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Role for nuclear matrix proteins and DNA methylation for XCI maintenance in female lymphocytes

核基质蛋白和 DNA 甲基化在女性淋巴细胞 XCI 维持中的作用

基本信息

批准号：
10660313
负责人：
Montserrat C Anguera
金额：
$ 57.38万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-07 至 2028-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10660313
关键词：
Alleles Antibody Formation Autoantibodies Autoantigens Autoimmune Autoimmune Diseases Autoimmunity B cell differentiation B-Cell Activation B-Lymphocyte Subsets B-Lymphocytes Binding Binding Proteins Biological Assay Biotinylation Cell physiology ChIP-seq Chromatin Chronic Complex DNA DNA Methylation Data Disease Dosage Compensation (Genetics)Epigenetic Process Exhibits Female Functional disorder Gene Dosage Gene Expression Gene Modified Gene Silencing Genes Genetic Genetic Transcription Goals Hypermethylation Immune Immunity Impairment In Vitro Inflammation Interferon Type I Interferons Link Lupus Lupus erythematosus cell Lymphocyte Maintenance Mediating Methylation Modification Molecular Mus Nuclear Lamina Nuclear Matrix-Associated Proteins Oligonucleotides Pathway interactions Patients Predisposing Factor Pristane Process Production Protein Methylation Proteins Publishing RNA RNA Binding Regulator Genes Role SAFB gene Severity of illness Signal Transduction Somatic Cell Susceptibility Gene Systemic Lupus Erythematosus Testing Therapeutic Intervention Transcript Work X Chromosome X Inactivation autosome biological sex bisulfite sequencing cellular imaging cohesin cohesion ds-DNA gene repression in vivo innovation insight lupus-like male mouse model novel overexpression preservation promoter response whole genome

项目摘要

Females are predisposed for developing systemic lupus erythematosus (SLE), but the underlying mechanisms remain obscure. Females have two X-chromosomes and equilibration of X-linked gene dosage to that of males (XY) occurs by X-Chromosome Inactivation (XCI), initiated and maintained by Xist RNA. The X-chromosome is enriched for immunity-related genes and X-linked genes are typically overexpressed in female-biased SLE, suggesting that impaired XCI maintenance contributes to SLE. Unlike other somatic cells, we found that naïve B cells lack enrichment of Xist RNA and heterochromatic marks, and that these modifications return to the inactive X (Xi) upon in vitro stimulation, which we term `dynamic XCI maintenance'. Our published work demonstrates that SLE patient naïve B cells exhibit abnormal expression of X-linked genes, suggesting a dysfunctional Xist- independent mechanism involving DNA methylation for transcriptional silencing prior to autoantigen stimulation could predispose to lupus. DNA methylation, associated with transcriptional repression, is reduced in SLE patient B cells and is an important epigenetic modification for gene silencing on the Xi. XCI is also regulated by various nuclear matrix proteins, such as hnRNPU and SAFB, which bind to Xist RNA and function to anchor Xist to the Xi or the nuclear lamina in somatic cells. Our preliminary data support the novel concept that Xist RNA transcripts are retained at the nuclear lamina by SAFB in naïve B cells, and that Xist RNA binds to hnRNPU upon B cell activation, tethering Xist RNA to the Xi to regulate transcriptional repression. We have developed a novel Xist deletion mouse model to determine how impaired XCI maintenance contributes to lupus disease, and our exciting preliminary data show that increased dsDNA levels in type I interferon induced disease. Our central hypothesis is that Xist-independent DNA methylation maintains transcriptional repression of the Xi and autosomes in naïve B cells and that upon activation, alterations in nuclear matrix protein binding promotes dynamic XCI maintenance, the dysfunction of which exacerbates Type I IFN-driven lupus disease. We will test our hypotheses with the following aims: (1) Are reductions in Xist-independent DNA hypermethylation in naïve B cells associated with aberrant gene expression in lupus? (2) How do interactions between SAFB, hnRNPU protein and Xist RNA maintain XCI in B cells and does lupus disease impair cohesin eviction by hnRNPU-Xist RNA complexes? (3) How does impaired XCI maintenance impact Type I IFN-driven lupus-like disease in female mice? IMPACT: Our novel and innovative genetic and molecular approaches will yield unprecedented mechanistic insight into how biological sex contributes to immune dysregulation of SLE disease, and will enable the identification of new molecular pathways and targets of female-biased autoimmune disease that could be amenable for therapeutic intervention.

女性易患系统性红斑狼疮 (SLE)，但其潜在机制保持模糊。女性有两条 X 染色体，X 连锁基因剂量与男性平衡 (XY) 通过 X 染色体失活 (XCI) 发生，由 Xist RNA 启动和维持。 X染色体是富含免疫相关基因，而 X 连锁基因通常在女性偏向的 SLE 中过度表达，表明 XCI 维持受损会导致 SLE。与其他体细胞不同，我们发现幼稚 B 细胞缺乏 Xist RNA 和异染色质标记的富集，并且这些修饰返回到非活性状态体外刺激后的 X (Xi)，我们称之为“动态 XCI 维持”。我们发表的作品表明 SLE 患者的初始 B 细胞表现出 X 连锁基因的异常表达，表明 Xist 功能失调涉及自身抗原刺激前转录沉默的 DNA 甲基化的独立机制可能易患狼疮。 SLE 患者中与转录抑制相关的 DNA 甲基化水平降低 B 细胞上的 Xi 基因沉默是重要的表观遗传修饰。 XCI 还受到各种监管核基质蛋白，例如 hnRNPU 和 SAFB，它们与 Xist RNA 结合并发挥将 Xist 锚定到 Xi 或体细胞中的核层。我们的初步数据支持 Xist RNA 转录本的新概念在幼稚 B 细胞中，SAFB 保留在核纤层上，并且 Xist RNA 与 B 细胞上的 hnRNPU 结合激活，将 Xist RNA 与 Xi 结合以调节转录抑制。我们开发了一种新颖的 Xist 缺失小鼠模型来确定 XCI 维持受损如何导致狼疮疾病，我们令人兴奋的初步数据表明，I 型干扰素诱发的疾病中双链 DNA 水平增加。我们的中心假设是Xist独立的DNA甲基化维持了Xi和常染色体的转录抑制 B 细胞以及激活后核基质蛋白结合的改变促进动态 XCI 维持，其功能障碍会加剧 I 型 IFN 驱动的狼疮疾病。我们将测试我们的假设其目标如下：(1) 幼稚 B 细胞中 Xist 独立 DNA 高甲基化的减少是否与狼疮基因表达异常？ (2) SAFB、hnRNPU蛋白和Xist RNA之间如何相互作用维持 B 细胞中的 XCI，狼疮疾病是否会损害 hnRNPU-Xist RNA 复合物对粘连蛋白的驱逐？ (3) XCI 维持受损如何影响雌性小鼠 I 型 IFN 驱动的狼疮样疾病？影响：我们的新颖和创新的遗传和分子方法将产生前所未有的机制洞察力生物性别导致系统性红斑狼疮疾病的免疫失调，并将有助于识别新的女性偏向性自身免疫性疾病的分子途径和靶点可能适合治疗干涉。