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

描述（由申请人提供）：本K01申请旨在为申请人提供建立系统性红斑狼疮（SLE）独立研究项目所需的技能。SLE是一种病因不明的自身免疫性疾病，主要发生在育龄妇女身上，影响身体的多个器官系统。细胞内信号事件异常改变导致的T细胞功能障碍被认为是本病发病机制的核心。一种被称为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，这一缺陷导致SLE T细胞功能低下。半衰期较差的CD3 zeta mRNA的异常选择性剪接（AS）形式的表达是SLE T细胞中zeta链表达减少的一个因素。异常剪接是SLE的一个共同特征，它涉及到其他参与T细胞信号传导、迁移和细胞因子产生的关键基因，而这些基因的调控尚未被描述。为了了解CD3 zeta选择性剪接的调控，申请人使用发现方法并确定了几种与CD3 zeta mRNA结合的蛋白质。在这些蛋白中，一个有希望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）。申请人发表了在健康个体的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患者的T细胞检测显示，患者ASF/SF2表达降低，提示异常的ASF/SF2表达可能导致其T细胞功能缺陷。此外，ASF/SF2表达水平似乎与SLE患者的疾病活动性呈负相关。申请人的初步研究表明，剪接调节剂ASF/SF2在T细胞生理学中具有新的作用，可能在以下方面发挥作用

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.