Epigenetics of Severe Systemic Inflammation

摘要 表观遗传基因重编程是一个与人类疾病相关的新兴概念。这项建议是 目的探讨重症全身性炎症(SSI)的表观遗传学。SSI血液和组织白细胞 显示受抑制的急性促炎基因转录的基因特异性重新编程，如肿瘤坏死因子和 IL-1，并激活其他基因的转录。SSI基因重编程逆转与临床相关 进步。我们报道了肿瘤坏死因子转录抑制和核小体向赖氨酸9-H3二甲基化转变。 (H3K9me2)、H3丝氨酸10去磷酸化(H3S10)和抑制物异染色质蛋白1(HP-1) SSI细胞模型中的启动子结合。我们进一步发现组蛋白H3赖氨酸甲基转移酶G9a 在肿瘤坏死因子沉默过程中，胞嘧啶甲基转移酶DNMT 3a/b与HP-1和CpG甲基化偶联。 总之，这些结果支持表观遗传机制参与抑制急性炎症。 在SSI患者中观察到的基因和维持通常致命的免疫抑制。我们假设SSI 产生从常染色质(响应性)异染色质(沉默)在近端的启动子 急性促炎基因。目标1将测试常染色质到异染色质的转变是否需要 特定的H3丝氨酸激酶、赖氨酸甲基转移酶和去甲基酶，以及接头或连接物，以及 潜在的可逆转性。目的2将测试急性促炎基因启动子CpG甲基化是否 通过特定的调节蛋白偶联到核小体的移位。目标3将测试表观遗传学 在循环中的人SSI白细胞中，常染色质向异染色质转变。实验 基本目标(目标1和目标2)和翻译目标(目标3)的方法将包括遗传(转基因 SiRNA或表达载体)和生化(染色质免疫沉淀、DNA测序、DNA酶 足迹、免疫印迹和信使核糖核酸定量)分析。 *

Abstract Epigenetic gene reprogramming is an emerging concept relevant to human diseases. This proposal's objective is to investigate epigenetics of severe systemic inflammation (SSI). SSI blood and tissue leukocytes show gene-specific reprogramming with repressed transcription of acute proinflammatory genes like TNF and IL-1 and activated transcription of other gene sets. SSI gene reprogramming reversal correlates with clinical improvement. We reported TNF transcription repression and a nucleosome shift to lysine 9 H3 di-methylation (H3K9me2), H3 serine 10 dephosphorylation(H3S10), and repressor heterochromatin protein 1 (HP-1) promoter binding in an SSI cell model. We further implicate the histone H3 lysine methyltransferase G9a coupled to HP-1 and CpG methylation by cytosine methyltransferase DNMT 3a/b during TNF silencing. Together, these results support that epigenetic mechanisms participate in repressing acute inflammatory genes and sustaining often lethal immunosuppression observed in SSI patients. We hypothesize that SSI generates a shift from a euchromatin (responsive) heterochromatin (silenced) at the proximal promoter of acute proinflammatory genes. Aim 1 will test in whether the euchromatin to heterochromatin shift requires specific H3 serine kinases, lysine methyltransferases and demethylases, and adapters or linkers, and the potential for reversibility. Aim 2 will test whether acute proinflammatory gene promoter CpG methylation is coupled through specific regulatory proteins to the nucleosome shifts. Aim 3 will test whether the epigenetic shift from euchromatin to heterochromatin occurs in circulating human SSI leukocytes. Experimental approaches for the basic (Aims 1 and 2) and translational aims (Aim 3) will include genetic (transfection of siRNA or expression plasmids) and biochemical (chromatin immunoprecipitation, DNA sequencing, DNAase footprinting, immunoblots, and mRNA quantitation) analyses. *