DNA Methyltransferase HsdM Induce Drug Resistance on Mycobacterium tuberculosis via Multiple Effects.

Hongqian Chu,Yongfei Hu,Bing Zhang,Baoli Zhu,Zhaogang Sun

doi:10.3390/antibiotics10121544

Abstract

Besides the genomic variants, epigenetic mechanisms such as DNA methylation also have an effect on drug resistance. This study aimed to investigate the methylomes of totally/extensively drug-resistant M. tuberculosis clinical isolates using the PacBio single-molecule real-time technology. The results showed they were almost the same as the pan-susceptible ones. Genetics and bioinformatics analysis confirmed three DNA methyltransferases—MamA, MamB, and HsdM. Moreover, anti-tuberculosis drug treatment did not change the methylomes. In addition, the knockout of the DNA methyltransferase hsdM gene in the extensively drug-resistant clinical isolate 11826 revealed that the motifs of GTAYN4ATC modified by HsdM were completely demethylated. Furthermore, the results of the methylated DNA target analysis found that HsdM was mainly involved in redox-related pathways, especially the prodrug isoniazid active protein KatG. HsdM also targeted three drug-targeted genes, eis, embB, and gyrA, and three drug transporters (Rv0194, Rv1410, and Rv1877), which mildly affected the drug susceptibility. The overexpression of HsdM in M. smegmatis increased the basal mutation rate. Our results suggested that DNA methyltransferase HsdM affected the drug resistance of M. tuberculosis by modulating the gene expression of redox, drug targets and transporters, and gene mutation.

Highlights

Tuberculosis (TB) is caused by the pathogen Mycobacterium tuberculosis (M. tuberculosis) and remains one of the leading causes of death caused by infectious pathogens, resulting in 10 million cases annually and latently infecting up to a third of the world’s population [1]
Compared with the sequencing reads obtained for the reference strain genome H37Rv (NC_000962) [13], 1650–1689 SNPs were found in clinical strains
Consistent with a previous study that showed that M. tuberculosis complex (MTBC) lineages displayed genomic diversity [14], we found genetic variations among clinical strains and identified multiple SNPs (Table S3)

Summary

Introduction

Tuberculosis (TB) is caused by the pathogen Mycobacterium tuberculosis (M. tuberculosis) and remains one of the leading causes of death caused by infectious pathogens, resulting in 10 million cases annually and latently infecting up to a third of the world’s population [1]. The results of gene mutations include compensatory evolution, epistasis, clonal interference, decreased cell wall permeability, overexpression of efflux pumps, drug/target modification, and target mimicry, which enhances M. tuberculosis by modulating their fitness, enhancing their transmissibility, and stabilizing the resistance phenotype within their population [4,5]. Epigenetic mechanisms such as DNA methylation affect gene expression. Studies based on whole-genome sequencing (WGS) of bacterial pathogens have provided novel insights into the evolution of M. tuberculosis drug resistance [6]. Despite many WGS studies, few reports exist about the epigenetic mechanisms of M. tuberculosis drug resistance. Several pieces of evidence have revealed N6-methyladenine (m6A) and 5-methylcytosine (m5C) methylation mechanisms within M. tuberculosis genomes, and three DNA MTases, MamA, MamB, and HsdM, are responsible for m6A modification [9,10,11]. A recent study revealed 12 M. tuberculosis complex (MTBC) methylomes and analyzed the methylation at the genome level [10]

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Methods

Results