Transcriptional Bypass of DNA-Protein and DNA-Peptide Conjugates by T7 RNA Polymerase.
Transcriptional Bypass of DNA-Protein and DNA-Peptide Conjugates by T7 RNA Polymerase.
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DOI：
10.1021/acschembio.9b00365
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发表时间：
2019-12-20
影响因子：
4
通讯作者：
Tretyakova NY
中科院分区：
文献类型：
--
作者：
Ji S;Thomforde J;Rogers C;Fu I;Broyde S;Tretyakova NY
DNA - 蛋白质交联物（DPCs）是一种异常庞大的DNA加合物，它会阻碍蛋白质与DNA的接触，并干扰基因表达、复制及修复过程。我们之前曾阐述过，在经抗肿瘤氮芥和铂类化合物处理的人类细胞中，DNA的N7 - 鸟嘌呤位点会形成DPCs，并且还表明DPCs可在内源性条件下于DNA表观遗传标记5 - 甲酰基 - dC处形成。然而，关于这些结构各异的DPCs对转录的影响，目前可用信息尚不充分。在本研究中，我们综合运用体外实验、质谱分析以及分子动力学模拟等方法，来检测噬菌体T7 RNA聚合酶绕过与7 - 脱氮 - dG的C7位点和dC的C5位点结合的DPCs的能力。这些模型加合物分别代表因接触抗肿瘤药物而诱导产生的内源性DPCs以及在表观遗传DNA标记处形成的内源性DPCs。我们的研究结果显示，含有全长蛋白质的DPCs会显著抑制T7 RNA聚合酶的体外转录，而短链DNA - 肽交联物（DpCs）则可被绕过。与7 - 脱氮 - dG的C7位点结合的DpCs能够以高保真度进行转录，而连接到dC的C5位点的相同多肽则会诱导转录错误。对与模板链上dC的C5原子或7 - 脱氮 - dG的C7位点结合的DpCs在T7 RNA聚合酶中的分子动力学模拟，解释了结合的肽如何容纳在DNA - RNA杂交体狭窄的大沟中，以及修饰后的dC如何在聚合酶活性位点与进入的ATP形成稳定的错配，从而导致转录诱变。
DNA-protein cross-links (DPCs) are unusually bulky DNA adducts that block the access of proteins to DNA and interfere with gene expression, replication, and repair. We previously described DPC formation at the N7-guanine position of DNA in human cells treated with antitumor nitrogen mustards and platinum compounds and have shown that DPCs can form endogenously at DNA epigenetic mark 5-formyl-dC. However, insufficient information is available about the effects of these structurally distinct DPCs on transcription. In the present work, we employ a combination of in vitro assays, mass spectrometry, and molecular dynamics simulations to examine the ability of phage T7 RNA polymerase to bypass DPCs conjugated to the C7 position of 7-deaza-dG and the C5 position of dC. These model adducts represent endogenous DPCs induced by exposure to antitumor drugs and formed at epigenetics DNA marks, respectively. Our results reveal that DPCs containing full length proteins significantly inhibit in vitro transcription by T7 RNA polymerase, while short DNA-peptide cross-links (DpCs) are bypassed. DpCs conjugated to the C7 position of 7-deaza-dG are transcribed with high fidelity, while the same polypeptides attached to the C5 position of dC induce transcription errors. Molecular dynamics simulations of DpCs conjugated either to the C5 atom of dC or the C7 position of 7-deaza-dG on the template strand in T7 RNA polymerase explain how the conjugated peptide can be accommodated in the narrow major groove of the DNA-RNA hybrid and how the modified dC can form a stable mismatch with the incoming ATP in the polymerase active site, allowing for transcriptional mutagenesis.
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影响因子：
5.5
作者：
Goetz, Andreas W.;Williamson, Mark J.;Xu, Dong;Poole, Duncan;Le Grand, Scott;Walker, Ross C.
通讯作者：
Walker, Ross C.
抗肿瘤氮的DNA-蛋白交联的蛋白质组学分析。
影响因子：
4.1
作者：
Loeber, Rachel L.;Michaelson-Richie, Erin D.;Codreanu, Simona G.;Liebler, Daniel C.;Campbell, Colin R.;Tretyakova, Natalia Y.
通讯作者：
Tretyakova, Natalia Y.
影响因子：
14.9
作者：
Lavery R;Moakher M;Maddocks JH;Petkeviciute D;Zakrzewska K
通讯作者：
Zakrzewska K
影响因子：
64.5
作者：
Duxin JP;Dewar JM;Yardimci H;Walter JC
通讯作者：
Walter JC
影响因子：
4.8
作者：
Kalogeraki, VS;Tornaletti, S;Hanawalt, PC
通讯作者：
Hanawalt, PC