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Abstract：
UNASSIGNED: Cis-regulatory elements (CREs) play crucial roles in regulating gene expression during erythroid cell differentiation. Genome-wide erythroid-specific CREs have not been characterized in chicken erythroid cells, which is an organism model used to study epigenetic regulation during erythropoiesis.
UNASSIGNED: Analysis of public genome-wide accessibility (ATAC-seq) maps, along with transcription factor (TF) motif analysis, CTCF, and RNA Pol II occupancy, as well as transcriptome analysis in fibroblasts and erythroid HD3 cells, were used to characterize erythroid-specific CREs. An α-globin CRE was identified, and its regulatory activity was validated in vitro and in vivo by luciferase activity and genome-editing assays in HD3 cells, respectively. Additionally, circular chromosome conformation capture (UMI-4C) assays were used to distinguish its role in structuring the α-globin domain in erythroid chicken cells.
UNASSIGNED: Erythroid-specific CREs displayed occupancy by erythroid TF binding motifs, CTCF, and RNA Pol II, as well as an association with genes involved in hematopoiesis and cell differentiation. An α-globin CRE, referred to as CRE-2, was identified as exhibiting enhancer activity over αD and αA genes in vitro and in vivo. Induction of terminal erythroid differentiation showed that α-globin CRE-2 is required for the induction of αD and αA. Analysis of TF binding motifs at α-globin CRE-2 shows apparent regulation mediated by GATA-1, YY1, and CTCF binding.
UNASSIGNED: Our findings demonstrate that cell-specific CREs constitute a key mechanism that contributes to the fine-tuning gene regulation of erythroid cell differentiation and provide insights into the annotation and characterization of CREs in chicken cells.
UNASSIGNED: Analysis of public genome-wide accessibility (ATAC-seq) maps, along with transcription factor (TF) motif analysis, CTCF, and RNA Pol II occupancy, as well as transcriptome analysis in fibroblasts and erythroid HD3 cells, were used to characterize erythroid-specific CREs. An α-globin CRE was identified, and its regulatory activity was validated in vitro and in vivo by luciferase activity and genome-editing assays in HD3 cells, respectively. Additionally, circular chromosome conformation capture (UMI-4C) assays were used to distinguish its role in structuring the α-globin domain in erythroid chicken cells.
UNASSIGNED: Erythroid-specific CREs displayed occupancy by erythroid TF binding motifs, CTCF, and RNA Pol II, as well as an association with genes involved in hematopoiesis and cell differentiation. An α-globin CRE, referred to as CRE-2, was identified as exhibiting enhancer activity over αD and αA genes in vitro and in vivo. Induction of terminal erythroid differentiation showed that α-globin CRE-2 is required for the induction of αD and αA. Analysis of TF binding motifs at α-globin CRE-2 shows apparent regulation mediated by GATA-1, YY1, and CTCF binding.
UNASSIGNED: Our findings demonstrate that cell-specific CREs constitute a key mechanism that contributes to the fine-tuning gene regulation of erythroid cell differentiation and provide insights into the annotation and characterization of CREs in chicken cells.
摘要：
■顺式调节元件（CREs）在红系细胞分化过程中调节基因表达中起着至关重要的作用。在鸡红系细胞中尚未表征全基因组红系特异性CRE，这是一种用于研究红细胞生成过程中表观遗传调控的生物体模型。
■对公共全基因组可及性（ATAC-seq）图谱的分析，连同转录因子（TF）基序分析，CTCF,和RNAPolII占用率，以及成纤维细胞和红系HD3细胞的转录组分析，用于表征红系特异性CRE。确定了α-珠蛋白CRE，在HD3细胞中通过荧光素酶活性和基因组编辑实验在体外和体内验证了其调节活性，分别。此外,环形染色体构象捕获（UMI-4C）测定用于区分其在红系鸡细胞中构建α-珠蛋白结构域中的作用。
■红系特异性CREs被红系TF结合基序占据，CTCF,和RNAPolII,以及与造血和细胞分化相关的基因。α-珠蛋白CRE，CRE-2被鉴定为在体外和体内对αD和αA基因表现出增强子活性。诱导终末红细胞分化表明α-珠蛋白CRE-2是诱导αD和αA所必需的。对α-珠蛋白CRE-2上的TF结合基序的分析显示了由GATA-1，YY1和CTCF结合介导的明显调节。
■我们的发现表明，细胞特异性CREs构成了一个关键机制，有助于微调红系细胞分化的基因调控，并提供了对鸡细胞CREs的注释和表征的见解。
■对公共全基因组可及性（ATAC-seq）图谱的分析，连同转录因子（TF）基序分析，CTCF,和RNAPolII占用率，以及成纤维细胞和红系HD3细胞的转录组分析，用于表征红系特异性CRE。确定了α-珠蛋白CRE，在HD3细胞中通过荧光素酶活性和基因组编辑实验在体外和体内验证了其调节活性，分别。此外,环形染色体构象捕获（UMI-4C）测定用于区分其在红系鸡细胞中构建α-珠蛋白结构域中的作用。
■红系特异性CREs被红系TF结合基序占据，CTCF,和RNAPolII,以及与造血和细胞分化相关的基因。α-珠蛋白CRE，CRE-2被鉴定为在体外和体内对αD和αA基因表现出增强子活性。诱导终末红细胞分化表明α-珠蛋白CRE-2是诱导αD和αA所必需的。对α-珠蛋白CRE-2上的TF结合基序的分析显示了由GATA-1，YY1和CTCF结合介导的明显调节。
■我们的发现表明，细胞特异性CREs构成了一个关键机制，有助于微调红系细胞分化的基因调控，并提供了对鸡细胞CREs的注释和表征的见解。
