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Deciphering the response of Mycobacterium smegmatis to nitrogen stress using bipartite active modules.

ABSTRACT: BACKGROUND:The ability to adapt to environments with fluctuating nutrient availability is vital for bacterial survival. Although essential for growth, few nitrogen metabolism genes have been identified or fully characterised in mycobacteria and nitrogen stress survival mechanisms are unknown. RESULTS:A global transcriptional analysis of the mycobacterial response to nitrogen stress, showed a significant change in the differential expression of 16% of the Mycobacterium smegmatis genome. Gene expression changes were mapped onto the metabolic network using Active Modules for Bipartite Networks (AMBIENT) to identify metabolic pathways showing coordinated transcriptional responses to the stress. AMBIENT revealed several key features of the metabolic response not identified by KEGG enrichment alone. Down regulated reactions were associated with the general reduction in cellular metabolism as a consequence of reduced growth rate. Up-regulated modules highlighted metabolic changes in nitrogen assimilation and scavenging, as well as reactions involved in hydrogen peroxide metabolism, carbon scavenging and energy generation. CONCLUSIONS:Application of an Active Modules algorithm to transcriptomic data identified key metabolic reactions and pathways altered in response to nitrogen stress, which are central to survival under nitrogen limiting environments.

SUBMITTER: Williams KJ

PROVIDER: S-EPMC3706326 | biostudies-literature | 2013 Jul

REPOSITORIES: biostudies-literature

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Deciphering the response of Mycobacterium smegmatis to nitrogen stress using bipartite active modules.

Williams Kerstin J KJ Bryant William A WA Jenkins Victoria A VA Barton Geraint R GR Witney Adam A AA Pinney John W JW Robertson Brian D BD

BMC genomics 20130702

<h4>Background</h4>The ability to adapt to environments with fluctuating nutrient availability is vital for bacterial survival. Although essential for growth, few nitrogen metabolism genes have been identified or fully characterised in mycobacteria and nitrogen stress survival mechanisms are unknown.<h4>Results</h4>A global transcriptional analysis of the mycobacterial response to nitrogen stress, showed a significant change in the differential expression of 16% of the Mycobacterium smegmatis ge ...[more]

PMID: 23819599

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Project description:Nitrogen is essential for microbial growth and its importance is demonstrated by the complex regulatory systems used to control the transport, assimilation and metabolism of nitrogen. Recent studies are beginning to shed light on how mycobacteria respond to nitrogen limitation and several regulators (e.g., GlnR, PII) have been characterized at a molecular level. However, despite this progress, our knowledge of the transcriptional response of mycobacteria to nitrogen limitation and its regulation is confined to batch culture.To gain further insight into the response of mycobacteria to nitrogen limitation, we developed a nitrogen-limited chemostat. We compared the transcriptional response of nitrogen-limited cells to carbon-limited cells using RNA-seq analysis in a continuous culture model at a constant growth rate.Our findings revealed significant changes in the expression of 357 genes (208 upregulated, 149 downregulated; >2-fold change, false discovery rate <5 %) in response to nitrogen limitation in continuous culture. The vast majority of the GlnR regulon (68 %) was differentially expressed under nitrogen limitation in continuous culture and approximately 52 % of the 357 genes overlapped with a previously published study investigating the response of M. smegmatis to nitrogen limitation in batch culture, while expression of only 17 % of the genes identified in batch culture were affected in our chemostat model. Moreover, we identified a unique set of 45 genes involved in the uptake and metabolism of nitrogen that were exclusive to our chemostat model. We observed strong downregulation of pathways for amino acid catabolism (i.e., alanine, aspartate, valine, proline and lysine), suggesting preservation of these amino acids for critical cellular function. We found 16 novel transcriptional regulators that were directly or indirectly involved in the global transcriptomic response of M. smegmatis to nitrogen limitation and identified several non-coding RNAs that might be involved in the transcriptional or post-transcriptional regulation of nitrogen-regulated gene expression.Using nitrogen-limited continuous culture we identified the nitrogen-responsive transcriptome of M. smegmatis, including a number of small non-coding RNAs implicated in controlling nitrogen-regulated gene expression.

Project description:Stringent response is a conserved stress response mechanism in which bacteria employ the second messengers guanosine tetraphosphate and guanosine pentaphosphate [collectively termed (p)ppGpp] to reprogram their cellular processes under stress. In mycobacteria, these alarmones govern a multitude of cellular phenotypes, such as cell division, biofilm formation, antibiotic tolerance, and long-term survival. Mycobacterium smegmatis possesses the bifunctional RelMsm as a (p)ppGpp synthetase and hydrolase. In addition, it contains a short alarmone synthetase MS_RHII-RSD (renamed RelZ), which contains an RNase H domain in tandem with the (p)ppGpp synthetase domain. The physiological functions of RelMsm have been well documented, but there is no clear picture about the cellular functions of RelZ in M. smegmatis RelZ has been implicated in R-loop induced stress response due to its unique domain architecture. In this study, we elucidate the differential substrate utilization pattern of RelZ compared to that of RelMsm We unveil the ability of RelZ to use GMP as a substrate to synthesize pGpp, thereby expanding the repertoire of second messengers known in mycobacteria. We have demonstrated that the pGpp synthesis activity of RelZ is negatively regulated by RNA and pppGpp. Furthermore, we investigated its role in biofilm formation and antibiotic tolerance. Our findings highlight the complex role played by the RelZ in cellular physiology of M. smegmatis and sheds light upon its functions distinct from those of RelMsmIMPORTANCE Bacteria utilize nucleotide messengers to survive the hostile environmental conditions and the onslaught of attacks within the host. The second messengers guanosine tetraphosphate and pentaphosphate [(p)ppGpp] have a profound impact on the long-term survival, biofilm formation, antibiotic tolerance, virulence, and pathogenesis of bacteria. Therefore, understanding the stress response mechanism regulated by (p)ppGpp is essential for discovering inhibitors of stress response and potential drug targets. Mycobacterium smegmatis contains two (p)ppGpp synthetases: RelMsm and RelZ. Our study unravels the novel regulatory mechanisms of RelZ activity and its role in mediating antibiotic tolerance. We further reveal its ability to synthesize novel second messenger pGpp, which may have regulatory roles in mycobacteria.

Dataset Information

Deciphering the response of Mycobacterium smegmatis to nitrogen stress using bipartite active modules.

Publications

Deciphering the response of Mycobacterium smegmatis to nitrogen stress using bipartite active modules.

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