ABSTRACT

The presence of 5-hydroxymethyl cytosine in DNA has been previously associated with ageing. Using in silico analysis of normal liver samples we presently observed that in 5-hydroxymethyl cytosine sequences, DNA methylation is dependent on the co-presence of G-quadruplexes and palindromes. This association exhibits discrete patterns depending on G-quadruplex and palindrome densities. DNase-Seq data show that 5-hydroxymethyl cytosine sequences are common among liver nucleosomes (p < 2.2x10⁻¹⁶) and threefold more frequent than nucleosome sequences. Nucleosomes lacking palindromes and potential G-quadruplexes are rare in vivo (1%) and nucleosome occupancy potential decreases with increasing G-quadruplexes. Palindrome distribution is similar to that previously reported in nucleosomes. In low and mixed complexity sequences 5-hydroxymethyl cytosine is frequently located next to three elements: G-quadruplexes or imperfect G-quadruplexes with CpGs, or unstable hairpin loops (TCCCAY₆TGGGA) mostly located in antisense strands or finally A-/T-rich segments near these motifs. The high frequencies and selective distribution of pentamer sequences (including TCCCA, TGGGA) probably indicate the positive contribution of 5-hydroxymethyl cytosine to stabilize the formation of structures unstable in the absence of this cytosine modification. Common motifs identified in all total 5-hydroxymethyl cytosine-containing sequences exhibit high homology to recognition sites of several transcription factor families: homeobox, factors involved in growth, mortality/ageing, cancer, neuronal function, vision, and reproduction. We conclude that cytosine hydroxymethylation could play a role in the recognition of sequences with G-quadruplexes/palindromes by forming epigenetically regulated DNA ‘springs’ and governing expansions or compressions recognized by different transcription factors or stabilizing nucleosomes. The balance of these epigenetic elements is lost in hepatocellular carcinoma.

摘要

DNA中5-羟甲基胞嘧啶的存在以前与衰老有关。使用对正常肝脏样本的计算机分析，我们目前观察到，在 5-羟甲基胞嘧啶序列中，DNA 甲基化依赖于 G-四链体和回文的共存。这种关联表现出取决于 G-四链体和回文密度的离散模式。DNase-Seq 数据显示 5-羟甲基胞嘧啶序列在肝脏核小体中很常见 ( p < 2.2x10 ^-16) 并且比核小体序列的频率高出三倍。缺乏回文和潜在 G-四链体的核小体在体内很少见 (1%)，并且核小体占据潜力随着 G-四链体的增加而降低。回文分布与先前在核小体中报道的相似。在低复杂度和混合复杂度序列中，5-羟甲基胞嘧啶通常位于三个元素旁边：G-四链体或具有 CpG 的不完美 G-四链体，或不稳定的发夹环 (TCCCAY ₆TGGGA）主要位于这些基序附近的反义链或最后富含 A-/T 的片段中。五聚体序列（包括 TCCCA、TGGGA）的高频率和选择性分布可能表明 5-羟甲基胞嘧啶对稳定在没有这种胞嘧啶修饰的情况下不稳定结构的形成的积极贡献。在所有含有 5-羟甲基胞嘧啶的总序列中鉴定的共同基序与几个转录因子家族的识别位点表现出高度同源性：同源框、参与生长的因子、死亡率/衰老、癌症、神经元功能、视力和繁殖。我们得出结论，胞嘧啶羟甲基化可以通过形成表观遗传调控的 DNA“弹簧”和控制不同转录因子识别的扩张或压缩或稳定核小体，在识别具有 G-四链体/回文的序列中发挥作用。这些表观遗传元素的平衡在肝细胞癌中丧失。