18 research outputs found
Connected network of the enriched differentially expressed genes following exposure to 0.2 mM H<sub>2</sub>O<sub>2</sub>.
<p>(A) Connected network of enriched differentially expressed genes involved in fatty acid metabolism (RM018 and RM020). (B) Partial fatty acid metabolism in <i>M</i>. <i>smegmatis</i>. Genes expressed differentially after 0.2 mM H<sub>2</sub>O<sub>2</sub> treatment assigned to RM018 and RM020 are marked in red.</p
Distinct Responses of <i>Mycobacterium smegmatis</i> to Exposure to Low and High Levels of Hydrogen Peroxide
<div><p>Hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) is a natural oxidant produced by aerobic organisms and gives rise to oxidative damage, including DNA mutations, protein inactivation and lipid damage. The genus <i>Mycobacterium</i> utilizes redox sensors and H<sub>2</sub>O<sub>2</sub> scavenging enzymes for the detoxification of H<sub>2</sub>O<sub>2</sub>. To date, the precise response to oxidative stress has not been fully elucidated. Here, we compared the effects of different levels of H<sub>2</sub>O<sub>2</sub> on transcription in <i>M</i>. <i>smegmatis</i> using RNA-sequencing. A 0.2 mM H<sub>2</sub>O<sub>2</sub> treatment had little effect on the growth and viability of <i>M</i>. <i>smegmatis</i> whereas 7 mM H<sub>2</sub>O<sub>2</sub> was lethal. Analysis of global transcription showed that 0.2 mM H<sub>2</sub>O<sub>2</sub> induced relatively few changes in gene expression, whereas a large proportion of the mycobacterial genome was found to be differentially expressed after treatment with 7 mM H<sub>2</sub>O<sub>2</sub>. Genes differentially expressed following treatment with 0.2 mM H<sub>2</sub>O<sub>2</sub> included those coding for proteins involved in glycolysis-gluconeogenesis and fatty acid metabolism pathways, and expression of most genes encoding ribosomal proteins was lower following treatment with 7 mM H<sub>2</sub>O<sub>2</sub>. Our analysis shows that <i>M</i>. <i>smegmatis</i> utilizes the sigma factor MSMEG_5214 in response to 0.2 mM H<sub>2</sub>O<sub>2</sub>, and the RpoE1 sigma factors MSMEG_0573 and MSMEG_0574 in response to 7 mM H<sub>2</sub>O<sub>2</sub>. In addition, different transcriptional regulators responded to different levels of H<sub>2</sub>O<sub>2</sub>: MSMEG_1919 was induced by 0.2 mM H<sub>2</sub>O<sub>2</sub>, while high-level induction of DevR occurred in response to 7 mM H<sub>2</sub>O<sub>2</sub>. We detected the induction of different detoxifying enzymes, including genes encoding KatG, AhpD, TrxB and Trx, at different levels of H<sub>2</sub>O<sub>2</sub> and the detoxifying enzymes were expressed at different levels of H<sub>2</sub>O<sub>2</sub>. In conclusion, our study reveals the changes in transcription that are induced in response to different levels of H<sub>2</sub>O<sub>2</sub> in <i>M</i>. <i>smegmatis</i>.</p></div
Overview of the differential expression profiles in response to 0.2 mM H<sub>2</sub>O<sub>2</sub> in <i>M</i>. <i>smegmatis</i>.
<p>(A) Enrichment analysis. The differently colored bars indicate the gene number for the enrichment of the annotations. (B) Interaction network of the differentially expressed genes of <i>M</i>. <i>smegmatis</i> induced by 0.2 mM H<sub>2</sub>O<sub>2</sub> using STRING (9.1) at confidence scores ≥ 0.4. The network is enriched among the 634 differentially expressed genes and 111 interactions were observed (p value = 0).</p
Quantitative RT-PCR validation of RNA-sequencing results.
<p>(A) Quantitative RT-PCR analysis of the mRNA expression of genes differentially expressed after treatment with different levels of H<sub>2</sub>O<sub>2</sub>. <i>M</i>. <i>smegmatis</i> cultures were treated with 2 mM or 7 mM H<sub>2</sub>O<sub>2</sub> for 30 min before extraction of RNA for qRT-PCR. The data represent 3 independent experiments. (B) Fold changes of selected genes differentially expressed genes after treatment with 0.2 mM and 7 mM H<sub>2</sub>O<sub>2</sub> obtained by the RNA-sequencing.</p
Fold changes of genes differentially expressed after treatment with 0.2 mM and 7 mM H<sub>2</sub>O<sub>2</sub> (treated vs untreated).
<p>Fold changes of genes differentially expressed after treatment with 0.2 mM and 7 mM H<sub>2</sub>O<sub>2</sub> (treated vs untreated).</p
Connected network of the enriched differentially expressed genes following exposure to 0.2 mM H<sub>2</sub>O<sub>2</sub>.
<p>(A) Connected network of enriched differentially expressed genes involved in fatty acid metabolism (RM018 and RM020). (B) Partial fatty acid metabolism in <i>M</i>. <i>smegmatis</i>. Genes expressed differentially after 0.2 mM H<sub>2</sub>O<sub>2</sub> treatment assigned to RM018 and RM020 are marked in red.</p
Schematic representation of the variable regions of class 1 integrons identified in <i>K. pneumoniae isolates</i>.
<p>Integron type with different cassette arrays (type I to X) are arranged as identified in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0075805#pone-0075805-t001" target="_blank">Table 1</a>. Gene cassettes are shown as boxes, with arrows indicating the orientation of transcription and hollow circles indicating the 59-base elements. The 5’- and 3’-conserved segments (5’-CS and 3’-CS) are annotated.</p
Comparison of the type IV integron in this study with its <i>E. coli</i> homologue in database.
<p>Sequence of <i>E</i>. <i>coli</i> strain DJ33-7 class I integron (JF806489) was retrieved from NCBI. The type IV integron (JQ823011) is arranged as indicated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0075805#pone-0075805-t001" target="_blank">Table 1</a>. Gene cassettes are shown as boxes, with arrows indicating the orientation of transcription. The 5’- and 3’-conserved segments (5’-CS and 3’-CS), attC, attI sites are annotated. Dashed box indicates a 590-bp deletion in the type IV integron.</p
Dissemination and Mechanism for the MCR-1 Colistin Resistance
<div><p>Polymyxins are the last line of defense against lethal infections caused by multidrug resistant Gram-negative pathogens. Very recently, the use of polymyxins has been greatly challenged by the emergence of the plasmid-borne mobile colistin resistance gene (<i>mcr-1</i>). However, the mechanistic aspects of the MCR-1 colistin resistance are still poorly understood. Here we report the comparative genomics of two new <i>mcr</i>-<i>1</i>-harbouring plasmids isolated from the human gut microbiota, highlighting the diversity in plasmid transfer of the <i>mcr</i>-<i>1</i> gene. Further genetic dissection delineated that both the trans-membrane region and a substrate-binding motif are required for the MCR-1-mediated colistin resistance. The soluble form of the membrane protein MCR-1 was successfully prepared and verified. Phylogenetic analyses revealed that MCR-1 is highly homologous to its counterpart PEA lipid A transferase in <i>Paenibacili</i>, a known producer of polymyxins. The fact that the plasmid-borne MCR-1 is placed in a subclade neighboring the chromosome-encoded colistin-resistant <i>Neisseria</i> LptA (EptA) potentially implies parallel evolutionary paths for the two genes. In conclusion, our finding provids a first glimpse of mechanism for the MCR-1-mediated colistin resistance.</p></div