Targeted degradation of host messenger RNA by a bacterial pathogen [CLIP-seq]
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE281108 | GEO | 2024/11/10
REPOSITORIES: GEO
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Project description:Post-transcriptional processing and modification of messenger RNA (mRNA) regulates gene expression in eukaryotes by dictating the stability, localisation and translation of newly synthesised transcripts 1. These epitranscriptomic events are coordinated by an extensive network of RNA-binding proteins 2,3. Here we observed that the bacterial pathogen, Legionella pneumophila, caused the selective degradation of host cellular mRNAs encoding factors involved in glycolysis and related metabolic pathways. Screening of a library of L. pneumophila deletion mutant strains revealed a single effector protein, LegC4, that mediated the post-transcriptional degradation of host mRNAs encoding key glycolytic enzymes, thereby suppressing host glycolysis during infection. Using CLIP-seq and complementary methods, we observed that LegC4 bound to mature processed host mRNA recognising a guanine (G)-rich motif that was overrepresented within mRNAs targeted for degradation. In vitro activity assays showed that LegC4 harboured intrinsic RNase activity and structural determination of a catalytically inactive mutant of LegC4 in complex with single-stranded RNA revealed a unique RNA-binding domain 4. The selective binding and degradation of host mRNA by LegC4 reveals a previously undescribed mechanism of bacterial effector protein activity targeting the host epitranscriptome.
2024-11-09 | GSE280989 | GEO
Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis.
2011-01-08 | GSE26473 | GEO
Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis.
2011-01-08 | GSE26490 | GEO
Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis. Three-condition experiment: macrophages left uninfected (negative control), or infected with wildtype Legionella pneumophila, or the mutant Δ5, which lacks five bacterial effectors involved in inhibition of host protein synthesis (lgt1, lgt2, lgt3, sidI, sidL) (two experimental conditions). Biological replicates: two, independently infected, harvested, and hybridized to arrays. One technical replicate per array.
2011-01-08 | E-GEOD-26490 | biostudies-arrayexpress
Project description:Ribosomes are highly abundant cellular machines that perform the essential task of translating the genetic code into proteins. Cellular translation activity is finely tuned and proteostasis insults, such as those incurred upon viral infection, activate stress signaling pathways that result in translation reprogramming. Viral infection selectively shuts down host mRNA while redistributing ribosomes for selective translation of viral mRNAs. The intricacies of this selective ribosome shuffle from host to viral mRNAs are poorly understood. Here, we uncover a role for the ribosome associated quality control (RQC) factor ZNF598, a sensor for collided ribosomes, as a critical factor for vaccinia virus mRNA translation. Collided ribosomes are sensed by ZNF598, which ubiquitylates 40S subunit proteins uS10 and eS10 and thereby initiates RQC-dependent nascent chain degradation and ribosome recycling. We show that vaccinia infection in human cells enhances uS10 ubiquitylation indicating an increased burden on RQC pathways during viral propagation. Consistent with an increased RQC demand, we demonstrate that vaccinia virus replication is impaired in cells which either lack ZNF598 or contain a ubiquitylation deficient version of uS10. Using SILAC-based proteomics and concurrent RNAseq analysis, we determine that host translation of vaccinia virus mRNAs is compromised in cells that lack RQC activity as compared to control cells whereas there was little evidence of differences in host or viral transcription. Additionally, vaccinia virus infection resulted in a loss of cellular RQC activity, suggesting that ribosomes engaged in viral protein production recruit ZNF598 away from its function in host translation. Thus, co-option of ZNF598 by vaccinia virus plays a critical role in translational reprogramming that is needed for optimal viral propagation.
