Role of Sam68 in Proinflammatory Signaling

Sam68 在促炎信号传导中的作用

基本信息

批准号：
10446490
负责人：
Parameswaran Ramakrishnan
金额：
$ 56.51万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-01 至 2026-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10446490
关键词：
3-Dimensional Acute Address Adverse event Affect Apoptosis Binding Biochemical Biological Assay Biological Response Modifier Therapy Bone Marrow Transplantation CRISPR/Cas technology Cell Line Cells Chronic Coculture Techniques Colitis Colon Colonic inflammation Complex DNA-Protein Interaction Deletion Mutation Development Digestive System Disorders Disease remission Environmental Risk Factor Epithelial Cell Proliferation Epithelial Cells Future Gel Gene Expression Genes Genetic Genetic Transcription Growth Hematopoietic Human Immune Incidence Infection Inflammation Inflammation Mediators Inflammatory Inflammatory Bowel Diseases Intestinal permeability Intestines Knock-out Knockout Mice Knowledge Lead Malignant Neoplasms Mediating Mediator of activation protein Membrane Modeling Molecular Mucositis Mucous Membrane Mus Organoids Oxazolone Partial Remission Pathogenesis Pathogenicity Pathologic Pathway interactions Patients Permeability Persons Play Point Mutation Post-Translational Protein Processing Proctitis Proteins RNA Binding RNA-Binding Proteins Receptor Signaling Refractory Relapse Research Risk Role SRC-associated p68 protein Signal Pathway Signal Transduction Signaling Protein Sodium Dextran Sulfate Spontaneous colitis Structure-Activity Relationship System TLR2 gene TNF gene Testing Therapeutic Tissues Toll-Like Receptor Pathway Toll-like receptors Transcriptional Activation Tumor Necrosis Factor Receptor Ulcerative Colitis United States Wiskott-Aldrich Syndrome Work base chemically induced colitis chromatin immunoprecipitation chronic inflammatory disease cohort conditional knockout cytokine design experimental study gut inflammation improved in vivo intestinal epithelium intestinal homeostasis murine colitis new therapeutic target novel novel therapeutic intervention pre-clinical protein activation protein protein interaction targeted treatment tool transcription factor transcriptome sequencing treatment strategy vector

3-Dimensional Acute Address Adverse event Affect Apoptosis Binding Biochemical Biological Assay Biological Response Modifier Therapy Bone Marrow Transplantation CRISPR/Cas technology Cell Line Cells Chronic Coculture Techniques Colitis Colon Colonic inflammation Complex DNA-Protein Interaction Deletion Mutation Development Digestive System Disorders Disease remission Environmental Risk Factor Epithelial Cell Proliferation Epithelial Cells Future Gel Gene Expression Genes Genetic Genetic Transcription Growth Hematopoietic Human Immune Incidence Infection Inflammation Inflammation Mediators Inflammatory Inflammatory Bowel Diseases Intestinal permeability Intestines Knock-out Knockout Mice Knowledge Lead Malignant Neoplasms Mediating Mediator of activation protein Membrane Modeling Molecular Mucositis Mucous Membrane Mus Organoids Oxazolone Partial Remission Pathogenesis Pathogenicity Pathologic Pathway interactions Patients Permeability Persons Play Point Mutation Post-Translational Protein Processing Proctitis Proteins RNA Binding RNA-Binding Proteins Receptor Signaling Refractory Relapse Research Risk Role SRC-associated p68 protein Signal Pathway Signal Transduction Signaling Protein Sodium Dextran Sulfate Spontaneous colitis Structure-Activity Relationship System TLR2 gene TNF gene Testing Therapeutic Tissues Toll-Like Receptor Pathway Toll-like receptors Transcriptional Activation Tumor Necrosis Factor Receptor Ulcerative Colitis United States Wiskott-Aldrich Syndrome Work base chemically induced colitis chromatin immunoprecipitation chronic inflammatory disease cohort conditional knockout cytokine design experimental study gut inflammation improved in vivo intestinal epithelium intestinal homeostasis murine colitis new therapeutic target novel novel therapeutic intervention pre-clinical protein activation protein protein interaction targeted treatment tool transcription factor transcriptome sequencing treatment strategy vector

