The Mycobacterial DNA Methyltransferase HsdM Decreases Intrinsic Isoniazid Susceptibility

Abstract

Tuberculosis, caused by the pathogen Mycobacterium tuberculosis, is a serious infectious disease worldwide. Multidrug-resistant TB (MDR-TB) remains a global problem, and the understanding of this resistance is incomplete. Studies suggested that DNA methylation promotes bacterial adaptability to antibiotic treatment, but the role of mycobacterial HsdM in drug susceptibility has not been explored. Here, we constructed an inactivated Mycobacterium bovis (BCG) strain, ΔhsdM. ΔhsdM shows growth advantages over wild-type BCG under isoniazid treatment and hypoxia-induced stress. Using high-precision PacBio single-molecule real-time sequencing to compare the ΔhsdM and BCG methylomes, we identified 219 methylated HsdM substrates. Bioinformatics analysis showed that most HsdM-modified genes were enriched in respiration- and energy-related pathways. qPCR showed that HsdM-modified genes directly affected their own transcription, indicating an altered redox regulation. The use of the latent Wayne model revealed that ΔhsdM had growth advantages over wild-type BCG and that HsdM regulated trcR mRNA levels, which may be crucial in regulating transition from latency to reactivation. We found that HsdM regulated corresponding transcription levels via gene methylation; thus, altering the mycobacterial redox status and decreasing the bacterial susceptibility to isoniazid, which is closely correlated with the redox status. Our results provide valuable insight into DNA methylation on drug susceptibility.

Experimental Evolution Reveals Redox State Modulates Mycobacterial Pathogenicity

“…In a previous study, the mutated mycobacterial strain, mc 2 51, evolved into an M. tuberculosis -like strain that presented slow growth and improved growth fitness under stress ( Li et al, 2014b ), which H 2 O 2 resistance in mycobacteria linked to virulence. Previous studies including ours have associated isoniazid (INH) with H 2 O 2 resistance ( Timmins and Deretic, 2006 ; Hu et al, 2021 ). The MIC of isoniazid (INH) for mc 2 51 was dramatically reduced to ∼1% of the MIC for mc 2 155 ( Table 1 ).…”

“…Clinical investigation has shown that cells from TB patients produce less ROS than those of healthy individuals ( Jaswal et al, 1992 ; Kumar et al, 1995 ). In addition, ROS can also be associated with the treatment of TB ( Piccaro et al, 2014 ; Hu et al, 2021 ). To avoid ROS attack, an evolved detoxified system is essential for M. tuberculosis survival, persistence, and subsequent reactivation ( Piccaro et al, 2014 ).…”

Mycobacterium tuberculosis Methyltransferase Rv1515c Can Suppress Host Defense Mechanisms by Modulating Immune Functions Utilizing a Multipronged Mechanism

“…MamA, MamB, and HsdM are three known DNA MTases coded by M. tb, which target different sequence pattern for N6-adenine methylation (Shell et al, 2013;Zhu et al, 2016). Mycobacterial DNA methyltransferase HSdM regulates transcription and decreases susceptibility to isoniazid (Hu et al, 2021). Rv1988 is a secretory methyltransferase that enters the nucleus of the host and methylates histone H3 at arginine residues, regulating the expression of genes that neutralise reactive oxygen species (Yaseen et al, 2015).…”

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

“…The number of hsdM-methylated motifs is almost double in BCG (674) than in clinical XDR-TB isolates 11826 (368). In comparison to their parent strain, the knockout of hsdM in BCG did not reveal a growth difference, while the knockout of hsdM in XDR-TB 11826 showed a drop in growth [ 21 ]. There were also some reports on the HsdM homolog in other bacterial species, such as in streptococci (e.g., S. pyogenes and S. agalactiae), which might involve bacterial drug resistance and other physiological features such as growth, hypoxia, and UV stress [ 22 , 23 ].…”