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A MyD88-JAK1-STAT1 complex directly induces SOCS-1 expression in macrophages infected with Group A Streptococcus.

ABSTRACT: Some pathogens can use host suppressor of cytokine signaling 1 (SOCS-1), an important negative-feedback molecule, as the main mode of immune evasion. Here we found that group A Streptococcus (GAS) is capable of inducing SOCS-1 expression in RAW264.7 and BMDM macrophages. IFN-? plays a role in GAS-induced SOCS-1 expression in macrophages following the induction of cytokine expression by GAS, representing the classical pathway of SOCS-1 expression. However, GAS also induced STAT1 activation and SOCS-1 expression when GAS-infected cells were incubated with anti-IFN-? monoclonal antibody in this study. Moreover, upon comparing TLR4(-/-) BMDM macrophages with wild-type (WT) cells, we found that TLR4 also plays an essential role in the induction of SOCS-1. MyD88, which is an adaptor protein for TLR4, contributes to STAT1 activation and phosphorylation by forming a complex with Janus kinase 1 (JAK1) and signal transducer and activator of transcription 1 (STAT1) in macrophages. GAS-stimulated expression of STAT1 was severely impaired in MyD88(-/-) macrophages, whereas expression of JAK1 was unaffected, suggesting that MyD88 was involved in STAT1 expression and phosphorylation. Together, these data demonstrated that in addition to IFN-? signaling and MyD88 complex formation, JAK1 and STAT1 act in a novel pathway to directly induce SOCS-1 expression in GAS-infected macrophages, which may be more conducive to rapid bacterial infection.

SUBMITTER: Wu J

PROVIDER: S-EPMC4654310 | biostudies-literature | 2015 May

REPOSITORIES: biostudies-literature

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A MyD88-JAK1-STAT1 complex directly induces SOCS-1 expression in macrophages infected with Group A Streptococcus.

Wu Jinghua J Ma Cuiqing C Wang Haixin H Wu Shuhui S Xue Gao G Shi Xinli X Song Zhang Z Wei Lin L

Cellular & molecular immunology 20141117 3

Some pathogens can use host suppressor of cytokine signaling 1 (SOCS-1), an important negative-feedback molecule, as the main mode of immune evasion. Here we found that group A Streptococcus (GAS) is capable of inducing SOCS-1 expression in RAW264.7 and BMDM macrophages. IFN-β plays a role in GAS-induced SOCS-1 expression in macrophages following the induction of cytokine expression by GAS, representing the classical pathway of SOCS-1 expression. However, GAS also induced STAT1 activation and SO ...[more]

PMID: 25399770

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Project description:UnlabelledAs key components of innate immune defense, macrophages are essential in controlling bacterial pathogens, including group A Streptococcus(GAS). Despite this, only a limited number of studies have analyzed the recovery of GAS from within human neutrophils and macrophages. Here, we determined the intracellular fate of GAS in human macrophages by using several quantitative approaches. In both U937 and primary human macrophages, the appearance over time of long GAS chains revealed that despite GAS-mediated cytotoxicity, replication occurred in viable, propidium iodide-negative macrophages. Whereas the major virulence factor M1 did not contribute to bacterial growth, a GAS mutant strain deficient in streptolysin O (SLO) was impaired for intracellular replication. SLO promoted bacterial escape from the GAS-containing vacuole (GCV) into the macrophage cytosol. Up to half of the cytosolic GAS colocalized with ubiquitin and p62, suggesting that the bacteria were targeted by the autophagy machinery. Despite this, live imaging of U937 macrophages revealed proficient replication of GAS after GCV rupture, indicating that escape from the GCV is important for growth of GAS in macrophages. Our results reveal that GAS can replicate within viable human macrophages, with SLO promoting GCV escape and cytosolic growth, despite the recruitment of autophagy receptors to bacteria.ImportanceClassically regarded as an extracellular pathogen, GAS can persist within human epithelial cells, as well as neutrophils and macrophages. Some studies suggest that GAS can modulate its intracellular vacuole to promote survival and perhaps replicate in macrophages. However, an in-depth single-cell analysis of the dynamics of survival and replication is lacking. We used macrophage-like cell lines and primary macrophages to measure the intracellular growth of GAS at both the population and single-cell levels. While CFU counts revealed no increase in overall bacterial growth, quantitative fluorescence microscopy, flow cytometry, and time-lapse imaging revealed bacterial replication in a proportion of infected macrophages. This study emphasizes the importance of single-cell analysis especially when studying the intracellular fate of a pathogen that is cytotoxic and displays heterogeneity in terms of intracellular killing and growth. To our knowledge, this study provides the first direct visualization of GAS replication inside human cells.

Project description:Adaptation of group A Streptococcus (GAS) to its human host is mediated by two-component systems that transduce external stimuli to regulate bacterial physiology. Among such systems, CsrRS (also known as CovRS) is the most extensively characterized for its role in regulating ∼10% of the GAS genome, including several virulence genes. Here, we show that extracellular magnesium and the human antimicrobial peptide LL-37 have opposing effects on the phosphorylation of the response regulator CsrR by the receptor kinase CsrS. Genetic inactivation of CsrS phosphatase or kinase activity, respectively, had similar but more pronounced effects on CsrR phosphorylation compared to growth in magnesium or LL-37. These changes in CsrR phosphorylation were correlated with the repression or activation of CsrR-regulated genes as assessed by NanoString analysis. Chromatin immunoprecipitation and DNA sequencing (ChIP-seq) revealed CsrR occupancy at CsrRS-regulated promoters and lower-affinity associations at many other locations on the GAS chromosome. Because ChIP-seq did not detect CsrR occupancy at promoters associated with some CsrR-regulated genes, we investigated whether these genes might be controlled indirectly by intermediate regulators whose expression is modulated by CsrR. Transcriptional profiling of mutant strains deficient in the expression of either of two previously uncharacterized transcription regulators in the CsrR regulon indicated that one or both proteins participated in the regulation of 22 of the 42 CsrR-regulated promoters for which no CsrR association was detected by ChIP-seq. Taken together, these results illuminate CsrRS-mediated regulation of GAS gene expression through modulation of CsrR phosphorylation, CsrR association with regulated promoters, and the control of intermediate transcription regulators. IMPORTANCE Group A Streptococcus (GAS) is an important public health threat as a cause of sore throat, skin infections, life-threatening invasive infections, and the postinfectious complications of acute rheumatic fever, a leading cause of acquired heart disease. This work characterizes CsrRS, a GAS system for the detection of environmental signals that enables adaptation of the bacteria for survival in the human throat by regulating the production of products that allow the bacteria to resist clearance by the human immune system. CsrRS consists of two proteins: CsrS, which is on the bacterial surface to detect specific stimuli, and CsrR, which receives signals from CsrS and, in response, represses or activates the expression of genes coding for proteins that enhance bacterial survival. Some of the genes regulated by CsrR encode proteins that are themselves regulators of gene expression, thereby creating a regulatory cascade.

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A MyD88-JAK1-STAT1 complex directly induces SOCS-1 expression in macrophages infected with Group A Streptococcus.

Publications

A MyD88-JAK1-STAT1 complex directly induces SOCS-1 expression in macrophages infected with Group A Streptococcus.

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