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项目摘要

Abstract Ulcerative colitis (UC) is a chronic form of inflammatory bowel disease (IBD) with no cure. Current treatment strategies offer only partial remission, with many patients remaining refractory to treatment, and carry risks of significant adverse events including serious infections and cancer. Thus, an improved understanding of inflammatory signaling pathways and identification of novel preclinical mechanisms are critical challenges in IBD research. Signaling downstream of the proinflammatory cytokine tumor necrosis factor (TNF) and toll like receptor (TLR) pathways play major roles in IBD pathogenesis. In previous work, we discovered that the RNA binding protein Sam68 is required for both TNF- and TLR-induced activation of the transcription factor NF-κB, a master regulator of inflammation, suggesting that Sam68 contributes to NF-κB-dependent inflammation. Our new preliminary results show that Sam68 is prominently expressed in human and murine intestinal epithelial cells (IEC); Sam68 knockout (KO) mice are significantly protected from dextran sulfate sodium (DSS)- and oxazolone-induced colitis; and Sam68 protein is significantly elevated in inflamed colons of UC patients. Based on this, we hypothesize that Sam68 is a critical mediator of inflammation in IECs, and targeting IEC Sam68 will provide a novel therapeutic strategy in UC via dual inhibition of TNF- and TLR-mediated inflammatory pathways. We propose to study the molecular mechanisms of IEC-specific Sam68 signaling in human and experimental murine colitis using cutting edge tools such as 3D colonoids derived from UC patient colonocytes and novel IEC-specific Sam68 conditional KO (cKO) mice. Aim 1 will delineate molecular mechanisms of IEC-specific Sam68 signaling in TNF- and TLR-induced inflammatory signaling and identify the structural domains and posttranslational modifications of Sam68 required for its inflammatory functions. Aim 2 will study the in vivo role(s) of Sam68 in primary IECs under homeostatic and inflammatory conditions, using Sam68-cKO mice challenged with DSS and oxazolone as experimental colitis models, and using a spontaneous colitis model, Sam68-cKO / Wiscott Aldrich Syndrome Protein (WASP) KO double KO mice. This aim will utilize 3D colonoids prepared from wild type and Sam68-KO mice to study role of Sam68 in IEC proliferation, permeability and inflammatory signaling to study the homeostatic role of Sam68. Aim 3 will delineate the proinflammatory role of Sam68 in UC patients. This aim will study IEC-specific growth, proliferation, apoptosis, and expression of TJ proteins, and activation of proinflammatory signaling in IEC's and mucosal immune cells using co-culture assays of control and patient derived 3D intestinal organoids and immune cells. Successful completion of this study will fill critical knowledge gaps in understanding novel mechanisms underlying TNF- and TLR-dependent inflammatory signaling in UC and pave the way for the development of novel therapeutics targeting Sam68 to treat UC. Moreover, this study will also serve as the basis to explore the role of Sam68 in other TNF- and TLR-dependent chronic inflammatory diseases as well.

抽象的溃疡性结肠炎（UC）是炎症性肠病（IBD）的一种慢性形式，无法治愈。当前治疗策略仅提供部分缓解，许多患者仍然对治疗难治性，并承担重大不良事件，包括严重的感染和癌症。这是对炎症信号通路和新型临床前机制的识别是关键的挑战 IBD研究。促炎细胞因子肿瘤坏死因子（TNF）和TOLL的信号传导下游受体（TLR）途径在IBD发病机理中起主要作用。在以前的工作中，我们发现RNA 结合蛋白SAM68是TNF和TLR诱导的转录因子NF-κB激活所必需的，炎症的主要调节剂，表明SAM68有助于NF-κB依赖性炎症。我们的新初步结果表明，SAM68在人类和鼠肠中突出表达上皮细胞（IEC）； SAM68敲除（KO）小鼠受到明显保护的免受硫酸葡萄糖钠的保护（DSS）和恶唑龙诱导的结肠炎； SAM68蛋白在UC发炎的结肠中显着升高患者。基于此，我们假设SAM68是IEC炎症的关键调解人，并且靶向IEC SAM68将通过双重抑制TNF-和 TLR介导的炎症途径。我们建议研究IEC特异性的分子机制 SAM68使用尖端工具（例如3D菌囊）在人和实验鼠结肠炎中发出信号传导源自UC患者结肠细胞和新型IEC特异性SAM68条件KO（CKO）小鼠。目标1意志 TNF和TLR诱导的炎症中IEC特异性SAM68信号传导的分子机制信号传导并确定SAM68所需的结构域和翻译后修饰炎症功能。 AIM 2将研究SAM68在稳态下的主要IEC中的体内角色炎症条件，使用DSS和恶唑龙挑战的SAM68-CKO小鼠作为实验性结肠炎模型，并使用赞助结肠炎模型，SAM68-CKO / Wiscott Aldrich综合征蛋白（WASP）KO 双KO老鼠。该目标将利用从野生型和SAM68-KO小鼠制备的3D菌囊来研究角色 SAM68在IEC增殖，渗透性和炎症信号传导中研究SAM68的稳态作用。 AIM 3将描述SAM68在UC患者中的促炎作用。这个目标将研究IEC特定的增长， TJ蛋白的增殖，凋亡和表达以及IEC和IEC促炎信号的激活粘膜免疫细胞使用对照和患者衍生的3D肠癌的共培养评估和免疫细胞。这项研究的成功完成将填补理解小说的关键知识差距 UC中TNF和TLR依赖性炎症信号的基础机制，并为针对SAM68治疗UC的新型治疗的开发。此外，这项研究也将作为探索SAM68在其他TNF和TLR依赖性慢性炎症性疾病中的作用的基础。