Mechanisms Regulating GPCR Trafficking

GPCR 贩运的监管机制

• 批准号: 8209041
• 负责人: Joann Trejo
• 金额: $ 32.5万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8209041
• 关键词: Adaptor Protein Complex 3; Arrestins; Back; Binding; Biochemical; Biological Assay; Biological Models; Cardiovascular Diseases; Cell surface; Cells; Ceramides; Clathrin; Clathrin Adaptors; Complex; Cytoplasmic Tail; Degradation Pathway; Disease; Drug usage; Dynamin; Endosomes; Family; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Proteins; Gastrointestinal Diseases; Human; Immune System Diseases; Immunoelectron Microscopy; Kidney Diseases; Life; Lung diseases; Lysosomes; Mediating; Mental disorders; Molecular; Multivesicular Body; Nerve Degeneration; PAR-1 Receptor; PAR-2 Receptor; Pathway interactions; Phosphorylation; Physiological; Prevention strategy; Process; Proteins; Receptor Signaling; Recycling; Regulation; Signal Transduction; Small Interfering RNA; Sorting - Cell Movement; Stimulus; System; Tyrosine; Ubiquitin; Ubiquitination; Vesicle; base; coated pit; desensitization; fluorescence imaging; human disease; inhibitor/antagonist; insight; mammalian genome; mutant; nexin; novel; public health relevance; receptor; research study; response; trafficking; tumor progression

DESCRIPTION (provided by applicant): G protein-coupled receptors (GPCRs) comprise the largest family of cell surface signaling receptors in the mammalian genome, mediate cellular responses to diverse stimuli and control a vast physiological responses. Dysregulated GPCR signaling has been implicated in multiple human pathological conditions, making this receptor class the target of nearly half the drugs used clinically. In addition to rapid desensitization, GPCR trafficking is crucial for the temporal and spatial control of receptor signaling. However, the mechanisms responsible for trafficking of GPCRs through the endocytic system remains poorly understood. Internalization of activated GPCRs occurs through a clathrin- and dynamin-dependent pathway that requires arrestins. However, arrestins are not essential for internalization of all GPCRs. Several GPCRs including protease- activated receptor-1 (PAR1) internalize through a clathrin- and dynamin-dependent pathway independent of arrestins, suggesting that distinct mechanisms regulate internalization of different GPCRs. Once internalized, GPCRs are dephosphorylated and recycled back to the cell surface in a resensitized state competent to signal again or sorted to a lysosomal degradation pathway, a process critical for termination of receptor signaling. Other GPCRs remain in endocytic compartments and signal independent of G-proteins. We previously showed that unlike most GPCRs, which internalize and recycle, activated PAR1 is internalized, sorted directly to lysosomes and degraded; a process critical for shutting-off activated PAR1 signaling. The efficiency with which PARs are degraded makes this receptor class an excellent model system to investigate the molecular basis of GPCR lysosomal degradation. Many GPCRs including protease-activated receptor-2 (PAR2) are modified with ubiquitin, which facilitates lysosomal trafficking through the endosomal-sorting complex required for transport (ESCRT) pathway. However, several GPCRs sort to lysosomes independent of ubiquitination and some components of the ESCRT complex. Remarkably, we found that activated PAR1 trafficks from endosomes to lysosomes independent of ubiquitination and ubiquitin-binding ESCRT components. The molecular mechanism by which PAR1 and other GPCRs sort to lysosomes independent of ubiquitination is not known. This proposal is focused on delineating the molecular mechanisms that regulate ubiquitin-independent lysosomal sorting of GPCRs. The specific aims of the proposal are to: 1) determine whether GPCRs that sort through ubiquitin- dependent versus -independent lysosomal pathways internalize through the same or distinct clathrin-coated pits, and sort through the same or distinct early or late endosomal compartments, 2) define the sorting signals and mechanism for the novel ubiquitin-independent endosome-to-lysosome sorting of GPCRs, and 3) delineate the molecular pathway by which GPCRs uniquely sort to intralumenal vesicles (ILVs) of multivesicular bodies (MVBs) independent of ubiquitination and the ubiquitin-binding ESCRT components. PUBLIC HEALTH RELEVANCE: G protein-coupled receptors (GPCRs) signal in response to diverse stimuli, control vast physiological responses and are implicated in multiple human pathological diseases. In addition to rapid GPCR desensitization, GPCR trafficking is important for the spatial and temporal control of receptor signaling and dysregulated trafficking contributes to aberrant signaling and disease. Thus, understanding the mechanisms that control trafficking of GPCRs, and developing the ability to manipulate it, may provide new strategies for the prevention and treatment of a wide range of human diseases.

