Laboratory of Pharmaceutical Microbiology, Ghent University, Ghent, Belgium.
Department of Plant and Microbial Microbiology, University of Zurich, Zurich, Switzerland.
mSphere. 2020 Jul 15;5(4):e00455-20. doi: 10.1128/mSphere.00455-20.
Respiratory tract infections by the opportunistic pathogen often lead to severe lung damage in cystic fibrosis (CF) patients. New insights in how to tackle these infections might emerge from the field of epigenetics, as DNA methylation is an important regulator of gene expression. The present study focused on two DNA methyltransferases (MTases) in strains J2315 and K56-2 and their role in regulating gene expression. predicted DNA MTase genes BCAL3494 and BCAM0992 were deleted in both strains, and the phenotypes of the resulting deletion mutants were studied: deletion mutant ΔBCAL3494 showed changes in biofilm structure and cell aggregation, while ΔBCAM0992 was less motile. wild-type cultures treated with sinefungin, a known DNA MTase inhibitor, exhibited the same phenotype as DNA MTase deletion mutants. Single-molecule real-time sequencing was used to characterize the methylome of , including methylation at the origin of replication, and motifs CACAG and GTWWAC were identified as targets of BCAL3494 and BCAM0992, respectively. All genes with methylated motifs in their putative promoter region were identified, and qPCR experiments showed an upregulation of several genes, including biofilm- and motility-related genes, in MTase deletion mutants with unmethylated motifs, explaining the observed phenotypes in these mutants. In summary, our data confirm that DNA methylation plays an important role in regulating the expression of genes involved in biofilm formation, cell aggregation, and motility. CF patients diagnosed with infections often experience rapid deterioration of lung function, known as cepacia syndrome. has a large multireplicon genome, and much remains to be learned about regulation of gene expression in this organism. From studies in other (model) organisms, it is known that epigenetic changes through DNA methylation play an important role in this regulation. The identification of genes of which the expression is regulated by DNA methylation and identification of the regulatory systems involved in this methylation are likely to advance the biological understanding of cell adaptation via epigenetic regulation. In time, this might lead to novel approaches to tackle infections in CF patients.
机会性病原体引起的呼吸道感染常导致囊性纤维化 (CF) 患者肺部严重损伤。DNA 甲基化是基因表达的重要调节因子，在表观遗传学领域，可能会出现新的方法来应对这些感染。本研究重点关注 J2315 和 K56-2 两种菌株中的两个 DNA 甲基转移酶 (MTase)及其在调节基因表达中的作用。在这两种菌株中，都删除了预测的 DNA MTase 基因 BCAL3494 和 BCAM0992，研究了由此产生的缺失突变体的表型：缺失突变体 ΔBCAL3494 表现出生物膜结构和细胞聚集的变化，而 ΔBCAM0992 的运动性降低。用已知的 DNA MTase 抑制剂 sinefungin 处理 野生型培养物表现出与 DNA MTase 缺失突变体相同的表型。单分子实时测序用于表征 的甲基组，包括复制起点的甲基化以及识别出 CACAG 和 GTWWAC 基序分别是 BCAL3494 和 BCAM0992 的靶标。在它们假定的启动子区域具有甲基化基序的所有基因都被鉴定出来，qPCR 实验表明，在 MTase 缺失突变体中，具有未甲基化基序的几个基因（包括与生物膜和运动性相关的基因）的表达上调，这解释了这些突变体中观察到的表型。总之，我们的数据证实 DNA 甲基化在调节与生物膜形成、细胞聚集和运动性相关的 基因表达中起着重要作用。被诊断患有 感染的 CF 患者经常经历肺功能迅速恶化，称为洋葱伯克霍尔德菌综合征。 具有大型多复制子基因组，关于该生物体中基因表达的调控仍有许多需要了解。从其他（模型）生物体的研究中可知，通过 DNA 甲基化的表观遗传变化在这种调控中起着重要作用。鉴定受 DNA 甲基化调控的 基因和鉴定参与这种甲基化的调控系统，可能会促进通过表观遗传调控来理解 细胞适应的生物学。随着时间的推移，这可能会导致针对 CF 患者中 感染的新方法。
