BETA-ADRENERGIC RECEPTOR STRUCTURE AND DESENSITIZATION

β-肾上腺素能受体结构和脱敏

基本信息

项目摘要

The regulation of cellular processes in mammals by epinephrine stimulation of beta2-adrenergic receptors (betaAR) is extensive and includes important effects on the nervous, cardiovascular, and pulmonary systems. BetaAR antagonists are used as therapy for several diseases of the cardiovascular system such as hypertension, and betaAR agonists are a mainstay in the treatment of asthma. One of the most important factors that govern the effectiveness of betaAR agonists is the rate of development of tolerance, which is thought to be caused in part by desensitization of the betaAR. Desensitization, the loss of betaAR stimulation of adenylyl cyclase (AC), appears to be caused by phosphorylation of the betaAR by various protein kinases, by internalization or by downregulation. Our long-term goal is to determine the roles these mechanisms play in the regulation of betaAR responsivity in mammalian tissue. The major goal of this proposal is to identify the specific amino acids in the betaAR that are phosphorylated in response to stimuli that activate PKA-, PKC- or betaARK-mediated mechanisms of desensitization in intact human embryonic kidney (HEK 293) cells, and the consequences of the phosphorylations on betaAR activation of AC. The primary focus will be on the putative betaARK consensus phosphorylation sites. Site-directed mutagenesis of proposed protein kinase consensus sites in the betaAR and the characterization of the mutations' effects on betaAR stimulation of AC following expression in HEK 293 cells will localize the domains involved in the kinase-mediated desensitizations. This analysis will be coupled with peptide mapping and microsequencing of highly purified betaAR phosphopeptides to determine the specific amino acids phosphorylated by the various kinases. Purification of the betaAR for peptide mapping will be based on the use of hemagglutinin and 6-histidine epitopes which have been introduced into the betaAR. Secondary aims include: analysis of the relative roles of betaARK and internalization in homologous desensitization and their interdependence; exploration of internalization domains by site-directed mutagenesis; investigation of the role of the C-terminal PKA/PKC consensus site; tests of our proposal that partial agonists induce less homologous desensitization than do full agonists; and an exact quantitation of the effect of receptor number on desensitization.
肾上腺素刺激对哺乳动物细胞过程的调节 β 2-肾上腺素能受体(β AR)是广泛的,包括重要的 对神经、心血管和肺系统的影响。 BetaAR 拮抗剂被用作多种心血管疾病的治疗 系统,如高血压,和β AR激动剂是一个支柱, 治疗哮喘。 决定β AR有效性的最重要因素之一 激动剂是耐受性的发展速度,这被认为是 部分原因是β AR脱敏。 脱敏 腺苷酸环化酶(AC)β AR刺激的丧失似乎是由 通过各种蛋白激酶磷酸化β AR, 内化或下调。 我们的长期目标是确定 这些机制在调节β AR反应性中的作用 在哺乳动物组织中。 该提案的主要目标是确定特定的氨基酸, β-AR对刺激的反应是磷酸化, PKA、PKC或β-ARK介导的完整细胞脱敏机制 人胚肾(HEK 293)细胞,以及 磷酸化对AC的β AR活化的影响。 主要重点将放在 假定的betaARK共有磷酸化位点。 定点 β AR中建议的蛋白激酶共有位点的诱变, 突变对AC的β AR刺激作用的表征 在HEK 293细胞中表达后, 在激酶介导的脱敏中。 这一分析将与 通过肽图谱和高度纯化的β AR的微测序, 磷酸肽,以确定特定的氨基酸磷酸化, 各种激酶。 用于肽图谱的β AR的纯化将 基于使用血凝素和6-组氨酸表位, 被引入β AR。 次要目的包括:分析betaARK和 同源脱敏中的内化及其相互依赖; 通过定点诱变探索内化结构域; C-末端PKA/PKC共有位点作用的研究;试验 我们的建议,部分激动剂诱导较少的同源 比完全激动剂更敏感;以及 受体数目对脱敏的影响。

项目成果

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RICHARD B CLARK其他文献

RICHARD B CLARK的其他文献

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{{ truncateString('RICHARD B CLARK', 18)}}的其他基金

SMALL INSTRUMENTATION GRANT
小型仪器补助金
  • 批准号:
    3524967
  • 财政年份:
    1992
  • 资助金额:
    $ 29.55万
  • 项目类别:
STRUCTURE/FUNCTION OF THE TSH RECEPTOR
TSH 受体的结构/功能
  • 批准号:
    3023297
  • 财政年份:
    1991
  • 资助金额:
    $ 29.55万
  • 项目类别:
SMALL INSTRUMENTATION PROGRAM
小型仪器项目
  • 批准号:
    3524771
  • 财政年份:
    1989
  • 资助金额:
    $ 29.55万
  • 项目类别:
BETA-ADRENERGIC RECEPTOR STRUCTURE AND DESENSITIZATION
β-肾上腺素能受体结构和脱敏
  • 批准号:
    2684744
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
BETA-ADRENERGIC RECEPTOR STRUCTURE AND DESENSITIZATION
β-肾上腺素能受体结构和脱敏
  • 批准号:
    3279137
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
B-ADRENERGIC RECEPTOR STUCTURE & DESENSITIZATION
B-肾上腺素能受体结构
  • 批准号:
    3279136
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
BETA-ADRENERGIC RECEPTOR STRUCTURE AND DESENSITIZATION
β-肾上腺素能受体结构和脱敏
  • 批准号:
    6606900
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
Beta Andrenergic Receptor Structure and Desensitization
β 肾上腺素能受体结构和脱敏
  • 批准号:
    7937878
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
Beta Adrenergic Receptor Structure and Desensitization
β 肾上腺素能受体结构和脱敏
  • 批准号:
    6871479
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
B-ADRENERGIC RECEPTOR STUCTURE & DESENSITIZATION
B-肾上腺素能受体结构
  • 批准号:
    3279141
  • 财政年份:
    1983
  • 资助金额:
    $ 29.55万
  • 项目类别:
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