Regulation of Melanoma Cell Malignancy by Compartmentalized Chemokine Receptor Signaling

通过区室化趋化因子受体信号传导调节黑色素瘤细胞恶性肿瘤

• 批准号: 10528741
• 负责人: Alex Rojas Bie Thomsen
• 金额: $ 41.27万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-23 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10528741
• 关键词: Agonist; Anatomy; Arrestins; Binding Sites; CCL19 gene; CCL21 gene; Calcium; Cell Differentiation process; Cell Maturation; Cell Proliferation; Cell membrane; Cells; Cholesterol; Complex; Cryoelectron Microscopy; Deformity; Disease; Drug Prescriptions; Drug Targeting; Endocytosis; Endosomes; Environment; G protein coupled receptor kinase; G-Protein-Coupled Receptors; GTP-Binding Proteins; Gene Expression; Genes; Genetic Transcription; Growth; Heterotrimeric GTP-Binding Proteins; Human; Immune; Infection; Inflammation; Invaded; Kidney; Lead; Ligands; Malignant - descriptor; Malignant Neoplasms; Melanoma Cell; Metastatic Neoplasm to Lymph Nodes; Nature; Neoplasm Metastasis; Pathogenicity; Pathologic; Pathway interactions; Patients; Phosphorus; Phosphorylation; Physiological; Production; Proliferating; Proteins; Receptor Signaling; Regulation; Reporting; Resolution; Role; Serum; Signal Transduction; Signaling Protein; Skin Cancer; Structure; Testing; Tissues; Transforming Growth Factors; Tumor-Derived; Up-Regulation; Water; base; cancer cell; cell motility; chemokine; chemokine receptor; cholesterol biosynthesis; chronic pain; desensitization; drug discovery; endosome membrane; fighting; malignant phenotype; melanoma; member; novel; receptor; receptor expression; receptor internalization; recruit; response; targeted treatment; therapeutic target; tumor

Project Summary Melanoma is the deadliest form of skin cancer with few treatment options to patients with advanced metastatic disease. Melanoma cell proliferation, survival, invasion, and metastasis to lymph nodes correlates with expression of the chemokine receptor CCR7. CCR7 is a member of the highly druggable G protein-coupled receptors (GPCRs) superfamily and promotes cell migration and maturation of immune cells. Classically, upon agonist stimulation, GPCRs at the cell membrane activate heterotrimeric G proteins, causing downstream signaling throughout the cell. In order to terminate G protein signaling, cells have devised a specialized desensitization mechanism that includes receptor phosphorylation by GPCR kinases and subsequent recruitment of β-arrestins (βarrs) to the phosphorylated receptors. The GPCR–βarrs interaction both blocks the G protein-binding site and promotes receptor endocytosis. However, we recently discovered that some GPCRs interact with G proteins and βarrs simultaneously to form GPCR–G protein–βarr `megaplexes', which allows the receptor to continue to stimulate G protein signaling while being internalized into endosomes by βarrs. Our preliminary results suggest that CCR7 forms such megaplexes and continues to stimulate G protein signaling after having been internalized. The proposed project aims to explore the involvement of this endosomal CCR7 signaling in malignant progression of melanoma cells. To this end, we have developed an experimental approach to separate pathophysiological functions initiated by plasma membrane CCR7 signaling from endosomal CCR7 signaling. This approach leverages two natural chemokine ligands, CCL19 and CCL21, which predominately activate G proteins from endosomes or plasma membrane, respectively. Using these chemokines to stimulate CCR7 in HEK293 cells, we demonstrate that signaling from different cellular compartments promote distinct transcriptional changes. In particular, we observed upregulation of key genes involved in the cholesterol biosynthesis in response to endosomal CCR7 signaling whereas plasma membrane CCR7 signaling enhanced the expression of inhibitory SMADs and other genes related to the transforming growth factor-β (TGF-β) pathway. In melanoma, these two pathways are reported to have opposite effects with cholesterol production accelerating melanoma cell proliferation and inhibitory SMADs limiting growth. Therefore, we here propose to test the hypothesis that endosomal CCR7 signaling in melanoma specifically drives pathogenic cell proliferation through enhanced cholesterol biosynthesis. The results from this project will not only enhance our understanding of how chemokine receptor expression promotes melanoma malignancy but might also provide a new strategy to develop effective melanoma therapeutics by targeting chemokine receptors at endosomal compartments.

