Epigenetics of the autoantibody response in systemic lupus

项目摘要/摘要。系统性红斑狼疮自身抗体反应的表观遗传学。 就像对病毒和细菌的“成熟”抗体反应一样，狼疮自身抗体反应需要B细胞型- 开关DNA重组(CSR)、体细胞超突变(SHM)和浆细胞分化。正如我们所做的那样 显示，表观遗传因素，包括组蛋白修饰物(如Sirt1)和抑制剂(如丁酸盐)调节 B细胞表达AID(基因：AICDA/Aicda)和Blimp-1(PRDM1/Prdm1)，这是CSR/SHM的关键 和浆细胞分化。对B型狼疮表观遗传格局的更好定义 细胞，我们假设TET2，一个关键的表观遗传因子，在提示下介导狼疮自身抗体反应 TET2在CSR/SHM和浆细胞分化中的B细胞内在作用(我们最近的发现)。我们认为 TET2是由诱导B细胞CSR/SHM和浆细胞分化的刺激诱导的，并且上调 在狼疮B细胞中。我们还认为，TET2通过其Fe2和a-1促进Aicda和Prdm1的转录。 酮戊二酸(a-KG)依赖的DNA脱甲基化催化活性及其非催化功能，即， 将OGT(由X-连锁OGT/OGT编码)招募到这些基因座上，以影响组蛋白糖基化。最后，我们认为 B细胞TET2将激素、营养和代谢信号转化为表观遗传变化，以调节 自身抗体反应，如TET2转录可被雌激素(我们的初步数据)和 维生素A和TET2被维生素C激活，但被富马酸(一种代谢物和α-KG竞争对手)抑制-AS 我们发现，雌激素促进了艾滋病的表达，从而增强了女性的抗体和自身抗体反应。 在人类和小鼠狼疮的机械性理解方面有丰富的经验和承诺 自身抗体反应，我们独一无二地准备使用分子B细胞生物学系统来测试我们的假设， 尖端表观遗传工具(羟甲基DNA分析、亚硫酸氢盐和氧化亚硫酸氢盐转化、芯片 和ATAC-Seq)，转基因小鼠(TgAicda-creTet2fl/fl，Tet2HxD-mut/hxd-mut和Tet2ogt-mut/Ogt-mut敲门小鼠， TgAicda-creOgtfl/fl、Tet2HxD-mut/hxd-muTgAicda-creOgtfl/fl、MRL/LPR TgAicda-creTet2fl/fl、MRL/LPR Tet2HxD-mut/hxd-mut和MRL/LPR TgAicda-creOgtfl/fl)，专有的人性化H-MICE®和Lupus-H-MICES®型号。目标1针对人类和 小鼠B细胞分化阶段特异性(静息、激活、浆和记忆细胞)对TET2、TET2的调节 蛋白质稳定性、E2上调TET2转录本及其潜在机制。AIM 2针对B细胞- TET2在促进AID和Blimp-1表达中的内在作用及其潜在机制(活性DNA 去甲基化和OGT介导组蛋白糖基化)，以及TET2诱导剂维生素A和TET2的增强作用 激活剂维生素C和TET2催化抑制剂(富马酸盐、Tet-IN-C35或Bobcat339)单独的抑制作用 或联合OGT消融或Tet2Ogt-MUT/OGT-MUT消融。目的3分析TET2和TET2的失调- 在人类和小鼠狼疮B细胞中介导的表观遗传机制，解决了TET2在狼疮中的作用，以及 探讨TET2抑制剂和OGT抑制剂ST045849作为狼疮治疗药物。我们的实验将揭开新奇和 整合环境线索以告知狼疮自身抗体反应的有针对性的表观遗传机制。

PROJECT SUMMARY/ABSTRACT. Epigenetics of the autoantibody response in systemic lupus. Like “mature” antibody responses to viruses and bacteria, the lupus autoantibody response requires B cell class- switch DNA recombination (CSR), somatic hypermutation (SHM) and plasma cell differentiation. As we have shown, epigenetic factors, including histone modifiers (such as Sirt1) and inhibitors (such as butyrate) modulate B cell expression of AID (gene: AICDA/Aicda) and Blimp-1 (PRDM1/Prdm1), which are critically for CSR/SHM and plasma cell differentiation, respectively. Towards a better definition of the epigenetic landscape of lupus B cells, we hypothesize that Tet2, a key epigenetic factor, mediates the lupus autoantibody response, as prompted the B cell-intrinsic role of Tet2 in CSR/SHM and plasma cell differentiation (our recent findings). We argue that Tet2 is induced by the stimuli that induce B cell CSR/SHM and plasma cell differentiation, and it is upregulated in lupus B cells. We also argue that Tet2 boosts transcription of Aicda and Prdm1 through its Fe2+ and a- ketoglutarate (a-KG)-dependent catalytic activity for DNA demethylation as well as its non-catalytic function, i.e., recruiting Ogt (encoded by X-linked OGT/Ogt) to these loci to effect histone glycosylation. Finally, we contend that B cell Tet2 transduces hormonal, nutritional and metabolic cues into epigenetic changes to modulate autoantibody responses, as Tet2 transcription could be upregulated by estrogen (E2, our preliminary data) and vitamin A, and Tet2 is activated by vitamin C, but inhibited by fumarate (a metabolite and a-KG competitor) – as shown by us, E2 boosts AID expression, which would enhance females’ antibody and autoantibody responses. With extensive experience in and commitment to the mechanistic understanding of human and mouse lupus autoantibody responses, we are uniquely poised to test our hypotheses using molecular B cell biology systems, cutting-edge epigenetic tools (hydroxylmethyl DNA analysis, bisulfite and oxidative bisulfite conversion, ChIP and ATAC-Seq), genetically modified mice (TgAicda-creTet2fl/fl, Tet2HxD-mut/HxD-mut and Tet2Ogt-mut/Ogt-mut knockin mice, TgAicda-creOgtfl/fl, Tet2HxD-mut/HxD-mutTgAicda-creOgtfl/fl, MRL/Lpr TgAicda-creTet2fl/fl, MRL/Lpr Tet2HxD-mut/HxD-mut and MRL/Lpr TgAicda-creOgtfl/fl), proprietary humanized H-Mice® and Lupus-H-Mice® models. Aim 1 addresses human and mouse B cell differentiation stage-specific (resting, activated, plasma and memory cell) regulation of Tet2, Tet2 protein stability, E2 upregulation of Tet2 transcripts and underlying mechanisms. Aim 2 addresses the B cell- intrinsic role of Tet2 in promoting AID and Blimp-1 expression, the underlying mechanisms (active DNA demethylation and Ogt-mediated histone glycosylation), and potentiation effect of Tet2 inducer vitamin A and activator vitamin C, and suppressive effect of Tet2 catalytic inhibitor (fumarate, TET-IN-C35 or Bobcat339) alone or combined with Ogt ablation or Tet2Ogt-mut/Ogt-mut knockin. Aim 3 analyzes dysregulation of Tet2 and Tet2- mediated epigenetic mechanisms in human and mouse lupus B cells, addresses the role of Tet2 in lupus, and explores Tet2 inhibitors and Ogt inhibitor ST045849 as lupus therapeutics. Our experiments will unveil novel and targetable epigenetic mechanisms that integrate environmental cues to inform the lupus autoantibody response.