ChIP-chip of monocytes using H3K9Ac, H3K4me3, H3K9me2 and H4K16Ac antibodies

Both genetic and environmental factors are implicated in Type 1 Diabetes (T1D). Since environmental factors can trigger epigenetic changes, we hypothesized that variations in histone posttranslational modifications (PTMs) at the promoter/enhancer regions of T1D susceptible genes may be associated with T1D. We therefore evaluated histone PTM variations at known T1D susceptible genes in blood cells from T1D patients versus healthy non-diabetic controls, and explored their connections to T1D. We used the chromatin-immunoprecipitation-linked-to-microarray approach to profile key histone PTMs, including H3-lysine-4 trimethylation (H3K4me3), H3K27me3, H3K9me3, H3K9 acetylation (H3K9Ac) and H4K16Ac at genes within the T1D susceptible loci in lymphocytes, and H3K4me3, H3K9me2, H3K9Ac and H4K16Ac at the IDDM1 region in monocytes of T1D patients and healthy controls separately. We screened for potential variations in histone PTMs using computational methods to compare datasets from T1D and controls. Interestingly, we observed marked variations in H3K9Ac levels at the upstream regions of HLA-DRB1 and HLA-DQB1 within the IDDM1 locus in T1D monocytes relative to controls. Additional experiments with THP-1 monocytes demonstrated increased expression of HLA-DRB1 and HLA-DQB1 in response to interferon- and TNF-treatment that were accompanied by changes in H3K9Ac at the same promoter regions as that seen in the patient monocytes. These results suggest that the H3K9Ac status of HLA-DRB1 and HLA-DQB1, two genes highly associated with T1D, may be relevant to their regulation and transcriptional response towards external stimuli. Thus, the promoter/enhancer architecture and chromatin status of key susceptible loci could be important determinants in their functional association to T1D susceptibility. We used ChIP-chip to profile key histone PTMs, including H3K4me3, H3K9me2, H3K9Ac and H4K16Ac, at the IDDM1 region in monocytes of T1D patients and healthy controls.

遗传与环境因素均与1型糖尿病（Type 1 Diabetes, T1D）的发病机制相关。鉴于环境因素可诱发表观遗传改变，我们提出假说：1型糖尿病易感基因启动子/增强子区域的组蛋白翻译后修饰（histone posttranslational modifications, PTMs）变异或与1型糖尿病存在关联。为此，我们对比分析了1型糖尿病患者与健康非糖尿病对照者血细胞中已知1型糖尿病易感基因的组蛋白PTM变异情况，并探讨其与1型糖尿病的关联。我们采用染色质免疫沉淀微阵列联用（chromatin-immunoprecipitation-linked-to-microarray, ChIP-chip）技术，对淋巴细胞内1型糖尿病易感基因位点的关键组蛋白PTM进行图谱分析，包括H3赖氨酸4三甲基化（H3-lysine-4 trimethylation, H3K4me3）、H3K27me3、H3K9me3、H3K9乙酰化（H3K9 acetylation, H3K9Ac）及H4K16Ac；同时分别对1型糖尿病患者与健康对照者单核细胞内IDDM1区域的H3K4me3、H3K9me2、H3K9Ac及H4K16Ac进行图谱分析。我们通过计算方法对比1型糖尿病患者与对照组的数据集，筛选出潜在的组蛋白PTM变异。值得注意的是，相较于对照组，1型糖尿病患者单核细胞中IDDM1位点内HLA-DRB1与HLA-DQB1上游区域的H3K9Ac水平存在显著差异。后续针对THP-1单核细胞的补充实验显示，经干扰素与肿瘤坏死因子（tumor necrosis factor, TNF）处理后，HLA-DRB1与HLA-DQB1的表达水平升高，且其同一启动子区域的H3K9Ac发生改变，与患者单核细胞中的变化一致。上述结果表明，与1型糖尿病高度相关的HLA-DRB1与HLA-DQB1的H3K9Ac状态，或与其基因调控及对外界刺激的转录应答相关。因此，关键易感基因位点的启动子/增强子结构与染色质状态，可能是决定其与1型糖尿病易感性功能关联的重要因素。我们采用ChIP-chip技术，对1型糖尿病患者与健康对照者单核细胞内IDDM1区域的关键组蛋白PTM（包括H3K4me3、H3K9me2、H3K9Ac及H4K16Ac）进行了图谱分析。