The Incorporation of Ribonucleotides Induces Structural and Conformational Changes in DNA.
The Incorporation of Ribonucleotides Induces Structural and Conformational Changes in DNA.
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DOI：
10.1016/j.bpj.2017.07.013
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发表时间：
2017-10-03
影响因子：
3.4
通讯作者：
Podestà A
中科院分区：
文献类型：
--
作者：
Meroni A;Mentegari E;Crespan E;Muzi-Falconi M;Lazzaro F;Podestà A
核糖核苷酸掺入是DNA复制过程中最常见的错误。因此，细胞已经发展出从基因组中移除核糖核苷酸并恢复其完整性的机制。的确，核糖核苷酸进入DNA会导致严重的后果，如基因组不稳定和复制压力。因此，从它们对DNA结构和构象的影响入手，了解核糖核苷酸掺入的影响变得非常重要。在这里，我们提出了一个系统的研究，核糖核酸掺入DNA分子的影响。据我们所知，我们已经开发出一种新的方法，可以有效地合成含有核糖核苷酸的长DNA分子(数百个碱基对)，该方法基于一种改进的聚合酶链式反应方法。因此，通过原子力显微镜，我们可以在单分子水平上研究核糖核苷酸掺入后，众多DNA群体的结构和构象性质的变化。具体地，我们刻画了等高线长度与基点数目的比例以及端到端距离与曲线距离、弯曲角分布和持续长度的比例关系。我们的结果显示，核糖核苷酸对DNA结构和构象的影响远远超出了单一核糖核苷酸的典型维度。特别是，核糖核苷酸的存在导致分子的系统性缩短，以及持续长度的减少。这种结构变化也可能发生在体内，在体内，它们可能直接影响下游的DNA交易，以及干扰蛋白质的结合和识别。
Ribonucleotide incorporation is the most common error occurring during DNA replication. Cells have hence developed mechanisms to remove ribonucleotides from the genome and restore its integrity. Indeed, the persistence of ribonucleotides into DNA leads to severe consequences, such as genome instability and replication stress. Thus, it becomes important to understand the effects of ribonucleotides incorporation, starting from their impact on DNA structure and conformation. Here we present a systematic study of the effects of ribonucleotide incorporation into DNA molecules. We have developed, to our knowledge, a new method to efficiently synthesize long DNA molecules (hundreds of basepairs) containing ribonucleotides, which is based on a modified protocol for the polymerase chain reaction. By means of atomic force microscopy, we could therefore investigate the changes, upon ribonucleotide incorporation, of the structural and conformational properties of numerous DNA populations at the single-molecule level. Specifically, we characterized the scaling of the contour length with the number of basepairs and the scaling of the end-to-end distance with the curvilinear distance, the bending angle distribution, and the persistence length. Our results revealed that ribonucleotides affect DNA structure and conformation on scales that go well beyond the typical dimension of the single ribonucleotide. In particular, the presence of ribonucleotides induces a systematic shortening of the molecules, together with a decrease of the persistence length. Such structural changes are also likely to occur in vivo, where they could directly affect the downstream DNA transactions, as well as interfere with protein binding and recognition.
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影响因子：
16
作者：
Ghodgaonkar, Medini Manohar;Lazzaro, Federico;Olivera-Pimentel, Maite;Artola-Boran, Mariela;Cejka, Petr;Reijns, Martin A.;Jackson, Andrew P.;Plevani, Paolo;Muzi-Falconi, Marco;Jiricny, Josef
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Jiricny, Josef
影响因子：
3.3
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影响因子：
15
作者：
Herrero-Galan, Elias;Eugenia Fuentes-Perez, Maria;Ricardo Arias-Gonzalez, J.
通讯作者：
Ricardo Arias-Gonzalez, J.
影响因子：
3.4
作者：
Hormeno, Silvia;Ibarra, Borja;Ricardo Arias-Gonzalez, J.
通讯作者：
Ricardo Arias-Gonzalez, J.
影响因子：
3.4
作者：
Ivanov, VI;Minchenkova, LE;Zimmer, C
通讯作者：
Zimmer, C