描述（由申请人提供）：G蛋白偶联受体（GPCR）包括哺乳动物基因组中最大的细胞表面信号受体家族，介导细胞对各种刺激的反应并控制广泛的生理反应。 GPCR信号失调已与多种人类病理条件有关，使该受体类别成为临床上使用的几乎一半的药物的靶标。除了快速脱敏外，GPCR运输对于受体信号的时间和空间控制至关重要。但是，负责通过内吞系统贩运GPCR的机制仍然很少理解。激活的GPCR的内在化是通过需要阻止蛋白的网状蛋白和动力蛋白依赖性途径发生的。但是，逮捕蛋白对于所有GPCR的内在化并不是必不可少的。包括蛋白酶活化受体-1（PAR1）在内的几种GPCR通过甲状腺激素和动力蛋白依赖性途径内部化，独立于停滞蛋白，这表明不同的机制调节了不同GPCR的内在化。一旦内化，GPCR将被脱磷酸化，并以敏感的状态恢复回到细胞表面，该状态有能力再次信号或分类为溶酶体降解途径，这是终止受体信号传导的关键过程。其他GPCR仍保留在内吞区室和与G蛋白无关的信号。我们先前表明，与大多数内化和回收的GPCR不同，活化的PAR1被内在化，直接分类为溶酶体并降解。关闭激活的PAR1信号传导至关重要的过程。降解PAR的效率使该受体类成为研究GPCR溶酶体降解的分子基础的出色模型系统。许多GPCR（包括蛋白酶激活的受体-2（PAR2））通过泛素修饰，这有助于通过运输（ESCRT）途径所需的内体分类复合物进行溶酶体运输。但是，几个GPCR与泛素化和ESCRT复合物的某些成分无关。值得注意的是，我们发现，从内体到溶酶体激活的PAR1运输，独立于泛素化和泛素结合的ESCRT成分。尚不清楚pAR1和其他GPCR与独立于泛素化的溶酶体对溶酶体排序的分子机制。该提案的重点是描述调节GPCR的泛素依赖性溶酶体分选的分子机制。该提案的具体目的是：1）确定通过泛素依赖性与独立的溶酶体途径进行排序的GPCR是否通过相同或不同的网格蛋白涂层的凹坑进行内部化，并通过相同或不同的早期或晚期内体隔室进行分类，2）定义与依赖性的ubiquiquil-Intose-intose依赖性的ubiquiquil-Intose unientose-intosem-Intosem-Intosem-Intosem-Intosem-Intose-Intosemissismiss和GPCR和3）描绘了GPCR的分子途径，该途径独特地将其独特地排序到多囊体（MVB）的量没有与泛素化和泛素结合的ESCRT组件的多囊体囊泡（ILV）。 公共卫生相关性：G蛋白偶联受体（GPCR）响应各种刺激，控制广泛的生理反应，并与多种人类病理疾病有关。除了快速的GPCR脱敏之外，GPCR运输对于受体信号传导和失调运输失调的空间和时间控制也很重要，这会导致异常信号传导和疾病。因此，了解控制GPCR的运输的机制以及发展操纵它的能力，可以为预防和治疗广泛的人类疾病提供新的策略。