Mediator kinases as interferon antagonists in Down Syndrome

介导激酶作为唐氏综合症干扰素拮抗剂

• 批准号: 10677733
• 负责人: Dylan J Taatjes
• 金额: $ 46.5万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-22 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10677733
• 关键词: Acute; Address; Affect; Age; Area; Autoimmune Diseases; Automobile Driving; B lymphocyte immortalization; Basic Science; Biochemical; Biological Assay; Cells; Chromosome 21; Chronic; Classification; Clinical; Code; Complement; Complex; DNA Binding; Defect; Dependence; Development; Down Syndrome; Ensure; Enzymes; Equilibrium; Foundations; Gene Expression; General Population; Genes; Genetic Transcription; Goals; Human; Human Genome; Hyperactivity; Hypersensitivity; Immune response; Immunity; In Vitro; Individual; Infection; Inflammation; Inflammatory; Interferon Activation; Interferon Receptor; Interferons; Link; Mediator; Metabolism; Methods; Molecular; Nuclear; Pathway interactions; Phosphotransferases; Polymerase; Proteins; RNA Polymerase II; RNA Splicing; Reagent; Regulation; Repression; Research Personnel; Role; Runaway; STAT1 gene; Sampling; Siblings; Signal Transduction; Solid; Spliced Genes; Stimulus; System; Testing; Therapeutic; Transcription Initiation; Transcriptional Regulation; United States National Institutes of Health; Untranslated RNA; antagonist; biobank; comorbidity; cortistatin; cytokine; druggable target; experience; experimental study; genome-wide; high reward; high risk; immune function; inhibitor; insight; interferon antagonist; metabolomics; new therapeutic target; novel therapeutic intervention; paralogous gene; programs; promoter; public health relevance; response; scaffold; sex; therapeutic target; transcription factor; transcriptome; transcriptome sequencing; transcriptomics

SUMMARY This collaborative project seeks to gain a deep mechanistic understanding of the Mediator kinases (CDK8 and CDK19) and their roles in driving hyper-active interferon (IFN) signaling in Down syndrome (DS). A chronic inflammatory state underlies the DS condition and contributes to its co-morbidities; consequently, a means to dampen IFN activation could have broad therapeutic benefits. CDK8 and CDK19 each represent druggable targets and, as potent activators of IFN responses, inhibition of their function could effectively antagonize runaway IFN signaling in DS. CDK8 and its paralog, CDK19, are considered “Mediator kinases” because of their stable, but reversible, association with the 26-subunit Mediator complex. Mediator is a global regulator of RNA polymerase II (pol II) activity, which transcribes all protein-coding and many non-coding RNAs in the human genome. CDK8 and CDK19 have each been shown to help drive transcriptional responses to IFN cytokines, which are universal regulators of inflammation and immune function. Here, we seek to define their essential— yet poorly understood—cellular and mechanistic roles in the context of DS (Biobank-matched D21 and T21 cells) and IFN signaling. We will combine sophisticated cell-based methods (e.g. transcriptomics and metabolomics) with detailed, cutting-edge biochemical approaches to allow rigorous and reliable assessment of the mechanisms by which CDK8 vs. CDK19 control IFN responses in DS individuals. In Aim 1, we will probe the role of Mediator kinase activity, whereas in Aim 2 we will focus on the CDK19 protein, which serves key structural/scaffolding roles to enable robust transcriptional responses upon IFN stimulation. Transcriptomic (RNA-seq and PRO-seq) and metabolomics approaches will be applied in Aims 1 and 2, to assess T21-specific signatures (including potential splicing defects) and to identify how CDK8 and/or CDK19 impact cellular responses to IFN. These experiments will also uncover how sequence-specific, DNA-binding transcription factors (TFs) are differentially mobilized in T21 vs. D21 cells, including during IFN stimulation. As part of this, we will establish how CDK8 and CDK19 independently control TF activation or repression, through kinase-dependent or -independent mechanisms. In Aim 3, we will perform detailed in vitro studies to define the molecular mechanisms by which Mediator kinases regulate pol II transcription initiation, promoter-proximal pausing, or elongation. Mechanistic findings will then be probed further in cells, to better characterize their significance in DS and related IFN signaling cascades. Collectively, these experiments (Aims 1-3) may help establish Mediator kinases (CDK8 & CDK19) as viable therapeutic targets to antagonize hyper-active IFN signaling in DS. Moreover, the balance of targeted/mechanistic studies with discovery-based approaches may yield new and unanticipated therapeutic strategies with relevance for DS and its associated co-morbidities, which also affect the general population.

