TRIM27 is a new negative regulator of CD4 T cells and Mast cells.

TRIM27 是 CD4 T 细胞和肥大细胞的新型负调节因子。

基本信息

批准号：
8875012
负责人：
EDWARD Y SKOLNIK
金额：
$ 32.96万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-01-15 至 2017-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8875012
关键词：
1-Phosphatidylinositol 3-Kinase Affect Anaphylaxis Applications Grants Asthma Autoimmune Diseases B-Cell Activation Boxing CD4 Positive T Lymphocytes Calcium-Activated Potassium Channel Cells Coiled-Coil Domain Cutaneous Development Failure Feedback Fingers Generations Goals Graft Rejection Helper-Inducer T-Lymphocyte Histidine Human Hypersensitivity Immune system In Vitro Inflammation Lead Lymphocyte Lysine Mediating Mus Names Nucleoside diphosphate kinase B Pathway interactions Phosphoric Monoester Hydrolases Phosphotransferases Physiological Potassium Channel Protein Family Proteins Receptor Activation Recruitment Activity Regulation Regulatory T-Lymphocyte Role Signal Pathway Signaling Molecule Structure of thyroid parafollicular cell T-Cell Activation T-Cell Receptor T-Lymphocyte T-Lymphocyte Subsets TRIM Motif Th1 Cells Ubiquitination allergic response follow-up immunological synapse immunological synapse formation in vivo inhibitor/antagonist insight mast cell member myotubularin phosphatidylinositol 3-phosphate phosphohistidine prevent protein-histidine kinase response ubiquitin-protein ligase

项目摘要

DESCRIPTION (provided by applicant): The K+ channel KCa3.1 is required for Ca2+ influx and the subsequent activation of B, T, and mast cells. Our studies have found that T cell receptor (TCR) activation leads to activation of KCa3.1 by activating the class 2 phosphatidylinositol 3 kinase C2 Beta (PI3K-C2ß) leading to the generation of PI3P, which is required for the subsequent activation of a mammalian histidine kinase, Nucleoside Diphosphate Kinase B, that then activates KCa3.1. We identified the Tripartate Motif containing protein 27 (TRIM27) as a new negative regulator of PI3K-C2ß and KCa3.1. TRIM27 is a member of a large family of proteins characterized by the presence of a tripartite motif, consisting of a RING finger, B box, and coiled coil (CC) domains. In this proposal, we will determine the mechanisms whereby TRIM27 regulates PI3K-C2ß, its physiological significance, and subsequent role as a negative regulator of KCa3.1 in lymphocyte and mast cell activation in vitro and in vivo. We have evidence that TRIM27 functions an E3 ligase and ubiquitinates and inhibits PI3K-C2ß. To determine the mechanism(s) whereby TRIM27 regulates PI3K-C2ß and how this is affected by TCR activation, we will determine in SA1, (Ai) the type of TRIM27 mediated ubiquitination of PI3K-C2ß; (Aii) the lysine residues on TRIM27 and PI3K-C2ß that are ubiquitinated; and (Aiii) the regions or domains on TRIM27 and PI3K-C2ß that mediate their association. In 1B, we will determine: (Bi) if TRIM27 regulates TCR stimulated PI3K-C2ß's kinase activity and/or degradation, or whether (Bii) association of TRIM27 with PI3K-C2ß is affected by TCR stimulation; (Biii) whether TRIM27 is recruited to the immunological synapse (IS), its role in IS formation and/or the recruitment PI3K-C2ß to IS following TCR activation; (Biv) whether TRIM27 modulates the PI3P levels in activated T cells, or (Bv) whether TRIM27 autoubiquitination, or ubiquitination of PI3K-C2ß is modulated by TCR stimulation. In (C), the role of TRIM27 stimulated SUMOylation of PI3K-C2ß will be assessed. We have generated TRIM27-/- mice and have evidence that KCa3.1 channel activity and TCR- stimulated Ca2+ flux are increased in TRIM27-/- Th1 cells. In SA2 (A), we will extend these studies to other CD4 helper T cell subsets, and determine whether Th2, Th17, and Treg differentiation and/or function are altered in cells isolated from TRIM27-/- mice. In (B), we will determine the downstream signaling pathways regulated by TRIM27, and (C) whether TRIM27-/- mice are predisposed to autoimmune disease. In (D) we will undertake a nonbiased approach to identify other targets of TRIM27 ubiquitination and regulation We found that TRIM27 functions to negatively regulate FceR1 stimulated KCa3.1 channel activity and Ca2+ flux in mast cells. We will determine in SA3: (A) if PI3K-C2ß mediates FceR1 stimulated activation of KCa3.1 and whether this is modulated by TRIM27; (B) whether effectors functions of TRIM27-/- mast cells are increased; (3C) the signaling pathways regulated by TRIM27 in mast cells, and/or (3D) whether TRIM27-/- mice have an increased anaphylactic response to both passive cutaneous and systemic anaphylaxis.

