Role and regulation of the splicing factor ASF/SF2 in human T lymphocyte physiolo

剪接因子ASF/SF2在人T淋巴细胞生理中的作用及调控

• 批准号: 8843361
• 负责人: Vaishali Moulton
• 金额: $ 12.41万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8843361
• 关键词: Activities of Daily Living; Address; Affect; Alternative Splicing; American; Arginine; Arthritis; Autoantibodies; Autoimmune Diseases; B-Lymphocytes; Biochemical; Biological Markers; Brain; CD3 Antigens; CD44 gene; CREM protein; Cause of Death; Cell Adhesion Molecules; Cell Line; Cell physiology; Cellular Immunology; Characteristics; Child; Chronic Disease; Complex; Defect; Disease; Disease Management; Disease Marker; Disease remission; Environment; Event; Exhibits; Family member; Functional disorder; Gene Expression Regulation; Gene Silencing; Generations; Genes; Goals; Half-Life; Human; IL2 gene; Immune; Immunotherapeutic agent; Individual; Infection; Infiltration; Israel; Kidney; Longitudinal Studies; Lymphocyte Function; Medical center; Messenger RNA; Methods; Molecular; Molecular Immunology; Organ; Pain; Pathogenesis; Pathology; Patients; Phase; Phosphorylation; Physiological; Physiology; Production; Protein Binding; Protein Isoforms; Protein Splicing; Proteins; Publishing; RNA Splicing; Regulation; Relapse; Research; Resources; Role; Serine; Signal Transduction; Signaling Molecule; System; Systemic Lupus Erythematosus; T-Cell Activation; T-Cell Development; T-Cell Receptor; T-Lymphocyte; Testing; Therapeutic immunosuppression; Transducers; Variant; Woman; base; body system; child bearing; cytokine; design; medical schools; migration; molecular marker; novel; peripheral blood; programs; research study; skills; skin disorder; targeted treatment; transcription factor

DESCRIPTION (provided by applicant): This K01 application is designed to prepare the applicant with the skills necessary to establish an independent research program on the study of systemic lupus erythematosus (SLE). SLE is an autoimmune disease of unknown cause which mainly afflicts women in their child bearing years and affects multiple organ systems of the body. T cell dysfunction due to abnormally altered intracellular signaling events is thought to be central in the pathogenesis of this disease. A critical signaling molecule known as the CD3 zeta chain is expressed in abnormally low amounts in the T cells of SLE patients, and this defect contributes to the poor functional capacity of the SLE T cells. Expression of an abnormal alternatively spliced (AS) form of the CD3 zeta mRNA with poor half life is a contributor to the reduced expression of zeta chain in SLE T cells. Aberrant splicing is a common feature observed in SLE and involves other critical genes involved in T cell signaling, migration, and cytokine production and the regulation of these genes has not been characterized. To understand the regulation of the CD3 zeta alternative splicing, the applicant used a discovery approach and identified several proteins binding to the CD3 zeta mRNA. Among these proteins, a promising candidate is the serine arginine (SR) family member Alternative Splicing Factor/Splicing Factor 2 (ASF/SF2). The applicant published that in T cells from healthy individuals, ASF/SF2 regulates CD3 zeta chain expression by suppressing production of the AS isoform. Examination of T cells from SLE patients revealed that patients had reduced expression of ASF/SF2 suggesting that aberrant ASF/SF2 expression may contribute to their defective T cell function. Additionally, the ASF/SF2 expression levels appeared to inversely correlate with the disease activity of SLE patients. The applicant's preliminary studies have shown a novel role for the splicing regulator ASF/SF2 in T cell physiology with a potential role in SLE pathophysiology. Based on the preliminary evidence, the hypothesis of this project is that ASF/SF2 regulates human T cell function and may represent a contributor to the SLE T cell defect. Toward the hypothesis, this project is aimed to understand the role of ASF/SF2 in human T lymphocytes and to determine whether ASF/SF2 may represent a disease marker in SLE. Methods will include biochemical analyses of human T cells and the development of a cell line to enable controlled expression of ASF/SF2. The specific aims of the project are 1) To determine whether TCR stimulation regulates ASF/SF2 expression and activity in human T lymphocytes 2) To determine how ASF/SF2 regulates T cell function and 3) To determine whether ASF/SF2 expression levels in SLE T cells may represent a disease marker. To accomplish these aims, the applicant will need to expand her technical and intellectual skills in cellular and molecular immunology and become proficient in the design, interpretation and analysis of studies in SLE patients. The research environment at Beth Israel Deaconess Medical Center and the other affiliates of the Harvard Medical School system will provide the applicant with the resources to reach her goals within 5 years.

描述（由申请人提供）：本 K01 申请旨在帮助申请人具备建立系统性红斑狼疮 (SLE) 独立研究项目所需的技能。系统性红斑狼疮是一种原因不明的自身免疫性疾病，主要影响育龄妇女，影响身体多个器官系统。由于细胞内信号传导事件异常改变而导致的 T 细胞功能障碍被认为是该疾病发病机制的核心。一种称为 CD3 zeta 链的关键信号分子在 SLE 患者的 T 细胞中表达量异常低，这种缺陷导致 SLE T 细胞的功能能力较差。半衰期较短的 CD3 zeta mRNA 的异常选择性剪接 (AS) 形式的表达是导致 SLE T 细胞中 zeta 链表达减少的一个因素。异常剪接是在 SLE 中观察到的一个常见特征，并涉及参与 T 细胞信号传导、迁移和细胞因子产生的其他关键基因，但这些基因的调节尚未得到表征。为了了解 CD3 zeta 选择性剪接的调节，申请人使用了一种发现方法并鉴定了几种与 CD3 zeta mRNA 结合的蛋白质。在这些蛋白质中，一个有前途的候选者是丝氨酸精氨酸 (SR) 家族成员选择性剪接因子/剪接因子 2 (ASF/SF2)。申请人发表文章称，在健康个体的 T 细胞中，ASF/SF2 通过抑制 AS 亚型的产生来调节 CD3 zeta 链的表达。对 SLE 患者 T 细胞的检查显示，患者 ASF/SF2 表达减少，表明 ASF/SF2 表达异常可能导致其 T 细胞功能缺陷。此外，ASF/SF2 表达水平似乎与 SLE 患者的疾病活动度呈负相关。申请人的初步研究表明剪接调节因子 ASF/SF2 在 T 细胞生理学中具有新的作用，并具有潜在的作用 SLE 病理生理学。根据初步证据，该项目的假设是 ASF/SF2 调节人类 T 细胞功能，并可能是 SLE T 细胞缺陷的一个促成因素。针对这一假设，该项目旨在了解 ASF/SF2 在人类 T 淋巴细胞中的作用，并确定 ASF/SF2 是否可能代表 SLE 的疾病标志物。方法将包括人类 T 细胞的生化分析和细胞系的开发，以实现 ASF/SF2 的受控表达。该项目的具体目标是 1) 确定 TCR 刺激是否调节人 T 淋巴细胞中的 ASF/SF2 表达和活性 2) 确定 ASF/SF2 如何调节 T 细胞功能 3) 确定 SLE T 细胞中的 ASF/SF2 表达水平是否可以代表疾病标志物。为了实现这些目标，申请人需要扩展其在细胞和分子免疫学方面的技术和智力技能，并精通系统性红斑狼疮患者研究的设计、解释和分析。贝斯以色列女执事医疗中心和哈佛医学院系统其他附属机构的研究环境将为申请人提供在 5 年内实现其目标的资源。