Basic info of AT-rich region containing strains.

Human pathogen Streptococcus pneumoniae forms multiple epigenetically and phenotypically distinct intra-populations by invertase PsrA-driven inversions of DNA methyltransferase hsdS genes in the colony opacity-determinant (cod) locus. As manifested by phase switch between opaque and transparent colonies, different genome methylation patterns or epigenomes confer pathogenesis-associated traits, but it is unknown how the pathogen controls the hsdS inversion orientations. Here, we report our finding of the SpxA1-TenA toxin-antitoxin (TA) system that regulates the orientations of hsdS inversions, and thereby bacterial epigenome and associated traits (e.g., colony opacity) by targeting pneumococcal protein synthesis. SpxA1 and TenA were found to constitute a highly conserved type II TA system in S. pneumoniae, primarily based on the observation that overexpressing toxin TenA led to growth arrest in E. coli and enhanced autolysis in S. pneumoniae, and the antitoxin SpxA1 repressed the transcription of the spxA1-tenA operon. When the transcription of tenA was de-repressed by a spontaneous AT di-nucleotide insertion/deletion in the promoter region of the spxA1-tenA operon, TenA bound to the ribosome maturation factor RimM, and thereby reduced the cellular level of alternative sigma factor ComX (known for the activation of natural transformation-associated genes). Attenuation of ComX expression in turn enhanced the transcription of the invertase gene psrA, which favored the formation of the transparent colony phase-associated hsdS allelic configurations in the cod locus. Phenotypically, moderate expression of TenA dramatically reshaped pneumococcal epigenome and colony opacity. Because spontaneous variations frequently occur during bacterial growth in the number of the AT di-nucleotides in the promoter region of the spxA1-tenA operon, this locus acts as a programmed genetic switch that generates pneumococcal subpopulations with epigenetic and phenotypic diversity.人类病原体肺炎链球菌（Streptococcus pneumoniae）可通过反转酶PsrA介导的菌落透明度决定位点（colony opacity-determinant, cod）内DNA甲基转移酶hsdS基因的反转，在种群内形成多种表观遗传与表型特征迥异的亚群。正如不透明菌落与透明菌落间的相转变所体现的那样，不同的基因组甲基化模式或表观基因组可赋予致病菌致病相关表型，但目前尚不清楚该病原体如何调控hsdS基因的反转方向。本研究报道了SpxA1-TenA毒素-抗毒素（toxin-antitoxin, TA）系统，该系统通过靶向肺炎链球菌的蛋白质合成过程，调控hsdS基因的反转方向，进而调控细菌的表观基因组及相关表型（如菌落透明度）。研究发现，SpxA1与TenA构成肺炎链球菌中高度保守的II型毒素-抗毒素系统，主要依据如下：过表达毒素TenA可导致大肠杆菌（E. coli）生长停滞，并增强肺炎链球菌的自溶活性；而抗毒素SpxA1可抑制spxA1-tenA操纵子的转录。当spxA1-tenA操纵子启动子区域发生自发性AT二核苷酸插入/缺失，解除了tenA基因的转录抑制后，TenA可结合核糖体成熟因子RimM，进而降低细胞内可变σ因子ComX的水平——ComX已知可激活自然转化相关基因。ComX表达的减弱可进一步上调反转酶基因psrA的转录，这有利于cod位点内与透明菌落相型相关的hsdS等位基因构型的形成。表型实验显示，TenA的适度表达可显著重塑肺炎链球菌的表观基因组与菌落透明度表型。由于在细菌生长过程中，spxA1-tenA操纵子启动子区域的AT二核苷酸数目常发生自发性变异，该位点可作为程序性遗传开关，产生具有表观遗传与表型多样性的肺炎链球菌亚群。