Uppsala University, Department of Medical Biochemistry and Microbiology, Biomedical Center, (Box 582), Uppsala S-75123, Sweden.
J Antimicrob Chemother. 2021 May 12;76(6):1441-1447. doi: 10.1093/jac/dkab056.
Ribosomal protection proteins (RPPs) interact with bacterial ribosomes to prevent inhibition of protein synthesis by tetracycline. RPP genes have evolved from a common ancestor into at least 12 distinct classes and spread by horizontal genetic transfer into a wide range of bacteria. Many bacterial genera host RPP genes from multiple classes but tet(M) is the predominant RPP gene found in Escherichia coli.
We asked whether phenotypic barriers (low-level resistance, high fitness cost) might constrain the fixation of other RPP genes in E. coli.
We expressed a diverse set of six different RPP genes in E. coli, including tet(M), and quantified tetracycline susceptibility and growth phenotypes as a function of expression level, and evolvability to overcome identified phenotypic barriers.
The genes tet(M) and tet(Q) conferred high-level tetracycline resistance without reducing fitness; tet(O) and tet(W) conferred high-level resistance but significantly reduced growth fitness; tetB(P) conferred low-level resistance and while mutants conferring high-level resistance were selectable these had reduced growth fitness; otr(A) did not confer resistance and resistant mutants could not be selected. Evolution experiments suggested that codon usage patterns in tet(O) and tet(W), and transcriptional silencing associated with nucleotide composition in tetB(P), accounted for the observed phenotypic barriers.
With the exception of tet(Q), the data reveal significant phenotypic and genetic barriers to the fixation of additional RPP genes in E. coli.
核糖体保护蛋白（RPP）与细菌核糖体相互作用，以防止四环素抑制蛋白质合成。RPP 基因从一个共同的祖先进化成至少 12 个不同的类别，并通过水平基因转移传播到广泛的细菌中。许多细菌属宿主具有来自多个类别的 RPP 基因，但 tet(M) 是大肠杆菌中发现的主要 RPP 基因。
我们想知道表型障碍（低水平抗性、高适应成本）是否会限制其他 RPP 基因在大肠杆菌中的固定。
我们在大肠杆菌中表达了一组不同的六种不同的 RPP 基因，包括 tet(M)，并根据表达水平量化了四环素的敏感性和生长表型，以及克服已识别的表型障碍的可进化性。
基因 tet(M) 和 tet(Q) 赋予了高水平的四环素抗性，而不会降低适应性；tet(O) 和 tet(W) 赋予了高水平的抗性，但显著降低了生长适应性；tetB(P) 赋予了低水平的抗性，虽然可以选择产生高水平抗性的突变体，但它们的生长适应性降低；otr(A) 不赋予抗性，并且不能选择抗性突变体。进化实验表明，tet(O) 和 tet(W) 中的密码子使用模式，以及与 tetB(P) 核苷酸组成相关的转录沉默，解释了观察到的表型障碍。
除了 tet(Q) 之外，数据显示在大肠杆菌中固定其他 RPP 基因存在显著的表型和遗传障碍。
