The proline-rich antimicrobial peptide Api137 disrupts assembly of the large ribosomal subunit inducing immature dead-end particles
Ontology highlight
ABSTRACT:
ORGANISM(S): Escherichia Coli
SUBMITTER: 
Daniela Volke
PROVIDER: PXD051066 | panorama | Sun Jan 12 00:00:00 GMT 2025
REPOSITORIES: PanoramaPublic
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Project description:Ribosome profiling is a powerful method for globally assessing the activity of ribosomes in a cell. Despite its application in many organisms, ribosome profiling studies in bacteria have struggled to obtain the resolution necessary to precisely define translational pauses. Here we report improvements that yield much higher resolution in E. coli profiling data, enabling us to more accurately assess ribosome pausing and refine earlier studies of the impact of polyproline motifs on elongation. We comprehensively characterize pausing at proline-rich motifs in the absence of elongation factor EFP. We find that only a small fraction of genes with strong pausing motifs have reduced ribosome density downstream and identify features that explain this phenomenon. These features allow us to predict which proteins likely have reduced output in the efp knockout strain. Ribosome profiling of E. coli MG1655 and mutants lacking EFP or its three modifiying enzymes
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Project description:Purpose: We use the gene expression data to estimate the effects of tetracycline on gene expression and average ribosome density. Methods: The mRNAs were extracted with TRIzol reagent. The mRNAs were fragmented into 280 bp and the sequencing process was conducted on HiSeq 2500 platform. We use cutadapt, bowtie2, Plastid and DEseq2 software to analyze the expression levels of genes in two Escherichia coli strains. Results: The gene expression in EF4 knockout Escherichia coli strain was similar with BW25113 strain under normal condition. Under tetracycline treatment, many genes' expression were differentially regulated. Interestingly, we found that the gene expression of ribosomal proteins was up-regulated in WT strain comparing with EF4 knockout E. coli strain. Conclusions: Our results suggest that EF4 affects the average ribosome density and global gene expression in two Escherichia coli strain under tetracycline treatment.
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Project description:The current work aimed to detect and identify significant differentially expressed proteins between a virulent and an attenuated Histomonas meleagridis strain which were in vitro co-cultivated with Escherichia coli DH5alpha. This was done by applying 2D-DIGE and sequential window acquisition of all theoretical spectra (SWATH) MS as tools on the two well-defined strains. In the gel-based experiments, 49 identified protein spots were found to be differentially expressed, of which 37 belonged to the in vitro cultivated virulent strain and 12 to the attenuated one. The most frequently identified proteins in the virulent strain take part in cytoskeleton formation, carbohydrate metabolism and adaptation to stress. However, post-translationally modified or truncated ubiquitous cellular proteins such as actin and GAPDH were identified as upregulated in multiple gel positions. This indicated their contribution to processes not related to cytoskeleton and carbohydrate metabolism, such as fibronectin or plasminogen binding. On the other hand, proteins involved in cytoskeleton, cell division and cytoskeleton organization were frequently observed in the attenuated strain. The findings of the gel-based studies were supplemented by the gel-free SWATH MS analysis, which identified and quantified 42 significantly differentially regulated proteins. In this case proteins with peptidase activity, metabolic proteins and actin-regulating proteins were the most frequent findings in the virulent strain, while proteins involved in hydrogenosomal carbohydrate metabolism dominated the results in the attenuated one.
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Project description:The current work aimed to detect and identify significantly differentially expressed proteins between a virulent and an attenuated Histomonas meleagridis strain which were in vitro co-cultivated with Escherichia coli DH5alpha. This was done by applying 2D-DIGE and sequential window acquisition of all theoretical spectra (SWATH) MS as tools on the two well-defined strains. In the gel-based experiments, 49 identified protein spots were found to be differentially expressed, of which 37 belonged to the in vitro cultivated virulent strain and 12 to the attenuated one. The most frequently identified proteins in the virulent strain take part in cytoskeleton formation, carbohydrate metabolism and adaptation to stress. However, post-translationally modified or truncated ubiquitous cellular proteins such as actin and GAPDH were identified as upregulated in multiple gel positions. This indicated their contribution to processes not related to cytoskeleton and carbohydrate metabolism, such as fibronectin or plasminogen binding. On the other hand, proteins involved in cytoskeleton, cell division and cytoskeleton organization were frequently observed in the attenuated strain. The findings of the gel-based studies were supplemented by the gel-free SWATH MS analysis, which identified and quantified 42 significantly differentially regulated proteins. In this case proteins with peptidase activity, metabolic proteins and actin-regulating proteins were the most frequent findings in the virulent strain, while proteins involved in hydrogenosomal carbohydrate metabolism dominated the results in the attenuated one.
