Xiao Yao, Liu Huan
Chin J Dent Res. 2020;23(2):119-130. doi: 10.3290/j.cjdr.a44748.
To investigate and characterise the differences between the open chromatin regions of oral and epidermal keratinocytes.
Human immortalised oral epithelial cell lines (HIOECs) were used as the standard model for oral keratinocytes, and primary normal human epidermal keratinocytes (NHEKs) were chosen as the model for epidermal keratinocytes. Assay for transposase accessible chromatin using sequencing (ATAC-seq) and H3K27ac chromatin immunoprecipitation sequencing (ChIP-seq) were used to evaluate the dynamic changes in open chromatin regions and active enhancers during oral keratinocyte differentiation. In silico prediction and dual-luciferase assays were used to evaluate the enriched motifs and maintain enhancer activity in specific enriched HIOECs. Integration and comparison of HIOEC ATAC-seq with NHEK ATAC-seq were used to identify oral keratinocyte-enriched open chromatin regions along with key motifs governing differential enhancer activity. The genomic regulatory elements and GWAS overlap algorithm was used to compare the annotation rate of HIOEC-overlapped craniofacial enhancers with other craniofacial enhancers for orofacial cleft-associated variants.
During the differentiation of HIOECs, 14933 open chromatin regions became more accessible. Grainyhead-like (GRHL) and Krüppel-like factor (KLF) motifs were overrepresented in maintaining HIOEC-specific activity. Compared with NHEKs, 16161 open chromatin regions were uniquely accessible in HIOECs. Within these regions, the C/EBP motif governed HIOEC-specific enhancer regulating SOX2 and PITX2, which enhanced oral keratinocyte wound healing. When intersected with human craniofacial super-enhancers, open chromatin regions in HIOECS can better annotate the common variants associated with orofacial cleft.
The intrinsic differences between the open chromatin regions of human oral and epidermal keratinocytes are directly maintained by a set of transcription factors.
研究并表征口腔角质形成细胞和表皮角质形成细胞开放染色质区域之间的差异。
将人永生化口腔上皮细胞系（HIOECs）用作口腔角质形成细胞的标准模型，并选择原代正常人表皮角质形成细胞（NHEKs）作为表皮角质形成细胞的模型。采用转座酶可及染色质测序分析（ATAC-seq）和H3K27ac染色质免疫沉淀测序（ChIP-seq）来评估口腔角质形成细胞分化过程中开放染色质区域和活性增强子的动态变化。利用计算机预测和双荧光素酶测定来评估特定富集的HIOECs中富集基序并维持增强子活性。将HIOEC ATAC-seq与NHEK ATAC-seq进行整合和比较，以鉴定口腔角质形成细胞富集的开放染色质区域以及控制差异增强子活性的关键基序。使用基因组调控元件和GWAS重叠算法来比较HIOEC重叠的颅面增强子与其他颅面增强子对口腔颌面部裂隙相关变体的注释率。
在HIOECs分化过程中，14933个开放染色质区域变得更容易接近。颗粒头样（GRHL）和Krüppel样因子（KLF）基序在维持HIOEC特异性活性方面过度富集。与NHEKs相比，HIOECs中有16161个开放染色质区域是独特可及的。在这些区域内，C/EBP基序控制着调节SOX2和PITX2的HIOEC特异性增强子，从而增强口腔角质形成细胞的伤口愈合。当与人类颅面超级增强子相交时，HIOECS中的开放染色质区域可以更好地注释与口腔颌面部裂隙相关的常见变体。
人类口腔和表皮角质形成细胞开放染色质区域之间的内在差异由一组转录因子直接维持。
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