Fishery College, Guangdong Ocean University, Zhanjiang, China.
Pearl Breeding and Processing Engineering Technology Research Centre of Guangdong Province, Zhanjiang, China.
Front Immunol. 2023 Oct 3;14:1247544. doi: 10.3389/fimmu.2023.1247544. eCollection 2023.
In the pearl culture industry, a major challenge is the overactive immunological response in pearl oysters resulting from allotransplantation, leading to shell-bead rejection and death. To better understand the molecular mechanisms of postoperative recovery and the regulatory role of DNA methylation in gene expression, we analyzed the changes in DNA methylation levels after allotransplantation in pearl oyster , and elucidated the regulatory function of DNA methylation in promoter activity of () gene.
We constructed nine DNA methylomes at different time points after allotransplantation and used bisulfite genomic sequencing PCR technology (BSP) to verify the methylation status in the promoter of . We performed Dual luciferase assays to determine the effect of the dense methylation region in the promoter on transcriptional activity and used DNA pull-down and mass spectrometry analysis to assess the capability of transcription factor binding with the dense methylation region.
The DNA methylomes reveal that CG-type methylation is predominant, with a trend opposite to non-CG-type methylation. Promoters, particularly CpG island-rich regions, were less frequently methylated than gene function elements. We identified 5,679 to 7,945 differentially methylated genes (DMGs) in the gene body, and 2,146 to 3,385 DMGs in the promoter at each time point compared to the pre-grafting group. Gene ontology and pathway enrichment analyses showed that these DMGs were mainly associated with "cellular process", "Membrane", "Epstein-Barr virus infection", "Notch signaling pathway", "Fanconi anemia pathway", and "Nucleotide excision repair". Our study also found that the DNA methylation patterns of the promoter region of gene were consistent with the DNA methylomics data. We further demonstrated that the dense methylation region in the promoter of affects transcriptional activity, and that the methylation status in the promoter modulates the binding of different transcription factors, particularly transcriptional repressors.
These findings enhance our understanding of the immune response and regulation mechanism induced by DNA methylation in pearl oysters after allotransplantation.
在珍珠养殖行业，一个主要的挑战是珍珠贝异体移植后的过度免疫反应，导致壳珠排斥和死亡。为了更好地了解术后恢复的分子机制和 DNA 甲基化在基因表达中的调控作用，我们分析了珍珠贝异体移植后 DNA 甲基化水平的变化，并阐明了 DNA 甲基化对 （）基因启动子活性的调控功能。
我们构建了九个异体移植后不同时间点的 DNA 甲基组，并用亚硫酸氢盐基因组测序 PCR 技术（BSP）验证启动子中的甲基化状态。我们进行了双荧光素酶测定，以确定启动子中密集甲基化区域对转录活性的影响，并使用 DNA 下拉和质谱分析来评估转录因子与密集甲基化区域的结合能力。
DNA 甲基组揭示 CG 型甲基化占主导地位，与非 CG 型甲基化趋势相反。启动子，特别是富含 CpG 岛的区域，比基因功能元件的甲基化程度低。与术前相比，我们在基因体中鉴定到 5679 到 7945 个差异甲基化基因（DMGs），在启动子中鉴定到 2146 到 3385 个 DMGs。基因本体和通路富集分析表明，这些 DMGs 主要与“细胞过程”、“膜”、“Epstein-Barr 病毒感染”、“Notch 信号通路”、“范可尼贫血通路”和“核苷酸切除修复”有关。我们的研究还发现， 基因启动子区域的 DNA 甲基化模式与 DNA 甲基组学数据一致。我们进一步证明，启动子中 基因的密集甲基化区域影响转录活性，启动子中的甲基化状态调节不同转录因子的结合，特别是转录抑制因子。
这些发现增强了我们对珍珠贝异体移植后 DNA 甲基化诱导的免疫反应和调控机制的理解。
