Institute of Biotechnology, Life Sciences Center, Vilnius University, Sauletekio av. 7, 10257 Vilnius, Lithuania.
Laboratory of Algology and Microbial Ecology, Nature Research Centre, Akademijos str. 2, 08412 Vilnius, Lithuania.
Mol Cell. 2020 Dec 17;80(6):955-970.e7. doi: 10.1016/j.molcel.2020.11.034. Epub 2020 Dec 7.
Prokaryotic toxin-antitoxin (TA) systems are composed of a toxin capable of interfering with key cellular processes and its neutralizing antidote, the antitoxin. Here, we focus on the HEPN-MNT TA system encoded in the vicinity of a subtype I-D CRISPR-Cas system in the cyanobacterium Aphanizomenon flos-aquae. We show that HEPN acts as a toxic RNase, which cleaves off 4 nt from the 3' end in a subset of tRNAs, thereby interfering with translation. Surprisingly, we find that the MNT (minimal nucleotidyltransferase) antitoxin inhibits HEPN RNase through covalent di-AMPylation (diadenylylation) of a conserved tyrosine residue, Y109, in the active site loop. Furthermore, we present crystallographic snapshots of the di-AMPylation reaction at different stages that explain the mechanism of HEPN RNase inactivation. Finally, we propose that the HEPN-MNT system functions as a cellular ATP sensor that monitors ATP homeostasis and, at low ATP levels, releases active HEPN toxin.
原核生物毒素-抗毒素（TA）系统由能够干扰关键细胞过程的毒素及其中和解毒剂抗毒素组成。在这里，我们关注的是蓝藻鱼腥藻中临近 I-D 型 CRISPR-Cas 系统编码的 HEPN-MNT TA 系统。我们表明，HEPN 作为一种有毒的 RNase，从一组 tRNA 的 3' 端切割掉 4 个核苷酸，从而干扰翻译。令人惊讶的是，我们发现 MNT（最小核苷酸转移酶）抗毒素通过保守酪氨酸残基 Y109 在活性位点环中的共价二腺苷酸化（二腺苷酰化）抑制 HEPN RNase。此外，我们呈现了不同阶段的二腺苷酸化反应的晶体快照，解释了 HEPN RNase 失活的机制。最后，我们提出 HEPN-MNT 系统作为一种细胞 ATP 传感器发挥作用，监测 ATP 动态平衡，并在低 ATP 水平下释放活性 HEPN 毒素。