2021-01-05 | MSV000086660 | MassIVE
Project description:This SuperSeries is composed of the following subset Series: GSE26473: Secreted bacterial effectors that inhibit host protein synthesis are critical for induction of the innate immune response to virulent Legionella pneumophila [exp1] GSE26490: Secreted bacterial effectors that inhibit host protein synthesis are critical for induction of the innate immune response to virulent Legionella pneumophila [exp2] Refer to individual Series
2011-01-08 | E-GEOD-26491 | biostudies-arrayexpress
Project description:The intracellular bacterial pathogen Legionella pneumophila causes an inflammatory pneumonia called Legionnaires’ Disease. For virulence, L. pneumophila requires a Dot/Icm type IV secretion system that translocates bacterial effectors to the host cytosol. L. pneumophila lacking the Dot/Icm system is recognized by Toll-like receptors (TLRs), leading to a canonical NF-κB-dependent transcriptional response. In addition, L. pneumophila expressing a functional Dot/Icm system potently induces unique transcriptional targets, including proinflammatory genes such as Il23a and Csf2. Here we demonstrate that this Dot/Icm-dependent response, which we term the effector-triggered response (ETR), requires five translocated bacterial effectors that inhibit host protein synthesis. Upon infection of macrophages with virulent L. pneumophila, these five effectors caused a global decrease in host translation, thereby preventing synthesis of IκB, an inhibitor of the NF-κB transcription factor. Thus, macrophages infected with wildtype L. pneumophila exhibited prolonged activation of NF-κB, which was associated with transcription of ETR target genes such as Il23a and Csf2. L. pneumophila mutants lacking the five effectors still activated TLRs and NF-κB, but because the mutants permitted normal IκB synthesis, NF-κB activation was more transient and was not sufficient to fully induce the ETR. L. pneumophila mutants expressing enzymatically inactive effectors were also unable to fully induce the ETR, whereas multiple compounds or bacterial toxins that inhibit host protein synthesis via distinct mechanisms recapitulated the ETR when administered with TLR ligands. Previous studies have demonstrated that the host response to bacterial infection is induced primarily by specific microbial molecules that activate TLRs or cytosolic pattern recognition receptors. Our results add to this model by providing a striking illustration of how the host immune response to a virulent pathogen can also be shaped by pathogen-encoded activities, such as inhibition of host protein synthesis. Four-condition experiment: macrophages left uninfected (negative control), or infected with wildtype Legionella pneumophila, the flagellin-deficient mutant ΔflaA, or the secretion-deficient mutant ΔdotA (three experimental conditions). Biological replicates: two, independently infected, harvested, and hybridized to arrays. One to two technical replicates per array, as indicated in file titles.
2011-01-08 | E-GEOD-26473 | biostudies-arrayexpress
Project description:GAPDHs from human pathogens S. aureus and P. aeruginosa can be readily inhibited by ROS-mediated direct oxidation of their catalytic active cysteines. Because of the rapid degradation of H2O2 by bacterial catalase, only steady-state but not one-dose treatment of H2O2 induces rapid metabolic reroute from glycolysis to pentose phosphate pathway (PPP). We conducted RNA-seq analyses to globally profile the bacterial transcriptomes in response to a steady level of H2O2, which reveals profound transcriptional changes including the induced expression of glycolytic genes in both bacteria. Our results revealed that the inactivation of GAPDH by H2O2 induces a metabolic reroute from glycolysis to PPP; the elevated levels of fructose 1,6-biphosphate (FBP) and 2-keto-3-deoxy-6-phosphogluconate (KDPG) lead to dissociation of their corresponding glycolytic repressors (GapR and HexR, respectively) from their cognate promoters, thus resulting in derepression of the glycolytic genes to overcome H2O2-stalled glycolysis in S. aureus and P. aeruginosa, respectively. Given that H2O2 can be produced constitutively by the host immune response, exposure to the steady-state stress of H2O2 recapitulates more accurately bacterial responses to host immune system in vivo. RNA-seq in Pseudomonas aeruginosa and Staphylococus aureus under steady state of H2O2
2014-05-08 | E-GEOD-55528 | biostudies-arrayexpress