项目概要 黑色素瘤是最致命的皮肤癌，晚期转移性患者的治疗选择很少 疾病。黑色素瘤细胞增殖、存活、侵袭和转移至淋巴结与 趋化因子受体 CCR7 的表达。 CCR7 是高度药物化的 G 蛋白偶联蛋白的成员 受体（GPCR）超家族并促进细胞迁移和免疫细胞的成熟。经典地讲，根据 激动剂刺激下，细胞膜上的 GPCR 激活异三聚体 G 蛋白，导致下游 整个细胞内的信号传导。为了终止 G 蛋白信号传导，细胞设计了一种专门的 脱敏机制，包括 GPCR 激酶的受体磷酸化和随后的 将 β-抑制蛋白 (βarrs) 募集至磷酸化受体。 GPCR-βarrs 相互作用都阻断 G 蛋白结合位点并促进受体内吞作用。然而，我们最近发现一些 GPCR 同时与G蛋白和βarrs相互作用形成GPCR-G蛋白-βarr‘megaplexes’，这使得 受体继续刺激 G 蛋白信号传导，同时被 βarrs 内化到核内体中。我们的 初步结果表明 CCR7 形成这种巨复合物并继续刺激 G 蛋白信号传导 被内化后。拟议的项目旨在探索这种内体 CCR7 的参与 黑色素瘤细胞恶性进展中的信号传导。为此，我们开发了一种实验方法 将质膜 CCR7 信号传导启动的病理生理功能与内体 CCR7 分开 发信号。这种方法利用了两种天然趋化因子配体 CCL19 和 CCL21，它们主要 分别激活来自内体或质膜的 G 蛋白。利用这些趋化因子来刺激 HEK293 细胞中的 CCR7，我们证明来自不同细胞区室的信号传导促进不同的 转录变化。特别是，我们观察到与胆固醇相关的关键基因的上调 生物合成响应内体 CCR7 信号传导，而质膜 CCR7 信号传导增强 抑制性 SMAD 和其他与转化生长因子-β (TGF-β) 途径相关的基因的表达。 据报道，在黑色素瘤中，这两种途径具有相反的作用，会加速胆固醇的产生 黑色素瘤细胞增殖和抑制性 SMAD 限制生长。因此，我们在这里建议测试 假设黑色素瘤中的内体 CCR7 信号传导通过以下方式特异性驱动致病细胞增殖： 增强胆固醇的生物合成。该项目的结果不仅将增强我们对如何 趋化因子受体表达促进黑色素瘤恶性肿瘤，但也可能提供一种新策略 通过靶向内体区室的趋化因子受体来开发有效的黑色素瘤治疗方法。

项目成果

Arrestin-biased AT1R agonism induces acute catecholamine secretion through TRPC3 coupling.

抑制蛋白偏向的 AT1R 激动作用通过 TRPC3 偶联诱导急性儿茶酚胺分泌

• DOI: 10.1038/ncomms14335
• 发表时间: 2017-02-09
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Liu CH;Gong Z;Liang ZL;Liu ZX;Yang F;Sun YJ;Ma ML;Wang YJ;Ji CR;Wang YH;Wang MJ;Cui FA;Lin A;Zheng WS;He DF;Qu CX;Xiao P;Liu CY;Thomsen AR;Joseph Cahill T 3rd;Kahsai AW;Yi F;Xiao KH;Xue T;Zhou Z;Yu X;Sun JP
• 通讯作者: Sun JP

Novel strategies in drug discovery of the calcium-sensing receptor based on biased signaling.

基于偏向信号传导的钙敏感受体药物发现新策略。

• DOI: 10.2174/138945012802429642
• 发表时间: 2012
• 期刊: Current drug targets
• 影响因子: 3.2
• 作者: Thomsen,AlexRojasBie;Smajilovic,Sanela;Bräuner-Osborne,Hans
• 通讯作者: Bräuner-Osborne,Hans

G protein-coupled receptor signaling analysis using homogenous time-resolved Förster resonance energy transfer (HTRF®) technology.

• DOI: 10.3390/ijms15022554
• 发表时间: 2014-02-13
• 期刊: International journal of molecular sciences
• 影响因子: 5.6
• 作者: Nørskov-Lauritsen L;Thomsen AR;Bräuner-Osborne H
• 通讯作者: Bräuner-Osborne H

Gq activity- and β-arrestin-1 scaffolding-mediated ADGRG2/CFTR coupling are required for male fertility.

Gq 活性和 beta-arrestin-1 支架介导的 ADGRG2/CFTR 偶联是男性生育能力所必需的

• DOI: 10.7554/elife.33432
• 发表时间: 2018-02-02
• 期刊: eLife
• 影响因子: 7.7
• 作者: Zhang DL;Sun YJ;Ma ML;Wang YJ;Lin H;Li RR;Liang ZL;Gao Y;Yang Z;He DF;Lin A;Mo H;Lu YJ;Li MJ;Kong W;Chung KY;Yi F;Li JY;Qin YY;Li J;Thomsen ARB;Kahsai AW;Chen ZJ;Xu ZG;Liu M;Li D;Yu X;Sun JP
• 通讯作者: Sun JP