Department of Molecular, Cellular, and Developmental Biology, University of California at Santa Barbara, CA 93106, USA.
Genes Dev. 2012 Mar 1;26(5):515-25. doi: 10.1101/gad.182345.111. Epub 2012 Feb 14.
Bacterial contact-dependent growth inhibition (CDI) is mediated by the CdiB/CdiA family of two-partner secretion proteins. CdiA effector proteins are exported onto the surface of CDI(+) inhibitor cells, where they interact with susceptible bacteria and deliver effectors/toxins derived from their C-terminal regions (CdiA-CT). CDI(+) cells also produce an immunity protein that binds the CdiA-CT and blocks its activity to prevent autoinhibition. Here, we show that the CdiA-CT from uropathogenic Escherichia coli strain 536 (UPEC536) is a latent tRNase that requires activation by the biosynthetic enzyme CysK (O-acetylserine sulfhydrylase A). UPEC536 CdiA-CT exhibits no nuclease activity in vitro, but cleaves within transfer RNA (tRNA) anti-codon loops when purified CysK is added. CysK and CdiA-CT form a stable complex, and their binding interaction appears to mimic that of the CysK/CysE cysteine synthase complex. CdiA-CT activation is also required for growth inhibition. Synthesis of CdiA-CT in E. coli cysK(+) cells arrests cell growth, whereas the growth of ΔcysK mutants is unaffected by the toxin. Moreover, E. coli ΔcysK cells are completely resistant to inhibitor cells expressing UPEC536 CdiA, indicating that CysK is required to activate the tRNase during CDI. Thus, CysK acts as a permissive factor for CDI, providing a potential mechanism to modulate growth inhibition in target cells.
细菌接触依赖性生长抑制 (CDI) 是由双组分分泌蛋白家族 CdiB/CdiA 介导的。CdiA 效应蛋白被运送到 CDI(+)抑制剂细胞的表面，在那里它们与易感细菌相互作用，并传递来自其 C 末端区域 (CdiA-CT) 的效应物/毒素。CDI(+)细胞还产生一种免疫蛋白，该蛋白结合 CdiA-CT 并阻止其活性以防止自身抑制。在这里，我们表明来自尿路致病性大肠杆菌菌株 536 (UPEC536) 的 CdiA-CT 是一种潜伏的 tRNase，需要生物合成酶 CysK (O-乙酰丝氨酸硫基转移酶 A) 的激活。UPEC536 CdiA-CT 在体外没有核酸酶活性，但当加入纯化的 CysK 时，它会在转移 RNA (tRNA) 反密码环内切割。CysK 和 CdiA-CT 形成稳定的复合物，它们的结合相互作用似乎模拟了 CysK/CysE 半胱氨酸合酶复合物。CdiA-CT 的激活对于生长抑制也是必需的。在 cysK(+)细胞中合成 CdiA-CT 会导致细胞生长停滞，而毒素对 ΔcysK 突变体的生长没有影响。此外，大肠杆菌 ΔcysK 细胞对表达 UPEC536 CdiA 的抑制剂细胞完全具有抗性，这表明 CysK 在 CDI 过程中激活 tRNase 是必需的。因此，CysK 作为 CDI 的许可因子发挥作用，为调节靶细胞中的生长抑制提供了一种潜在的机制。
