Methylation patterns of papillomavirus DNA, its influence on E2 function, and implications in viral infection
Methylation patterns of papillomavirus DNA, its influence on E2 function, and implications in viral infection
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DOI：
10.1128/jvi.77.23.12450-12459.2003
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发表时间：
2003-12-01
影响因子：
5.4
通讯作者：
Lambert, PF
中科院分区：
文献类型：
--
作者：
Kim, K;Garner-Hamrick, PA;Lambert, PF
乳头瘤病毒E2蛋白在乳头瘤病毒基因组的转录、复制和维持中的生物活性依赖于E2蛋白特异性结合该基因组的能力。人乳头瘤病毒（HPV）基因组长控制区（LCR）内的E2结合位点（E2BS）含有潜在的胞嘧啶（CpG）5 ′端甲基化位点。E2蛋白在体外结合E2 BS的能力受到这些胞嘧啶甲基化的抑制（59）。在此，我们描述了实验来评估甲基化对细胞中E2功能的影响。E2激活转录的能力受到E2响应转录模板中CpG二核苷酸的全局甲基化的抑制，或者当转录模板的E2BS内只有CpG二核苷酸被甲基化时。因此，E2的至少一种生物活性取决于其以位点特异性方式结合DNA的能力，其同源结合位点的甲基化状态影响E2的生物活性。DNA甲基化酶的活性受哺乳动物细胞分化状态的影响。HPV的生命周期与其宿主细胞在复层鳞状上皮内的分化有关。为了研究乳头瘤病毒基因组的甲基化是否受宿主上皮细胞分化状态的影响，我们分析了从HPV 16感染患者分离的宫颈上皮细胞系中收获的HPV 16 DNA。我们发现，使用亚硫酸氢盐处理来区分甲基化和非甲基化胞嘧啶，HPV 16 LCR在高度分化的细胞群中选择性地低甲基化。相比之下，来自低分化基底细胞样细胞的HPV 16 LCR含有多个甲基化胞嘧啶，并且通常在E2BS处甲基化，特别是E2BS。这些实验表明，病毒基因组的甲基化状态，特别是E2BS的甲基化状态，可能在病毒生命周期中发生变化，提供了一种新的手段来调节E2功能。这些研究还揭示了在非CpG二核苷酸处指示从头甲基化的广泛甲基化模式。这种从头，甲基化模式的潜在影响进行了讨论。
The biological activities of the papillomavirus E2 protein in transcription, replication, and maintenance of the papillomavirus genome rely on the E2 protein's ability to bind that genome specifically. The E2 binding sites (E2BSs), located within the long control region (LCR) of human papillomavirus (HPV) genomes, contain potential sites for 5'methylation at cytosine (CpG) residues. The E2 protein's capacity to bind E2BS in vitro is inhibited by methylation of these cytosines (59). Herein, we describe experiments to assess the influence of methylation on E2 function in cells. E2's ability to activate transcription was inhibited by the global methylation of CpG dinucleotides in E2-responsive transcriptional templates or when only the CpG dinucleotides within the E2BSs of a transcriptional template were methylated. Thus at least one biological activity of E2 that is dependent on its ability to bind DNA in a site-specific manner is influenced by the methylation status of its cognate binding site. The activity of DNA methylases is influenced by the differentiation status of mammalian cells. The life cycle of HPVs is tied to the differentiation of its host cells within stratified squamous epithelia. To investigate whether methylation of the papillomavirus genomes is influenced by the differentiation status of host epithelial cells, we analyzed HPV16 DNA harvested from a cervical epithelial cell line that was isolated from an HPV16-infected patient. We found, using bisulfite treatment to discriminate between methylated and unmethylated cytosines, that the HPV16 LCR was selectively hypomethylated in highly differentiated cell populations. In contrast, the HPV16 LCR from poorly differentiated, basal cell-like cells contained multiple methylated cytosines and were often methylated at E2BSs, particularly E2BS. These experiments indicate that the methylation state of the viral genome, and particular that of E2BSs, may vary during the viral life cycle, providing a novel means for modulating E2 function. These studies also uncovered an extensive pattern of methylation at non-CpG dinucleotides indicative of de novo methylation. The potential implications of this de novo, methylation pattern are discussed.