Background

The i-motif is a tetrameric DNA structure based on the formation of hemiprotonated cytosine-cytosine (C⁺.C) base pairs. i-Motifs are widely used in nanotechnology. In biological systems, i-motifs are involved in gene regulation and in control of genome integrity. In vivo, the i-motif forming sequences are subjects of epigenetic modifications, particularly 5-cytosine methylation. In plants, natively occurring methylation patterns lead to a complex network of C⁺.C, ^5mC⁺.C and ^5mC⁺.^5mC base-pairs in the i-motif stem. The impact of complex methylation patterns (CMPs) on i-motif formation propensity is currently unknown.

Methods

We employed CD and UV-absorbance spectroscopies, native PAGE, thermal denaturation and quantum-chemical calculations to analyse the effects of native, native-like, and non-native CMPs in the i-motif stem on the i-motif stability and pK_a.

Results

CMPs have strong influence on i-motif stability and pK_a and influence these parameters in sequence-specific manner. In contrast to a general belief, i) CMPs do not invariably stabilize the i-motif, and ii) when the CMPs do stabilize the i-motif, the extent of the stabilization depends (in a complex manner) on the number and pattern of symmetric ^5mC⁺.^5mC or asymmetric ^5mC⁺.C base pairs in the i-motif stem.

Conclusions

CMPs can be effectively used to fine-tune i-motif properties. Our data support the notion of epigenetic modifications as a plausible control mechanism of i-motif formation in vivo.

General Significance

Our results have implications in epigenetic regulation of telomeric DNA in plants and highlight the potential and limitations of engineered patterning of cytosine methylations on the i-motif scaffold in nanotechnological applications.

中文翻译：

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胞嘧啶的复合5-甲基化以序列特异性方式调节i-基序稳定性：对DNA纳米技术和植物端粒DNA的表观遗传调控的影响。

背景

i-基序是基于半质子化胞嘧啶-胞嘧啶（C ⁺ .C）碱基对形成的四聚体DNA结构。i-Motifs广泛用于纳米技术。在生物系统中，i-基序参与基因调节和基因组完整性控制。在体内，i-基序形成序列是表观遗传修饰，特别是5-胞嘧啶甲基化的对象。在植物中，天然存在的甲基化模式导致C ⁺ .C，^5m C ⁺ .C和^5m C ⁺形成复杂的网络。i-基序茎中有^5m个C碱基对。目前尚不清楚复杂甲基化模式（CMP）对i-基序形成倾向的影响。

方法

我们采用了CD和UV吸收光谱，天然PAGE，热变性和量子化学计算来分析i-基序茎中的天然，类天然和非天然CMP对i-基序稳定性和pK _{a的影响。}。

结果

CMP对i-基序稳定性和pK _a有很大影响，并以序列特异性方式影响这些参数。与一般的看法相反，i）CMP不会始终稳定i-基元，并且ii）当CMP确实稳定了i-基元时，稳定程度取决于（复杂的方式）取决于i-基元的数量和样式。对称^5m C ⁺。i-基序茎中有^5m C或不对称的^5m C ⁺ .C碱基对。

结论

CMP可以有效地用于微调i-motif属性。我们的数据支持表观遗传修饰的概念作为体内i-基序形成的合理控制机制。

一般意义

我们的研究结果对植物端粒DNA的表观遗传调控有重要意义，并突出了纳米技术应用中i-基序支架上胞嘧啶甲基化工程化模式的潜力和局限性。