cd28/ctla4/icos基因簇内功能性SNP的鉴定

cd28/ctla4/icos region is very important in immune regulation and immune diseases. SNPs in this region have been reported as disease risk loci in plenty of association studies, but so far the functional SNP in this region has not been identified. We found one DNase hypersensitive site (DHS) in the intron of ctla4 has different open chromatin structure during T cells activation and is related with CTLA4 expression. This region was enriched for H3K4me1 modification, suggested it may function as enhancer during gene regulation.?It located in the same LD block with other published disease associated SNPs and has its genetics polymorphism as seeing from dbSNP database. We proposed that the functional SNP for this region is located in this DHS site. We will first sequencing and construct the haplotype of this region in Chinese Han population. And then using DNase-qPCR, qRT-PCR, Reporter assay, EMSA and ChIP assay to identify the functional SNP and study the mechanism of its functional during gene regulation. Our results will identify the functional SNP in this important region and reveal its mechanism and also may be able to provide more direct markers for immune related diseases risk prediction and evaluation.

cd28/ctla4/icos是重要的免疫功能区域，其SNP位点已被大量研究证实与多种疾病的发病风险有关，但该区域功能性SNP未得到鉴定。我们前期发现ctla4内含子的一段区域在CTLA4表达的过程中存在染色质结构开放状态的变化，ChIP结果表明该区域具有增强子元件的H3K4me1组蛋白修饰模式。该区域为唯一与ctla4表达有关的功能元件，并且与已报道的关联性SNP位于同一LD block，dbSNP数据显示其具有遗传多态性。为此，我们认为基因簇内的功能性SNP可能存在于该区域。本课题将通过重测序构建汉族人群该区域遗传多态性位点的单体型；然后通过DNase-qPCR、定量PCR、报告基因、EMSA、ChIP等技术对其功能性SNP进行研究。结果将有望找到该区域功能性SNP位点并阐明其影响基因表达的机制，亦可为相关疾病风险评估提供更直接靶点。

鉴定与疾病风险相关的功能性SNP是功能基因组时代重要的研究内容。cd28/icos/ctla4是重要的免疫功能区域，该区域多个SNP位点被认为与包括自身免疫性疾病在内的多种疾病相关，但功能性SNP还未得到明确鉴定。本研究通过重测序构建单体型，结合开放染色质结构和表观遗传学修饰的变化，以及报告基因、定量PCR、ChIP和CRISPR等实验，发现CTLA-4一内含子区域位点可能是cd28/icos/ctla4基因簇内的功能性SNP位点，该位点影响基因表达可能与隔离子活性和CTCT结合有关。该区域功能性位点的明确有望解释其附近基因表达调控的机制，亦可为相关疾病风险评估提供靶点。

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