Transcriptional Profiling of Mycobacterium tuberculosis Exposed to In Vitro Lysosomal Stress.
Increasing experimental evidence supports the idea that Mycobacterium tuberculosis has evolved strategies to survive within lysosomes of activated macrophages. To further our knowledge of M. tuberculosis response to the hostile lysosomal environment, we profiled the global transcriptional activity of M. tuberculosis when exposed to the lysosomal soluble fraction (SF) prepared from activated macrophages. Transcriptome sequencing (RNA-seq) analysis was performed using various incubation conditions, ranging from noninhibitory to cidal based on the mycobacterial replication or killing profile. Under inhibitory conditions that led to the absence of apparent mycobacterial replication, M. tuberculosis expressed a unique transcriptome with modulation of genes involved in general stress response, metabolic reprogramming, respiration, oxidative stress, dormancy response, and virulence. The transcription pattern also indicates characteristic cell wall remodeling with the possible outcomes of increased infectivity, intrinsic resistance to antibiotics, and subversion of the host immune system. Among the lysosome-specific responses, we identified the glgE-mediated 1,4 α-glucan synthesis pathway and a defined group of VapBC toxin/anti-toxin systems, both of which represent toxicity mechanisms that potentially can be exploited for killing intracellular mycobacteria. A meta-analysis including previously reported transcriptomic studies in macrophage infection and in vitro stress models was conducted to identify overlapping and nonoverlapping pathways. Finally, the Tap efflux pump-encoding gene Rv1258c was selected for validation. An M. tuberculosis ΔRv1258c mutant was constructed and displayed increased susceptibility to killing by lysosomal SF and the antimicrobial peptide LL-37, as well as attenuated survival in primary murine macrophages and human macrophage cell line THP-1.
Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore Infectious Disease Interdisciplinary Research Group, Singapore-MIT Alliance for Research and Technology, Singapore.
Wenwei
W
Lin
Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore.
Paola Florez
PF
de Sessions
Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore.
Garrett Hor Keong
GH
Teoh
Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore.
Ahmad Naim Nazri
AN
Mohamed
Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore.
Yuan O
YO
Zhu
Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore.
Vanessa Hui Qi
VH
Koh
Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore.
Michelle Lay Teng
ML
Ang
Infectious Disease Interdisciplinary Research Group, Singapore-MIT Alliance for Research and Technology, Singapore.
Peter C
PC
Dedon
Genome Institute of Singapore, Agency for Science, Technology and Research, Singapore Department of Pathogen Molecular Biology, London School of Hygiene & Tropical Medicine, London, United Kingdom.
Martin Lloyd
ML
Hibberd
Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore Infectious Disease Interdisciplinary Research Group, Singapore-MIT Alliance for Research and Technology, Singapore micas@nus.edu.sg.
Sylvie
S
Alonso
D000818:Animals; D023181:Antimicrobial Cationic Peptides; D054804:Cathelicidins / Q000235:genetics; D002460:Cell Line; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D054884:Host-Pathogen Interactions / Q000235:genetics; D006801:Humans; D008247:Lysosomes / Q000235:genetics*; D008264:Macrophages / Q000382:microbiology; D051379:Mice; D008807:Mice, Inbred BALB C; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D018384:Oxidative Stress / Q000235:genetics*; D014158:Transcription, Genetic / Q000235:genetics*; D059467:Transcriptome / Q000235:genetics; D014376:Tuberculosis / Q000382:microbiology; D014774:Virulence / Q000235:genetics
D023181:Antimicrobial Cationic Peptides; D054804:Cathelicidins