The autoregulator Aca2 mediates anti-CRISPR repression
The autoregulator Aca2 mediates anti-CRISPR repression
复制标题
DOI：
10.1093/nar/gkz721
复制
发表时间：
2019-10-10
影响因子：
14.9
通讯作者：
Fineran, Peter C.
中科院分区：
文献类型：
--
作者：
Birkholz, Nils;Fagerlund, Robert D.;Fineran, Peter C.
CRISPR-CAS系统是广泛存在的细菌适应性防御机制，提供对噬菌体的保护。作为回应，噬菌体进化出了抗CRISPR蛋白，使宿主的CRISPR-CAS系统失活，从而使成功感染成为可能。抗CRISPR基因常见于编码转录调控基因的操纵子中。这些抗CRISPR相关(AcA)基因在抗CRISPR调控中的作用尚不清楚。在这里，我们证明了由温和胡萝卜杆菌噬菌体ZF40编码的Aca2是一种自动调节因子，它抑制了抗CRISPR-aca2操纵子。Aca2是一个螺旋-转角-螺旋结构域蛋白，它形成一个同源二聚体，并与抗CRISPR启动子中的两个反向重复序列相互作用。反向重复序列在序列上相似，但在Aca2亲和力上不同，我们认为它们已经进化成在噬菌体生命周期的不同阶段微调和下调抗CRISPR的产物。Aca2与第一个反向重复序列特异的高亲和力结合阻止了启动子并诱导DNA弯曲。第二个反向重复序列只在体内高Aca2浓度时起到抑制作用，在体外没有检测到DNA结合。我们的研究揭示了ACA蛋白调节其相关的抗CRISPR表达的机制。
CRISPR-Cas systems are widespread bacterial adaptive defence mechanisms that provide protection against bacteriophages. In response, phages have evolved anti-CRISPR proteins that inactivate CRISPR-Cas systems of their hosts, enabling successful infection. Anti-CRISPR genes are frequently found in operons with genes encoding putative transcriptional regulators. The role, if any, of these anti-CRISPR-associated (aca) genes in anti-CRISPR regulation is unclear. Here, we show that Aca2, encoded by the Pectobacterium carotovorum temperate phage ZF40, is an autoregulator that represses the anti-CRISPR-aca2 operon. Aca2 is a helix-turn-helix domain protein that forms a homodimer and interacts with two inverted repeats in the anti-CRISPR promoter. The inverted repeats are similar in sequence but differ in their Aca2 affinity, and we propose that they have evolved to fine-tune, and downregulate, anti-CRISPR production at different stages of the phage life cycle. Specific, high-affinity binding of Aca2 to the first inverted repeat blocks the promoter and induces DNA bending. The second inverted repeat only contributes to repression at high Aca2 concentrations in vivo, and no DNA binding was detectable in vitro. Our investigation reveals the mechanism by which an Aca protein regulates expression of its associated anti-CRISPR.