Department of Microbiology, Institute of Plant and Microbial Biology, University of Zürich, Zürich, Switzerland.
J Bacteriol. 2021 May 20;203(12):e0068320. doi: 10.1128/JB.00683-20. Epub 2021 Mar 22.
Bacterial genomes can be methylated at particular motifs by methyltransferases (MTs). This DNA modification allows restriction endonucleases (REs) to discriminate between self and foreign DNA. While the accepted primary function of such restriction modification (RM) systems is to degrade incoming foreign DNA, other roles of RM systems and lone RE or MT components have been found in genome protection, stability, and the regulation of various phenotypes. The Burkholderia cepacia complex (Bcc) is a group of closely related opportunistic pathogens with biotechnological potential. Here, we constructed and analyzed mutants lacking various RM components in the clinical Bcc isolate Burkholderia cenocepacia H111 and used single-molecule, real-time (SMRT) sequencing of single mutants to assign the B. cenocepacia H111 MTs to their cognate motifs. DNA methylation is shown to affect biofilm formation, cell shape, motility, siderophore production, and membrane vesicle production. Moreover, DNA methylation had a large effect on the maintenance of the Bcc virulence megaplasmid pC3. Our data also suggest that the MT-encoding gene, which is essential in H111 and is located within a prophage, is required for maintaining the bacteriophage in a lysogenic state, thereby ensuring a constant, low level of phage production within the bacterial population. While the genome sequence determines an organism's proteins, methylation of the nucleotides themselves can confer additional properties. In bacteria, MTs modify specific nucleotide motifs to allow discrimination of "self" from "nonself" DNA, e.g., from bacteriophages. Restriction enzymes detect "nonself" methylation patterns and cut foreign DNA. Furthermore, methylation of promoter regions can influence gene expression and hence affect various phenotypes. In this study, we determined the methylated motifs of four strains from the Burkholderia cepacia complex of opportunistic pathogens. We deleted all genes encoding the restriction and modification components in one of these strains, Burkholderia cenocepacia H111. It is shown that DNA methylation affects various phenotypic traits, the most noteworthy being lysogenicity of a bacteriophage and maintenance of a virulence megaplasmid.
细菌基因组可以通过甲基转移酶（MTs）在特定基序上甲基化。这种 DNA 修饰允许限制性内切酶（REs）区分自身和外源 DNA。虽然这种限制修饰（RM）系统的公认主要功能是降解进入的外源 DNA，但在基因组保护、稳定性和各种表型的调控中，已经发现了 RM 系统和单独的 RE 或 MT 成分的其他作用。洋葱伯克霍尔德氏菌复合群（Bcc）是一组密切相关的机会性病原体，具有生物技术潜力。在这里，我们构建并分析了临床分离株伯克霍尔德氏菌 cenocepacia H111 中各种 RM 成分缺失的突变体，并使用单分子实时（SMRT）测序对单个突变体进行分析，以确定 B. cenocepacia H111 MT 与其同源基序的关系。结果表明，DNA 甲基化会影响生物膜形成、细胞形态、运动性、铁载体产生和膜囊泡产生。此外，DNA 甲基化对伯克霍尔德氏菌毒力大质粒 pC3 的维持有很大影响。我们的数据还表明，在 H111 中必不可少且位于一个前噬菌体内的 MT 编码基因对于维持噬菌体处于溶原状态是必需的，从而确保噬菌体在细菌群体中持续产生低水平的噬菌体。虽然基因组序列决定了生物体的蛋白质，但核苷酸本身的甲基化可以赋予其额外的特性。在细菌中，MT 修饰特定的核苷酸基序，以区分“自身”和“非自身” DNA，例如噬菌体。限制酶检测“非自身”甲基化模式并切割外源 DNA。此外，启动子区域的甲基化可以影响基因表达，从而影响各种表型。在这项研究中，我们确定了来自机会性病原体洋葱伯克霍尔德氏菌复合群的四个菌株的甲基化基序。我们删除了其中一个菌株，即伯克霍尔德氏菌 cenocepacia H111 中所有编码限制和修饰成分的基因。结果表明，DNA 甲基化会影响各种表型特征，最显著的是噬菌体的溶原性和毒力大质粒的维持。
