Probing the Functions of KLF2 Downregulation

探讨 KLF2 下调的功能

• 批准号: 9247134
• 负责人: Geoffrey S. Kansas
• 金额: $ 19.31万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9247134
• 关键词: Address; Adipocytes; Anti-Inflammatory Agents; Anti-inflammatory; Arginine; B-Lymphocytes; Biochemical; Biological Process; Blood Vessels; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CD8B1 gene; Cell Compartmentation; Cell Count; Cells; Communities; Data; Defect; Development; Disease; Down-Regulation; Endothelial Cells; Endothelium; Engineering; Enterobacteria phage P1 Cre recombinase; Erythrocytes; Erythropoiesis; Family; Generations; Genes; Genetic Transcription; Helper-Inducer T-Lymphocyte; Hour; Immune system; Inflammatory; Leukocytes; Lung; Lymphocyte; Lysine; Maintenance; Mature T-Lymphocyte; Mediating; Memory; Messenger RNA; Mouse Strains; Mus; Mutant Strains Mice; Mutation; Myeloid Cells; Process; Proteins; Regulation; Resources; Stimulus; System; T cell regulation; T-Lymphocyte; Tissues; Ubiquitin; Ubiquitination; adipocyte differentiation; cell motility; cell type; lung development; member; multicatalytic endopeptidase complex; mutant; novel; public health relevance; receptor; response; success; thymocyte; transcription factor; ubiquitin-protein ligase

 DESCRIPTION (provided by applicant): KLF2 is a member of the Kruppel-like family of transcription factors, which control diverse processes in many different cell types. KLF2 has documented functions in a wide variety of cells and tissues, including lung development, vascular mural cell migration, adipocyte development, and erythropoiesis. Within the immune system, KLF2 also subserves multiple functions, including regulation of T and B cell traffic, maintenance of myeloid cell quiescence, and anti-inflammatory actions in endothelial cells. More recently, KLF2 has been implicated in regulation of T helper differentiation, but mechanisms underlying this function are not yet clear. KLF2 is somewhat unique among lymphocyte transcription factors in that its expression is rapidly lost upon activation through Ag receptors (TCR and BCR), and this loss of expression is at both the mRNA and protein levels. A similar process occurs in both myeloid cells and in endothelium, where conditional inactivation of KLF2 leads to an activated, pro-inflammatory state. Whether downregulation of KLF2 is an essential aspect of the development or function of other cell types, including lung, blood vessels, adipocytes, and early erythrocytes, is unknown. Downregulation of KLF2 following Ag receptor engagement occurs rapidly at both the transcriptional and post-translational levels, generally within ~8-12 hours. Mechanisms which mediate transcriptional silencing have not been identified. In contrast, it is well established that KLF2 downregulation at the protein level s via the well studied ubiquitin-proteosome system, and several E3 ubiquitin ligases which mediate this effect in various non-hematopoietic cells have been identified. Further, the key lysine residue essential for ubiquitination has been identified, and mutation of this lysine to arginine completely blocks proteolytic destruction of KLF2. However, none of these studies have addressed the basic question of why this occurs, and no biological functions in any cell types have been identified which require KLF2 downregulation. We will address this issue by producting a novel mouse strain in which KLF2 expression can be enforced in any cell type which expresses the cre recombinase (Aim 1), and use this mouse to understand the functions of KLF2 downregulation in the T cell compartment (Aim 2). Further, the mutant mice which we produce will be ideal for approaching this question in numerous other cell types as well. As KLF2 downregulation is a feature of many, if not most or all cell types in which it is expressed, this project will make a major impact in the field.

 描述（由申请人提供）：KLF2是Kruppel样转录因子家族的成员，其控制许多不同细胞类型中的不同过程。 KLF2 已记录在多种细胞和组织中的功能，包括肺发育、血管壁细胞迁移、脂肪细胞发育和红细胞生成。在免疫系统中，KLF2 还具有多种功能，包括调节 T 和 B 细胞运输、维持骨髓细胞静止以及内皮细胞的抗炎作用。最近，KLF2 与 T 辅助细胞分化的调节有关，但该功能的机制尚不清楚。 KLF2 在淋巴细胞转录因子中有些独特，因为它的表达在通过 Ag 受体（TCR 和 BCR）激活后迅速丧失，并且这种表达丧失发生在 mRNA 和蛋白质水平。类似的过程发生在骨髓细胞和内皮细胞中，其中 KLF2 的条件性失活会导致激活的促炎状态。 KLF2 的下调是否是其他细胞类型（包括肺、血管、脂肪细胞和早期红细胞）发育或功能的重要方面尚不清楚。 Ag 受体结合后 KLF2 的下调在转录和翻译后水平上迅速发生，通常在约 8-12 小时内。介导转录沉默的机制尚未确定。相比之下，众所周知，KLF2 通过经过充分研究的泛素-蛋白酶体系统在蛋白质水平上下调，并且已鉴定出几种在各种非造血细胞中介导这种作用的 E3 泛素连接酶。此外，已经确定了泛素化所必需的关键赖氨酸残基，并且该赖氨酸突变为精氨酸完全阻止了 KLF2 的蛋白水解破坏。然而，这些研究都没有解决为什么会发生这种情况的基本问题，也没有发现任何细胞类型中需要下调 KLF2 的生物学功能。 我们将通过生产一种新的小鼠品系来解决这个问题，其中 KLF2 表达可以在表达 cre 重组酶的任何细胞类型中强制执行（目标 1），并使用该小鼠来了解 T 细胞区室中 KLF2 下调的功能（目标 2）。此外，我们生产的突变小鼠也将是在许多其他细胞类型中解决这个问题的理想选择。由于 KLF2 下调是许多（如果不是大多数或所有）表达 KLF2 的细胞类型的一个特征，因此该项目将在该领域产生重大影响。