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Project description:The current work aimed to detect and identify significant differentially expressed proteins between a virulent and an attenuated Histomonas meleagridis strain which was in vitro co-cultivated with Escherichia coli DH5alpha. This was done by applying 2D-DIGE and sequential window acquisition of all theoretical spectra (SWATH) MS as tools on the two well-defined strains. In the gel-based experiments, 49 identified protein spots were found to be differentially expressed, of which 37 belonged to the in vitro cultivated virulent strain and 12 to the attenuated one. The most frequently identified proteins in the virulent strain take part in cytoskeleton formation, carbohydrate metabolism and adaptation to stress. However, post-translationally modified or truncated ubiquitous cellular proteins such as actin and GAPDH were identified as upregulated in multiple gel positions. This indicated their contribution to processes not related to cytoskeleton and carbohydrate metabolism, such as fibronectin or plasminogen binding. On the other hand, proteins involved in cytoskeleton, cell division and cytoskeleton organization were frequently observed in the attenuated strain. The findings of the gel-based studies were supplemented by the gel-free SWATH MS analysis, which identified and quantified 42 significantly differentially regulated proteins. In this case proteins with peptidase activity, metabolic proteins and actin-regulating proteins were the most frequent findings in the virulent strain, while proteins involved in hydrogenosomal carbohydrate metabolism dominated the results in the attenuated one.
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Project description:The current work aimed to detect and identify significantly differentially expressed proteins between a virulent and an attenuated Histomonas meleagridis strain which were in vitro co-cultivated with Escherichia coli DH5alpha. This was done by applying 2D-DIGE and sequential window acquisition of all theoretical spectra (SWATH) MS as tools on the two well-defined strains. In the gel-based experiments, 49 identified protein spots were found to be differentially expressed, of which 37 belonged to the in vitro cultivated virulent strain and 12 to the attenuated one. The most frequently identified proteins in the virulent strain take part in cytoskeleton formation, carbohydrate metabolism and adaptation to stress. However, post-translationally modified or truncated ubiquitous cellular proteins such as actin and GAPDH were identified as upregulated in multiple gel positions. This indicated their contribution to processes not related to cytoskeleton and carbohydrate metabolism, such as fibronectin or plasminogen binding. On the other hand, proteins involved in cytoskeleton, cell division and cytoskeleton organization were frequently observed in the attenuated strain. The findings of the gel-based studies were supplemented by the gel-free SWATH MS analysis, which identified and quantified 42 significantly differentially regulated proteins. In this case proteins with peptidase activity, metabolic proteins and actin-regulating proteins were the most frequent findings in the virulent strain, while proteins involved in hydrogenosomal carbohydrate metabolism dominated the results in the attenuated one.
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Project description:Ribosome profiling is a powerful method for globally assessing the activity of ribosomes in a cell. Despite its application in many organisms, ribosome profiling studies in bacteria have struggled to obtain the resolution necessary to precisely define translational pauses. Here we report improvements that yield much higher resolution in E. coli profiling data, enabling us to more accurately assess ribosome pausing and refine earlier studies of the impact of polyproline motifs on elongation. We comprehensively characterize pausing at proline-rich motifs in the absence of elongation factor EFP. We find that only a small fraction of genes with strong pausing motifs have reduced ribosome density downstream and identify features that explain this phenomenon. These features allow us to predict which proteins likely have reduced output in the efp knockout strain.
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