Project description:Wild-type (JRS4) and irr mutant (JRS550) Group A Streptococcus (GAS) strains growing in either early or late exponential phase Keywords = Human Keywords = Neutrophils Keywords = Bacterial Keywords = Gene Regulation Keywords = Inflammation Keywords: other
Project description:Group A Streptococcus (GAS, aka Streptococcus pyogenes) causes an array of human diseases from mild pharyngitis to life-threatening necrotizing fasciitis. Invading host cells is a strategy for GAS to avoid antibiotic killing and immune system clearance. Our previous study showed that GAS is able to multiply in human microvascular endothelial cell line-1 (HMEC-1), and has higher survival in HMEC-1 cells due to insufficient acidification in lysosomes. GAS peroxide response regulator (PerR) modulates not only peroxide stress, but also metal homeostasis and bacterial virulence. Therefore, we aimed to investigate the role of PerR during GAS invading endothelial cells. First, we found that ΔperR mutant was more tolerant to H2O2 in vitro using hydrogen peroxide sensitivity assay. We further used cDNA-qPCR analysis to clarify the resistance mechanism of ΔperR mutant. The gene expressions of dpr, ahpC, and ahpF were up-regulated, explaining the enhanced resistance of the ΔperR mutant against peroxide stress. However, the proliferation of the ΔperR mutant in HMEC-1 cells was significantly lower than wild type strain after 5 h post-infection. To explore the underlying mechanisms of ΔperR mutant during infection, we performed dual RNAseq analysis to identify differentially expressed genes, and validated them using cDNA-qPCR analysis. The resulting up-regulated genes of iron efflux pump pmtA, iron/zinc chelating protein dpr, and zinc acquisition system (adcA, lmb/adcAII, phtD), and down-regulated zinc efflux pump czcD gave rise to impaired metal homeostasis in the ΔperR mutant. Furthermore, in vitro growth curve assays showed that the ΔperR mutant was sensitive to zinc deficiency and resistant to zinc toxicity. Taken together, this study has demonstrated the critical roles that GAS PerR plays in protecting from peroxide stress of host innate immune responses and zinc sequestration of nutritional immunity. Our novel findings open a new avenue of strategies in the development of antimicrobial agents. Our study demonstrates the importance of PerR. It aids GAS virulence in immune evasion during the LC3-associated phagocytosis in endothelial cells.
Project description:This transcriptional analysis is a follow up to a population genomic investigation of 3615 Streptococcus pyogenes serotype M1 strains whch are responsible for an epidemic of human invasive infections (www.pnas.org/cgi/doi/10.1073/pnas.1403138111), The goal was to assess gene expression differences between predecessor pre-epidemic M1 strains and their descendent epidemic M1 strains to gain insights into the underlying genetic basis for the shift in the frequency and severity of human infections caused by these pathogenic bacteria The transcriptomes of 7 GAS M1 strains, 4 pre-epidemic and 3 epidemic, were compared at two phases of growth, mid-exponential and early-stationary, using 3 biologial replicates, to identify genes differentially expressed between the pre-epidemic and epidemic isolates with the goal of to gaining insight into the underlying genetic basis for the evolutionary emergence, increased frequency and severity of the epidemic strains relative to the pre-epidemic strains
Project description:Transcriptional profiling of Streptococcus pyogenes MGAS5005 cells comparing control untreated GAS cells with GAS cells exposed to 4uM heme for 1.5 h
Project description:The nasopharynx and the skin are the major oxygen-rich anatomical sites for colonization by the human pathogen Streptococcus pyogenes (group A Streptococcus, GAS). To establish infection, GAS must survive oxidative stress generated during aerobic metabolism and the release of reactive oxygen species (ROS) by host innate immune cells. Glutathione is the major host antioxidant molecule while GAS is glutathione-auxotrophic. Here we report the molecular characterization of the ABC transporter substrate binding protein GshT in the GAS glutathione salvage pathway. We demonstrate that glutathione uptake is critical for aerobic growth of GAS and that impaired import of glutathione induces oxidative stress that triggers enhanced production of the reducing equivalent NADPH. Our results highlight the interrelationship between glutathione assimilation, carbohydrate metabolism, virulence factor production and innate immune evasion. Together, these findings suggest an adaptive strategy employed by extracellular bacterial pathogens to exploit host glutathione stores for their own benefit.
