Engineering Transcriptional Interference through RNA Polymerase Processivity Control

Engineering Transcriptional Interference through RNA Polymerase Processivity Control
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通过 RNA 聚合酶持续过程控制工程转录干扰

DOI:
10.1021/acssynbio.0c00534
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发表时间:
2021
影响因子:
4.7
通讯作者:
Chatterjee, Anushree
Chatterjee, Anushree
中科院分区:
生物学2区
文献类型:
--
作者:
O’Connor, Nolan J.;Bordoy, Antoni E.;Chatterjee, Anushree

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反义转录广泛存在于生命的各个领域,并已被证明可以通过转录干扰(TI)影响基因表达,这是一种顺式转录过程对另一个转录过程产生负面影响的现象。 RNA 聚合酶 (RNAP) 的持续性或不间断转录与细菌基因组中的反义转录水平密切相关,但它对 TI 的影响虽然可能很重要,但尚未得到充分表征。在这里,我们表明可以通过三种不同的抗终止策略通过持续性控制来调整 TI:抗生素双环霉素、噬菌体蛋白 Psu 和核糖体-RNAP 偶联。我们将这些方法应用于 TI 并调整核糖体-RNAP 耦合,由于 RNAP 碰撞和反义 RNA 干扰而产生 38 倍的转录水平基因抑制。然后,我们将蛋白质路障和 TI 结合起来,设计最小的遗传 NAND 和 NOR 逻辑门。总之,这些结果表明了强 TI 的持续性控制的重要性,并证明了 TI 在合成生物学方面的潜力。
Antisense transcription is widespread in all kingdoms of life and has been shown to influence gene expression through transcriptional interference (TI), a phenomenon in which one transcriptional process negatively influences anotherin cis. The processivity, or uninterrupted transcription, of an RNA polymerase (RNAP) is closely tied to levels of antisense transcription in bacterial genomes, but its influence on TI, while likely important, is not well-characterized. Here, we show that TI can be tuned through processivity control via three distinct antitermination strategies: the antibiotic bicyclomycin, phage protein Psu, and ribosome-RNAP coupling. We apply these methods toward TI and tune ribosome-RNAP coupling to produce 38-fold transcription-level gene repression due to both RNAP collisions and antisense RNA interference. We then couple protein roadblock and TI to design minimal genetic NAND and NOR logic gates. Together, these results show the importance of processivity control for strong TI and demonstrate TI’s potential for synthetic biology.
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