilson;N. Worman;S. Campbell;M. Fidock;L. Furness;D. Parry-Smith