Project description:Abstract Legionella pneumophila, the causative agent of Legionnaire’s disease, grows within macrophages and manipulates target cell signaling. Formation of a Legionella-containing replication vacuole requires the function of the bacterial type IV secretion system (Dot/Icm), which transfers protein substrates into the host cell cytoplasm. A global microarray analysis was used to examine the response of human macrophage-like U937 cells to low dose infections with L. pneumophila. The most striking change in expression was the Dot/Icm-dependent up-regulation of anti-apoptotic genes positively controlled by the transcriptional regulator NF-?B. Consistent with this finding, L. pneumophila triggered nuclear localization of NF-?B in human and mouse macrophage in a Dot/Icm-dependent manner. The mechanism of activation at low dose infections involved a signaling pathway that occurred independently of the TLR adaptor MyD88, and cytoplasmic sensor Nod1. In contrast, high MOI conditions caused a host cell response that masked the unique Dot/Icm-dependent activation of NF-?B. Inhibition of NF-?B translocation into the nucleus resulted in premature host cell death and termination of bacterial replication. In the absence of one anti-apoptotic protein, PAI-2, host cell death increased in response to L. pneumophila infection, indicating that induction of anti-apoptotic genes is critical for host cell survival. Keywords: time course, dose response
2006-08-17 | GSE5551 | GEO
Project description:Bacterial competence is a genetically programmed process that is triggered as a response to environmental stimuli. DNA uptake from the extracellular milieu by competence requires the expression of a complex machinery composed of a competence pseudopilus and a DNA translocase. In L. pneumophila, it has been reported that microaerophilic conditions and genetic alteration of the competence repressors ComR and ProQ result in competence. We have analyzed the gene expression profiles of the proQ and comR mutants using the L. pneumophila Philadelphia-1 microarray. Many competence genes were found to be induced in the comR and proQ mutants and represent a defined, regulon. The gene expression profiling of the proQ and comR mutants was part of a study aimed at understanding the function of the processive 3'-5' exoribonuclease RNase R (rnr gene) in L. pneumophila. We found that the gene expression profile of the rnr mutant shares similarities with the gene expression profiles of the comR and proQ mutants and that loss of the exoribonuclease activity of RNase R was sufficient to induce competence development. RNase R is a processive 3'-5' exoribonuclease with a high degree of conservation in prokaryotes. Although some bacteria possess additional hydrolytic 3´-5-exoribonucleases such as RNase II, RNase R was found to be the only one predicted in the Legionella pneumophila genome. This provided a unique opportunity to study the role of RNase R in the absence of an additional RNase with similar enzymatic activity. We investigated the role of RNase R in the biology of Legionella pneumophila under various conditions and performed gene expression profiling using microarrays. At optimal growth temperature, loss of RNase R had no major consequence on bacterial growth and had a moderate impact on normal gene regulation. However, at lower temperatures loss of RNase R has a significant impact on bacterial growth and resulted in the accumulation of structured RNA degradation products. Concurrently, gene regulation was affected and specifically resulted in increased expression of the competence regulon. Loss of the exoribonuclease activity of RNase R was sufficient to induce competence development, a genetically-programmed process normally triggered as a response to environmental stimuli. Temperature-dependent expression of competence genes in the rnr mutant was found to be independent of the previously identified competence regulators comR and ProQ. comR: Three test samples and three reference samples were analyzed. The test sample being the strain KS79, a comR mutant of L. pneumophila strain JR32 which is used as a reference. proQ: Three test samples and three reference samples were analyzed. The test sample being the strain LELA3825, a proQ mutant of L. pneumophila strain JR32 which is used as a reference. rnr: Six test samples and six reference samples grown at 30°C or 37°C were analyzed. The test sample being the strain LELA559, a rnr mutant of L. pneumophila strain JR32 which is used as a reference.
2010-05-20 | E-GEOD-12796 | biostudies-arrayexpress