Role of the L1 retrotransposon in interferon-positive SLE

L1 逆转录转座子在干扰素阳性 SLE 中的作用

• 批准号: 10603189
• 负责人: Tomas M Mustelin
• 金额: $ 54.75万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-21 至 2028-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10603189
• 关键词: Adjuvant; Autoantibodies; Autoantigens; Autoimmunity; Automobile Driving; B-Lymphocytes; Biological Markers; Biology; Blocking Antibodies; Cell Aging; Cell Death; Cells; Childhood; Complement; Complex; Consumption; Crohn' s disease; DNA; Data; Dependence; Development; Disease; Elements; Event; FDA approved; Flare; Funding; Gender; Genetic Transcription; Goals; HIV; Human; IFNAR1 gene; IgG autoantibodies; Immune; Immune system; Interferon Type I; Interferon-beta; Interferons; Journals; L1 Elements; Lupus; Measures; Mediating; Methods; Molecular; Nephritis; Nuclear; Paper; Pathogenesis; Pathway interactions; Patients; Phase; Phase II Clinical Trials; Phase III Clinical Trials; Play; Process; Production; Protein Family; Proteins; RNA; RNA-Directed DNA Polymerase; Research; Retrotransposon; Reverse Transcription; Role; Serum; Signal Pathway; Signal Transduction; Stimulator of Interferon Genes; Systemic Lupus Erythematosus; Tenofovir; Testing; Transcript; Virus; Virus Diseases; Work; cancer cell; cell type; clinical development; density; ds-DNA; emtricitabine; experimental study; gene induction; granulocyte; immunogenic; inhibitor; monocyte; neutralizing monoclonal antibodies; neutrophil; protein complex; sensor; stress granule; targeted treatment; transcriptome sequencing; type I interferon receptor

Project Summary/Abstract The overall goal of our research is to elucidate the proximal molecular drivers and mechanisms of systemic lupus erythematosus (SLE) to enable the development of new and better therapies that target these processes but spare the broader immune system. This proposal will explore the role of the Long Interspersed Nuclear Element-1 (LINE-1, or L1) in type I interferon (IFN)-positive SLE. We have discovered that most SLE patients have high-titer autoantibodies against the first protein encoded by L1, ORF1p and that the titers of these autoantibodies correlate with disease activity as measured by SLEDAI score, complement consumption, presence of nephritis, anti- dsDNA and other autoantibodies, as well as with type I interferon gene induction. Our working hypothesis is that the biology of L1 is intimately connected with the pathogenesis of SLE by two mechanisms: i) driving autoimmunity to the virus-like L1 protein complexes that contain RNA and DNA; and ii) generating aberrant DNA by reverse-transcribing RNAs (including its own) and triggering DNA-sensor signaling to induce type I IFNs. AIM 1. To characterize the expression of L1 in SLE. The cell types that express the highest levels of L1 in SLE are the neutrophil and the low-density granulocyte. The L1-expressing neutrophils show signs of activation, and biomarkers of neutrophil NETosis are elevated in patient serum. By RNA-Seq we have identified one dominant (chr4q22.1) and six additional distinct human-specific L1 elements as the origins of the elevated transcripts and ORF1p protein. We will solidify these observations, detect and analyze their dependence on patient gender and disease parameters. We will examine factors that influence the transcription of these loci, and we will ask if stress granules containing ORF1p are released from neutrophils undergoing NETosis or other forms of cell death. These experiments will be complemented by asking if other proteins present in stress granules are also autoantigens in SLE. AIM 2. To elucidate the role of L1 ORF2p in type I IFN induction in SLE. Several recent papers in top journals have demonstrated a connection between L1 expression and the induction of IFN in cellular senescence, Crohn's disease, and in cancer cells. To determine how these findings apply to SLE, we will analyze L1-expressing neutrophils for activated DNA sensors, the signaling pathways from these sensors, and the type I IFN themselves. We will use an ultrasensitive method to quantitate ORF2p and then use inhibitors of the its RT activity to determine if it drives type I IFN production. Lastly, we will ask if silencing the DNA sensor DAI/ZBP1 blocks this IFN production.

项目总结/摘要 我们研究的总体目标是阐明近端分子驱动因素和机制 系统性红斑狼疮（SLE）的研究，以开发新的更好的治疗方法 针对这些过程，但不影响更广泛的免疫系统。本提案将探讨 长散布核元件-1（LINE-1，或L1）在I型干扰素（IFN）阳性 SLE。我们已经发现，大多数SLE患者都有高滴度的自身抗体， L1编码的蛋白质，ORF 1 p，这些自身抗体的滴度与疾病相关 通过SLEDAI评分、补体消耗、肾炎的存在、抗- dsDNA和其他自身抗体，以及与I型干扰素基因诱导。 我们的工作假设是，L1的生物学与以下疾病的发病机制密切相关： SLE通过两种机制：i）驱动对病毒样L1蛋白复合物的自身免疫， 含有RNA和DNA;和ii）通过逆转录RNA（包括RNA）产生异常DNA， 其自身的）和触发DNA传感器信号传导以诱导I型IFN。 AIM 1.探讨L1在SLE中的表达特点。表达最高的细胞类型 SLE患者的L1水平为中性粒细胞和低密度粒细胞。L1表达 中性粒细胞显示活化迹象，患者中性粒细胞NETosis的生物标志物升高 血清的通过RNA-Seq，我们已经确定了一个显性的（chr4q22.1）和六个额外的不同的 人类特异性L1元件作为升高的转录物和ORF 1 p蛋白的起源。我们将 巩固这些观察结果，检测并分析它们对患者性别和疾病的依赖性， 参数我们将研究影响这些基因座转录的因素， 如果含有ORF 1 p的应激颗粒从经历NETosis或其他的中性粒细胞释放， 细胞死亡的形式。这些实验将通过询问是否存在其他蛋白质来补充 也是SLE的自身抗原。 AIM 2.目的探讨L1 ORF 2 p在SLE患者I型IFN诱导中的作用。最近的几 顶级期刊上的论文已经证明了L1表达和归纳之间的联系 IFN γ在细胞衰老、克罗恩病和癌细胞中的作用。为了确定这些 研究结果适用于SLE，我们将分析L1表达的中性粒细胞的激活DNA传感器， 来自这些传感器的信号通路，以及I型IFN本身。我们将使用 超灵敏的方法来定量ORF 2 p，然后使用其RT活性的抑制剂， 确定它是否驱动I型干扰素的产生。最后，我们会问，如果沉默的DNA传感器， DAI/ZBP 1阻断这种IFN的产生。