描述（由申请人提供）：K+ 通道 KCa3.1 是 Ca2+ 流入以及随后 B、T 和肥大细胞激活所必需的。我们的研究发现，T 细胞受体 (TCR) 激活会导致 KCa3.1 激活。通过激活 2 类磷脂酰肌醇 3 激酶 C2 Beta (PI3K-C2ß)，导致生成 PI3P，这是后续激活所必需的哺乳动物组氨酸激酶，核苷二磷酸激酶 B，然后激活 KCa3.1，我们鉴定出含有蛋白 27 (TRIM27) 的三部分基序为 PI3K-C2ß 的新负调节因子，并且 KCa3.1 是一个大 TRIM27 的成员。以存在三部分基序为特征的蛋白质家族，由环指、B 盒和卷曲螺旋 (CC) 结构域组成。我们将确定 TRIM27 调节 PI3K-C2ß 的机制、其生理意义以及随后在体外和体内淋巴细胞和肥大细胞激活中作为 KCa3.1 负调节因子的作用。我们有证据表明 TRIM27 具有 E3 连接酶和泛素化作用。并抑制 PI3K-C2ß 确定 TRIM27 调节 PI3K-C2ß 的机制以及 TCR 如何影响这一机制。激活，我们将在 SA1 中确定 TRIM27 介导的 PI3K-C2ß 泛素化类型；(Aii) TRIM27 和 PI3K-C2ß 上泛素化的赖氨酸残基；以及 (Aiii) TRIM27 和 PI3K 上的区域或结构域； -C2ß 介导它们的关联在 1B 中，我们将确定： (Bi) 如果。 TRIM27 调节 TCR 刺激的 PI3K-C2ß 的激酶活性和/或降解，或者 (Bii) TRIM27 与 PI3K-C2ß 的关联是否受到 TCR 刺激的影响；(Biii) TRIM27 是否被招募到免疫突触 (IS)，其在TCR 激活后 IS 的形成和/或 PI3K-C2ß 向 IS 的募集 (Biv) TRIM27 是否调节激活的 T 细胞中的 PI3P 水平，或 (Bv) TRIM27 自身泛素化或 PI3K-C2ß 泛素化是否受 TCR 刺激调节。在 (C) 中，我们将评估 TRIM27 刺激 PI3K-C2ß 的 SUMO 化的作用。 -/- 小鼠，并有证据表明 KCa3.1 通道活性和 TCR 刺激的 Ca2+ 通量在TRIM27-/- Th1 细胞。在 SA2 (A) 中，我们将这些研究扩展到其他 CD4 辅助 T 细胞亚群，并确定从 TRIM27-/- 分离的细胞中 Th2、Th17 和 Treg 分化和/或功能是否发生改变。在 (B) 中，我们将确定 TRIM27 调节的下游信号通路，以及 (C) TRIM27-/- 小鼠是否易患自身免疫性疾病。确定 TRIM27 泛素化和调节的其他靶标的无偏方法我们发现 TRIM27 具有负调节肥大细胞中 FceR1 刺激的 KCa3.1 通道活性和 Ca2+ 通量的功能。我们将在 SA3 中确定：(A) PI3K-C2ß 是否介导 FceR1 刺激的激活。 KCa3.1 的影响以及其是否受 TRIM27 调节；(B) TRIM27-/- 肥大细胞的效应子功能是否起作用；增加；(3C) 肥大细胞中 TRIM27 调节的信号通路，和/或 (3D) TRIM27-/- 小鼠对被动皮肤和全身过敏反应是否有增加的过敏反应。