概括 这个协作项目旨在获得对介体激酶的深刻理解（CDK8和 CDK19）及其在唐氏综合征（DS）中驱动超活动干扰素（IFN）信号传导中的作用。慢性 炎症状态是DS状况的基础，并导致其合并症；因此，一种手段 抑制IFN激活可能具有广泛的治疗益处。 CDK8和CDK19每个代表可吸毒 目标，作为IFN响应的潜在激活剂，其功能的抑制可能有效地拮抗 DS中的IFN信号失控。 CDK8及其旁系同源物CDK19被视为“介体激酶”，因为它们的 稳定但可逆的与26个亚基调解人建筑群相关。介体是RNA的全球调节器 聚合酶II（POL II）活性，它在人类中转录所有蛋白质编码和许多非编码RNA 基因组。 CDK8和CDK19已被证明有助于驱动对IFN细胞因子的转录反应， 是炎症和免疫功能的普遍调节剂。在这里，我们试图定义它们的基本 - 然而，了解不足 - 在DS（生物库匹配的D21和T21细胞）的情况下，细胞和机械作用 和IFN信号。我们将结合基于细胞的复杂方法（例如转录组学和代谢组学） 采用详细的尖端生化方法，允许对机制进行严格和可靠的评估 CDK8与CDK19控制DS个体中的IFN响应。在AIM 1中，我们将探究调解员的作用 激酶活性，而在AIM 2中，我们将专注于CDK19蛋白，该蛋白提供关键的结构/脚手架 在IFN刺激时可以发挥强大的转录反应。转录组（RNA-Seq和Pro-Seq） AIM 1和2将应用代谢组学方法，以评估T21特异性签名（包括 潜在的剪接缺陷）并确定CDK8和/或CDK19如何影响对IFN的细胞反应。这些 实验还将发现序列特异性，DNA结合转录因子（TF）如何差异 在T21与D21细胞中动员，包括在IFN模拟过程中。为此，我们将确定CDK8和 CDK19通过激酶依赖或非依赖性独立控制TF激活或表达 机制。在AIM 3中，我们将进行详细的体外研究，以定义分子机制 介体激酶调节Pol II转录启动，启动子抗暂停或伸长。机理 然后将进一步探测细胞的发现，以更好地表征其在DS和相关IFN中的重要性 信号级联。总的来说，这些实验（目标1-3）可能有助于建立介体激酶（CDK8＆ CDK19）作为可行的治疗靶标，可以拮抗DS中的高活动IFN信号。而且，平衡 具有基于发现的方法的有针对性/机械研究可能会产生新的和意外的疗法 与DS及其相关的合并症相关的策略，这也会影响普通人群。

项目成果

Transcription dosage compensation does not occur in Down syndrome.

• DOI: 10.1186/s12915-023-01700-4
• 发表时间: 2023-11-10
• 期刊: BMC BIOLOGY
• 影响因子: 5.4
• 作者: Hunter, Samuel;Hendrix, Jo;Freeman, Justin;Dowell, Robin D.;Allen, Mary A.
• 通讯作者: Allen, Mary A.

Metabolic and Transcriptomic Effects of Mediator Kinase Inhibition on the Interferon Response in Down Syndrome.

介导激酶抑制对唐氏综合症干扰素反应的代谢和转录组学影响。

• 发表时间: 2022
• 期刊: FASEB journal : official publication of the Federation of American Societies for Experimental Biology
• 作者: Cozzolino,KiraA;Jones,Taylor;Ajit,Deepa;Dowell,Robin;Taatjes,Dylan
• 通讯作者: Taatjes,Dylan