Role of NFAT in the Skeleton

NFAT 在骨骼中的作用

• 批准号: 9146275
• 负责人: Ernesto Canalis
• 金额: $ 39.63万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-18 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9146275
• 关键词: Accounting; Binding Proteins; Biomechanics; Bone Resorption; Bone remodeling; Cell model; Cells; Collagen; Collagen Type I; Disease; Elements; Event; Exhibits; Failure; Gene Expression; Gene Expression Regulation; Goals; Half-Life; Health; Homeostasis; Hour; Immune response; In Vitro; Inflammatory Response; Investigation; Laboratories; Messenger RNA; MicroRNAs; Modeling; Mus; Mutant Strains Mice; Mutation; Nucleic Acid Regulatory Sequences; Osteoblasts; Osteogenesis; Osteopenia; Phenotype; Physiology; Play; Positioning Attribute; Post-Transcriptional Regulation; Property; Proteins; RNA Sequences; RNA-Binding Proteins; Regulation; Research; Role; Signal Transduction; Skeleton; T-Cell Activation; Transcript; Transgenic Organisms; Uncertainty; Up-Regulation; Work; bone; bone mass; cytokine; gain of function; in vivo; inhibitor/antagonist; mouse model; mutant; notch protein; novel; nuclear factors of activated T-cells; osteoblast differentiation; osteoclastogenesis; promoter; response; skeletal; tomography; transcription factor

 DESCRIPTION (provided by applicant): Investigations from our laboratory revealed that skeletal cells synthesize Nuclear factor of activated T- cells (NFAT)c1 through c4. Characterization of cells expressing constitutively active (ca) Nfatc1 or Nfatc2 indicates an inhibitory role of Nfat in osteoblastic differentiation and function. This was confirmed in vivo, ad transgenics expressing caNfatc2 in osteoblasts under the control of the type I α1 collagen promoter (Col3.6- Nfatc2) are osteopenic and have impaired bone formation. Studies in mouse models of global Nfatc2 inactivation are difficult to interpret because they have non-specific inflammatory responses and upregulation of various cytokines. To understand the function of Nfatc2 in osteoblasts, we obtained conditional Nfatc1loxP/loxP and created conditional Nfatc2loxP/loxP mice placing us in a unique position to establish its function, singly and in the context of the Nfatc1 inactivation. In accordance with the osteopenic phenotype of the Col3.6-Nfatc2 transgenics, inactivation of Nfatc1 or Nfatc2 in Osterix expressing cells resulted in an increase in cancellous bone volume. In independent studies, we discovered that Notch induces Nfatc2 expression in osteoblasts selectively and acts by post-transcriptional mechanisms. In a Notch2 gain-of-function model, Notch2Q2319X mutant mice exhibited osteopenia due to increased bone resorption and lack of a compensatory bone forming response. In this model, Nfatc2 is induced in osteoblasts and may account for the uncoupling of the two remodeling events. The goal of this project is to understand the function of Nfatc2 in cells of the osteoblastc lineage. For this purpose, we will study mouse and cellular models misexpressing Nfatc2 in cells of the osteoblastic lineage. Our specific aims are: 1) To establish the function of Nfatc2 in osteoblasts. To this end, the skeletal phenotype of conditional deletion mutants of Nfatc2, singly and in combination with Nfatc1, will be established in cells of the osteoblastic lineage using microcomputed tomography and bone histomorphometric analysis. In addition, we will examine the biomechanical properties of bone and study mechanisms responsible for the effects; 2) To establish the post-transcriptional control of Nfatc2 by Notch in osteoblasts. This Aim will define the control of Nfatc2 expression by Notch and identify RNA sequences, binding proteins and microRNAs responsible for Nfatc2 post-transcriptional regulation; and 3) To establish the contribution of Nfatc2 to Notch actions in osteoblasts and determine whether the Nfatc2 induction by Notch is responsible for the failure of Notch2Q2319X mutants to manifest a bone forming response to enhance bone resorption. To this end, the Notch2Q2319X phenotype will be examined in the context of the Nfatc2 inactivation. These investigations should clarify the function of Nfatc2 in cells of the osteoblastic lineage, and identify novel mechanisms by which the skeleton integrates distinct signals to regulate bone remodeling.

 描述（由申请人提供）：我们实验室的研究显示骨骼细胞合成活化T细胞核因子（NFAT）c1至c4。表达组成性活性（ca）Nfatc 1或Nfatc 2的细胞的表征表明Nfat在成骨细胞分化和功能中的抑制作用。这在体内得到证实，在I型α1胶原启动子（Col3.6-Nfatc 2）控制下在成骨细胞中表达caNfatc 2的ad转基因是骨质减少的并且具有受损的骨形成。在小鼠模型中的整体Nfatc 2失活的研究很难解释，因为它们具有非特异性炎症反应和各种细胞因子的上调。为了了解Nfatc 2在成骨细胞中的功能，我们获得了条件性Nfatc 1 loxP/loxP，并建立了条件性Nfatc 2loxP/loxP小鼠，使我们处于一个独特的位置，以单独和在Nfatc 1失活的背景下建立其功能。根据Col3.6-Nfatc 2转基因的骨质减少表型，Osterix表达细胞中Nfatc 1或Nfatc 2的失活导致松质骨体积增加。在独立的研究中，我们发现Notch选择性地诱导成骨细胞中Nfatc 2的表达，并通过转录后机制起作用。在Notch 2功能获得性模型中，Notch 2 Q2319 X突变小鼠由于骨吸收增加和缺乏代偿性骨形成反应而表现出骨质减少。在该模型中，Nfatc 2在成骨细胞中被诱导，并且可以解释两个重构事件的解偶联。该项目的目标是了解Nfatc 2在成骨细胞谱系中的功能。为此，我们将研究成骨细胞系细胞中错误表达Nfatc 2的小鼠和细胞模型。本研究的具体目的是：1）研究Nfatc 2在成骨细胞中的功能。为此，Nfatc 2的条件性缺失突变体的骨骼表型，单独和与Nfatc 1组合，将建立在成骨细胞谱系的细胞中使用微计算机断层扫描和骨组织形态学分析。此外，我们还将研究骨的生物力学特性，并研究其作用机制; 2）建立Notch对成骨细胞中Nfatc 2的转录后调控。该目的将定义Notch对Nfatc 2表达的控制，并鉴定负责Nfatc 2转录后调节的RNA序列、结合蛋白和microRNA;以及3）确定Nfatc 2对成骨细胞中Notch作用的贡献，并确定Notch对Nfatc 2的诱导是否是Notch 2 Q2319 X突变体未能表现出骨形成应答以增强骨吸收的原因。为此，将在Nfatc 2失活的情况下检查Notch 2 Q2319 X表型。这些研究应该阐明Nfatc 2在成骨细胞谱系细胞中的功能，并确定骨骼整合不同信号以调节骨重建的新机制。