Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Newcastle University, Baddiley-Clark Building, Richardson Road, NE2 4AX Newcastle upon Tyne, UK.
Mol Cell. 2013 Oct 24;52(2):248-54. doi: 10.1016/j.molcel.2013.08.045. Epub 2013 Oct 3.
HipA of Escherichia coli is a eukaryote-like serine-threonine kinase that inhibits cell growth and induces persistence (multidrug tolerance). Previously, it was proposed that HipA inhibits cell growth by the phosphorylation of the essential translation factor EF-Tu. Here, we provide evidence that EF-Tu is not a target of HipA. Instead, a genetic screen reveals that the overexpression of glutamyl-tRNA synthetase (GltX) suppresses the toxicity of HipA. We show that HipA phosphorylates conserved Ser(239) near the active center of GltX and inhibits aminoacylation, a unique example of an aminoacyl-tRNA synthetase being inhibited by a toxin encoded by a toxin-antitoxin locus. HipA only phosphorylates tRNA(Glu)-bound GltX, which is consistent with the earlier finding that the regulatory motif containing Ser(239) changes configuration upon tRNA binding. These results indicate that HipA mediates persistence by the generation of "hungry" codons at the ribosomal A site that trigger the synthesis of (p)ppGpp, a hypothesis that we verify experimentally.
大肠杆菌的 HipA 是一种具有真核生物样丝氨酸-苏氨酸激酶活性的蛋白，它能抑制细胞生长并诱导持久性（多药耐受）。先前有研究表明 HipA 通过磷酸化必需的翻译因子 EF-Tu 来抑制细胞生长。在此，我们提供的证据表明 EF-Tu 不是 HipA 的靶标。相反，通过遗传筛选发现，谷氨酰-tRNA 合成酶（GltX）的过表达能抑制 HipA 的毒性。我们发现 HipA 磷酸化 GltX 活性中心附近保守的丝氨酸（Ser239），并抑制其氨酰化，这是毒素-抗毒素基因座编码的毒素抑制氨酰-tRNA 合成酶的独特例子。HipA 仅磷酸化与 tRNA(Glu)结合的 GltX，这与先前发现的含有 Ser239 的调节基序在与 tRNA 结合时改变构象的结论一致。这些结果表明，HipA 通过在核糖体 A 位产生“饥饿”密码子来介导持久性，从而触发（p）ppGpp 的合成，我们通过实验验证了这一假说。
