Importance of the tmRNA system for cell survival when transcription is blocked by DNA-protein cross-links.
Importance of the tmRNA system for cell survival when transcription is blocked by DNA-protein cross-links.
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DOI：
10.1111/j.1365-2958.2010.07355.x
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发表时间：
2010-11
影响因子：
3.6
通讯作者：
Kreuzer KN
中科院分区：
文献类型：
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作者：
Kuo HK;Krasich R;Bhagwat AS;Kreuzer KN
抗癌药物5-氮杂胞苷（aza-C）可诱导胞嘧啶甲基转移酶和DNA之间的DNA-蛋白质交联（DPC），因为该药物抑制甲基化。我们发现在tmRNA翻译质量控制系统中有缺陷的突变体对aza-C过敏。超敏反应需要活性甲基转移酶的表达，表明DPC形成的重要性。此外，在表达甲基转移酶的细胞中，经aza-C处理后，tmRNA途径被激活，导致SsrA标记蛋白的水平增加。这些结果表明，tmRNA途径清除了aza-C诱导的DPC转录阻断后产生的停滞的核糖体-mRNA复合物。作为支持，ssrA突变体对利迪链菌素也是超敏感的，利迪链菌素通过不同的机制阻断RNA聚合酶延伸。tmRNA途径被认为仅作用于含有靠近A位点的3' RNA末端的核糖体，并且已知的用于从封闭的聚合酶释放RNA 3'末端的途径涉及Mfd解旋酶。然而，Mfd敲除突变体对氮杂-C诱导的DPC形成或链球菌溶血素不敏感，表明Mfd不参与。转录终止因子Rho也可能不参与，因为Rho特异性抑制剂双环霉素未能显示与aza-C或streptolydigin的协同作用。基于这些发现，我们讨论了模型如何E。大肠杆菌处理转录/翻译复合物，其在DPC处被阻断。
Anticancer drug 5-azacytidine (aza-C) induces DNA-protein crosslinks (DPCs) between cytosine methyltransferase and DNA as the drug inhibits methylation. We found that mutants defective in the tmRNA translational quality control system are hypersensitive to aza-C. Hypersensitivity requires expression of active methyltransferase, indicating the importance of DPC formation. Furthermore, the tmRNA pathway is activated upon aza-C treatment in cells expressing methyltransferase, resulting in increased levels of SsrA tagged proteins. These results argue that the tmRNA pathway clears stalled ribosome-mRNA complexes generated after transcriptional blockage by aza-C-induced DPCs. In support, an ssrA mutant is also hypersensitive to streptolydigin, which blocks RNA polymerase elongation by a different mechanism. The tmRNA pathway is thought to act only on ribosomes containing a 3’ RNA end near the A site, and the known pathway for releasing RNA 3’ ends from a blocked polymerase involves Mfd helicase. However, an mfd knockout mutant is not hypersensitive to either aza-C-induced DPC formation or streptolydigin, indicating that Mfd is not involved. Transcription termination factor Rho is also likely not involved, because the Rho-specific inhibitor bicyclomycin failed to show synergism with either aza-C or streptolydigin. Based on these findings, we discuss models for how E. coli processes transcription/translation complexes blocked at DPCs.
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