Institute of Medical Microbiology and Hospital Epidemiology, Hannover Medical School, Carl-Neuberg-Straße 1, 30625 Hannover, Germany, German Center for Infection Research, Hannover-Braunschweig Site, Carl-Neuberg-Straße 1, 30625 Hannover, Germany, New England Biolabs, 240 County Road, Ipswich, MA 01938, USA, Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures, Inhoffenstraße 7B, 38124 Braunschweig, Germany and Pacific Biosciences, 1380 Willow Road, Menlo Park, CA 94025, USA.
Nucleic Acids Res. 2014 Feb;42(4):2415-32. doi: 10.1093/nar/gkt1201. Epub 2013 Dec 2.
The genome of Helicobacter pylori is remarkable for its large number of restriction-modification (R-M) systems, and strain-specific diversity in R-M systems has been suggested to limit natural transformation, the major driving force of genetic diversification in H. pylori. We have determined the comprehensive methylomes of two H. pylori strains at single base resolution, using Single Molecule Real-Time (SMRT®) sequencing. For strains 26695 and J99-R3, 17 and 22 methylated sequence motifs were identified, respectively. For most motifs, almost all sites occurring in the genome were detected as methylated. Twelve novel methylation patterns corresponding to nine recognition sequences were detected (26695, 3; J99-R3, 6). Functional inactivation, correction of frameshifts as well as cloning and expression of candidate methyltransferases (MTases) permitted not only the functional characterization of multiple, yet undescribed, MTases, but also revealed novel features of both Type I and Type II R-M systems, including frameshift-mediated changes of sequence specificity and the interaction of one MTase with two alternative specificity subunits resulting in different methylation patterns. The methylomes of these well-characterized H. pylori strains will provide a valuable resource for future studies investigating the role of H. pylori R-M systems in limiting transformation as well as in gene regulation and host interaction.
幽门螺杆菌的基因组以其大量的限制修饰（R-M）系统为特征，并且已经表明 R-M 系统在菌株特异性上的多样性限制了自然转化，这是幽门螺杆菌遗传多样化的主要驱动力。我们使用单分子实时（SMRT®）测序技术，以单碱基分辨率确定了两种幽门螺杆菌菌株的综合甲基组。对于菌株 26695 和 J99-R3，分别鉴定出了 17 个和 22 个甲基化序列基序。对于大多数基序，在基因组中发生的几乎所有位点都被检测为甲基化。检测到了 12 种新的甲基化模式，对应于 9 个识别序列（26695，3；J99-R3，6）。功能失活、纠正移码以及候选甲基转移酶（MTase）的克隆和表达不仅允许对多个尚未描述的 MTase 进行功能表征，而且还揭示了 I 型和 II 型 R-M 系统的新特征，包括移码介导的序列特异性变化以及一个 MTase 与两个替代特异性亚基的相互作用，导致不同的甲基化模式。这些经过良好表征的幽门螺杆菌菌株的甲基组将为未来研究幽门螺杆菌 R-M 系统在限制转化以及基因调控和宿主相互作用中的作用提供有价值的资源。
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