Department of Chemical and Biological Engineering, University of Colorado Boulder, Boulder, Colorado 80303, United States.
Antimicrobial Regeneration Consortium, Boulder, Colorado 80301, United States.
ACS Synth Biol. 2021 Apr 16;10(4):737-748. doi: 10.1021/acssynbio.0c00534. Epub 2021 Mar 12.
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 another . 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.
反义转录在所有生命领域都很普遍，通过转录干扰 (TI) 影响基因表达，其中一个转录过程对另一个转录过程产生负面影响。RNA 聚合酶 (RNAP) 的连续性或不间断转录与细菌基因组中反义转录的水平密切相关，但它对 TI 的影响虽然可能很重要，但尚未得到很好的描述。在这里，我们通过三种不同的终止抑制策略（抗生素双环霉素、噬菌体蛋白 Psu 和核糖体-RNAP 偶联）展示了通过连续性控制来调节 TI。我们将这些方法应用于 TI，并调节核糖体-RNAP 偶联，以产生由于 RNAP 碰撞和反义 RNA 干扰导致的 38 倍转录水平基因抑制。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。这些结果共同表明了连续性控制对强 TI 的重要性，并展示了 TI 在合成生物学中的潜力。
