ENDOPEPTIDASES AFFECT G-PROTEIN COUPLED RECEPTOR SIGNALING AND RESENSITIZATION

内肽酶影响 G 蛋白偶联受体信号传导和再敏化

• 批准号: 8363772
• 负责人: NIGEL W BUNNETT
• 金额: $ 0.02万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363772
• 关键词: Affect; Agonist; Big Endothelin; Biological; Bradykinin; Cell membrane; Cell surface; Complex; Degradation Pathway; Disease; Dissociation; Endocytosis; Endopeptidases; Endosomes; Endothelin; Endothelin-1; Environment; Family; Functional disorder; Funding; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Grant; Heterotrimeric GTP-Binding Proteins; In Vitro; Inflammation; Mass Spectrum Analysis; National Center for Research Resources; Neprilysin; Neuropeptides; Peptide Hydrolases; Principal Investigator; Process; Receptor Activation; Recycling; Research; Research Infrastructure; Resources; Role; Signal Transduction; Sorting - Cell Movement; Source; Substance P; Substance P Receptor; Substrate Specificity; United States National Institutes of Health; Vasoconstrictor Agents; cost; desensitization; member; prevent; receptor; receptor recycling; research study

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. G-protein coupled receptor (GPCR) activation by neuropeptides is terminated by either cell-surface peptidase activity or receptor desensitization, and thereby prevents uncontrolled signaling that may cause dysfunction and disease. Cell-surface peptidases such as neutral endopeptidase (NEP) degrade extra-cellular neuropeptides and terminate their biological actions. For example, NEP degrades substance P (SP) and terminates inflammation by limiting the activation of the neurokinin-1 receptor (NK1R). Alternatively, activated GPCR signaling is terminated by desensitization, a process that entails uncoupling of heterotrimeric G-proteins and endocytosis of the receptor-agonist complex into endosomes where the acidic environment of the endosomes facilitates dissociation of the receptor and agonist. While the receptor is sorted to either recycling or degradative pathways by poorly understood mechanisms, the ultimate fate of the agonist after endocytosis is unknown. Endothelin-converting enzyeme-1 (ECE-1) is a member of the M-13 family of endopeptidases that also includes NEP. While NEP is confined to the plasma membrane, ECE-1 has a broad subcellular distribution that includes both the plasma membrane and endosomes. The primary natural substrate of ECE-1 expressed at the plasma membrane is big endothelin (big-ET). Big-ET is hydrolyzed by ECE-1 at the plasma membrane to form endothelin-1 (ET), a potent vasoconstrictor. While ECE-1 hydrolyzes a variety of neuropeptides such as substance-P and bradykinin in in-vitro experiments, the biological significance of this activity is unknown. Furthermore, the role of ECE-1 contained in endosomes has not been explored. The aim of this project is to study the role of ECE-1 in GPCR signaling. Activation of GPCRs results in endocytosis of the receptor and agonist into acidified endosomes. We propose that endopeptidases such as ECE-1 are also present in these acidified endosomes where they degrade the neuropeptide agonists and thereby control receptor recycling. Mass spectrometry will allow for characterization of endosomal ECE-1 activity and determine its substrate specificity.

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 神经肽对G蛋白偶联受体的激活可通过细胞表面多肽酶活性或受体脱敏来终止，从而防止可能导致功能障碍和疾病的失控信号。中性内肽酶(NEP)等细胞表面肽酶能降解胞外神经肽并终止其生物学作用。例如，NEP通过限制神经激肽-1受体(NK1R)的激活来降解P物质(SP)并终止炎症。或者，激活的GPCR信号通过脱敏终止，这一过程需要异源三聚体G蛋白的解偶联和受体-激动剂复合体的内吞作用，在内体的酸性环境中有利于受体和激动剂的解离。虽然受体通过鲜为人知的机制被分类为循环或降解途径，但激动剂在内吞作用后的最终命运尚不清楚。内皮素转换酶-1(ECE1)是M-13内肽酶家族中的一员，也包括NEP。虽然NEP仅限于质膜，但ECA-1具有广泛的亚细胞分布，包括质膜和内小体。ECE1在细胞膜上表达的主要天然底物是大内皮素(BIG-ET)。BIG-ET被ECE1在质膜上水解，形成内皮素-1(ET)，这是一种有效的血管收缩因子。在体外实验中，ECE1对P物质和缓激肽等多种神经肽进行了水解性研究，但其生物学意义尚不清楚。此外，内容体中所含的ECA-1的作用还没有被探索过。本项目的目的是研究ECE1在GPCR信号转导中的作用。GPCRs的激活导致受体和激动剂的内吞作用进入酸化的内涵体。我们认为，在这些酸化的内体中也存在内肽酶，如ECA-1，它们在那里降解神经肽激动剂，从而控制受体的循环。质谱学将能够表征内体ECE1的活性，并确定其底物专一性。