Novel roles of C1q tumor necrosis factor-related proteins (CTRPs) in relaxin receptor biology

C1q 肿瘤坏死因子相关蛋白 (CTRP) 在松弛素受体生物学中的新作用

• 批准号: RGPIN-2017-05485
• 负责人: Klonisch, Thomas
• 金额: $ 2.48万
• 依托单位: University of Manitoba
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=711669
• 关键词: Novel; roles; C1q; tumor; necrosis

I study the role of relaxin receptor and binding partners on cell differentiation. My lab established relaxin (RLX) and INSL3 as novel mediators of cell motility and extracellular matrix (ECM) invasion. We also demonstrated a new function of Insl5, another relaxin-like peptide, in glucose homeostasis in mice and identified enteroendocrine cells of the human and mouse large intestine as specific producers of INSL5. Recently, my lab discovered a new binding partner to relaxin receptor RXFP1. The secreted C1qTNF-related peptide CTRP8 protein is structurally distinct from RLX. CTRP8 binds to RXFP1 and activates signaling pathways resulting in enhanced cell migration and matrix invasion. This proposal is a continuation of my pioneering CTRP8-RXFP1 work and uses molecular modelling and targeted mutagenesis to identify specific CTRP8 residues important for the binding of RXFP1. I use in-vivo labeling to capture novel binding factors to intracellular domains of RXFP1 that trigger CTRP8-induced signaling events. We were the first to show a novel role of CTRP8-RXFP1 in the protection against DNA damage by increased expression and activity of defined DNA repair proteins. I will expand on this finding and determine the impact of CTRP8-RXFP1 on oxidative stress responses which occur frequently in cells. I will measure mitochondrial production of reactive oxidative radicals (ROS), assess mitochondrial genes, and quantify ROS-induced oxidative DNA base lesions using 3D nuclear imaging, mitochondrial gene expression profiling and protein analysis. I showed that CTRP8-RXFP1 engages STAT3 signaling and increases the expression of key proteins promoting DNA repair and cell survival. I will study the molecular mechanisms by which CTRP8-RXFP1 promotes cell survival and mutagenic DNA damage repair. My lab has compelling evidence implicating CTRP1, a close relative of CTRP8, as a new RXFP1 agonist. CTRP1 and CTRP8 share the RXFP1 binding domain and we showed that CTRP1 requires RXFP1 to enhance cell motility and activate specific signaling pathways. The broad tissue expression of CTRP1 may have important implications for RXFP1 signaling in tissues. Like for CTRP8-RXFP1, we will demonstrate the physical CTRP1-RXFP1 interaction and identify the role of CTRP1 in cell motility, matrix invasion and DNA protection. We use new Ctrp1 mouse strains (overexpressing and KO) to assess the protective functions of Ctrp1-Rxfp1 in bleomycin-induced lung fibrosis. We employ our ocular wound healing mouse model and determine the ability of CTRP1 and CTRP8 to repair ocular surface lesions. My proposal will generate key advances in this exciting emerging field of RXFP research and, for the tenure of this grant, will provide training opportunities for a total of three graduate students and up to five summer/COOP students.

我研究松弛素受体和结合伴侣在细胞分化中的作用。我的实验室建立了松弛素（RLX）和INSL 3作为细胞运动和细胞外基质（ECM）侵袭的新介质。我们还证明了另一种松弛素样肽Insl 5在小鼠葡萄糖稳态中的新功能，并鉴定了人和小鼠大肠的肠内分泌细胞作为INSL 5的特异性生产者。最近，我的实验室发现了松弛素受体RXFP 1的新结合伴侣。分泌的C1 qTNF相关肽CTRP 8蛋白在结构上不同于RLX。CTRP 8结合RXFP 1并激活信号通路，导致增强的细胞迁移和基质侵袭。该提案是我开创性的CTRP 8-RXFP 1工作的延续，并使用分子建模和靶向诱变来鉴定对RXFP 1结合重要的特定CTRP 8残基。我使用体内标记来捕获触发CTRP 8诱导的信号传导事件的RXFP 1细胞内结构域的新型结合因子。我们是第一个显示CTRP 8-RXFP 1在通过增加定义的DNA修复蛋白的表达和活性来保护DNA损伤中的新作用的人。我将扩展这一发现，并确定CTRP 8-RXFP 1对细胞中经常发生的氧化应激反应的影响。我将测量反应性氧化自由基（ROS）的线粒体产生，评估线粒体基因，并使用3D核成像，线粒体基因表达谱和蛋白质分析来量化ROS诱导的氧化DNA碱基损伤。我发现CTRP 8-RXFP 1参与STAT 3信号传导，并增加促进DNA修复和细胞存活的关键蛋白的表达。我将研究CTRP 8-RXFP 1促进细胞存活和诱变DNA损伤修复的分子机制。我的实验室有令人信服的证据表明CTRP 1是CTRP 8的近亲，是一种新的RXFP 1激动剂。CTRP 1和CTRP 8共享RXFP 1结合结构域，我们发现CTRP 1需要RXFP 1来增强细胞运动性并激活特定的信号通路。CTRP 1的广泛组织表达可能对组织中的RXFP 1信号传导具有重要意义。与CTRP 8-RXFP 1一样，我们将证明CTRP 1-RXFP 1的物理相互作用，并确定CTRP 1在细胞运动性、基质侵袭和DNA保护中的作用。我们使用新的Ctrp 1小鼠品系（过表达和KO）来评估Ctrp 1-Rxfp 1在博来霉素诱导的肺纤维化中的保护功能。我们采用我们的眼创伤愈合小鼠模型并确定CTRP 1和CTRP 8修复眼表病变的能力。我的提议将在RXFP研究这一令人兴奋的新兴领域取得关键进展，并在此赠款的任期内，将为总共三名研究生和多达五名暑期/COOP学生提供培训机会。