Project description:We developed a surface-displayed library of 117 human antimicrobial peptides (AMPs) and over 3000 human AMP variants in a laboratory E. coli strain expressing PhoP and PhoQ from Salmonella Typhimurium. We used sort-seq (fluorescence-activated cell sorting followed by next-generation sequencing) to characterize PhoPQ activation by these surface-displayed AMPs.
Project description:Transfer RNAs (tRNAs) maintain translational fidelity through strict charging by their cognate aminoacyl-tRNA synthetase and codon:anticodon base pairing with the mRNA at the ribosome. Mistranslation occurs when an amino acid not specified by the genetic code is incorporated into a protein. Since alanyl-tRNA synthetase uniquely recognizes a G3:U70 base pair in alanine tRNAs and the anticodon plays no role in charging, alanine tRNA variants with anticodon mutations have the potential to mistranslate alanine. Our goal was to quantify mis-incorporation of alanine into proteins in Saccharomyces cerevisiae strains expressing one of 57 different alanine tRNA anticodon variants. Using mass spectrometry, we observed mistranslation for 45 of the variants when expressed on single-copy plasmids.
Project description:Salmonella causes a range of diseases in different hosts, including enterocolitis and systemic infection. Lysine acetylation regulates many eukaryotic cellular processes, but its function in prokaryotes is largely unknown. Reversible acetylation in Salmonella Typhimurium depends on acetyltransferase Pat and nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase CobB. Here, we used cell and animal models to evaluate the virulence of pat and cobB deletion mutants in S. Typhimurium, and found that pat is essential for bacterial intestinal colonization, systemic infection and host inflammation response. Next, to understand the underlying mechanism, genome-wide transcriptome was analyzed. RNA-seq data showed the expression of Salmonella pathogenicity islands 1 (SPI-1) is partially dependent on pat. In addition, we found that HilD is a substrate of Pat, which is essential for maintaining HilD protein level. Taken together, these results suggested that protein acetylation system regulates SPI-1 expression by controlling HilD in a post-translational manner to mediate S. Typhimurium virulence.
Project description:Salmonella causes a range of diseases in different hosts, including enterocolitis and systemic infection. Lysine acetylation regulates many eukaryotic cellular processes, but its function in prokaryotes is largely unknown. Reversible acetylation in Salmonella Typhimurium depends on acetyltransferase Pat and nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase CobB. Here, we used cell and animal models to evaluate the virulence of pat and cobB deletion mutants in S. Typhimurium, and found that pat is essential for bacterial intestinal colonization, systemic infection and host inflammation response. Next, to understand the underlying mechanism, genome-wide transcriptome was analyzed. RNA-seq data showed the expression of Salmonella pathogenicity islands 1 (SPI-1) is partially dependent on pat. In addition, we found that HilD is a substrate of Pat, which is essential for maintaining HilD protein level. Taken together, these results suggested that protein acetylation system regulates SPI-1 expression by controlling HilD in a post-translational manner to mediate S. Typhimurium virulence. To use RNA-seq to analyze the transcriptome patterns of pat or cobB mutation in Salmonella Typhimurium 14028s.
Project description:The GeneChip Porcine Genome Array was used to identify the transcriptional response upon either Salmonella typhimurium (ST) or Salmonella choleraesuis (SC) infection in two porcine epithelial cell lines (IPEC-J2, from jejunum and IPI-2I, from ileum) during 2 and 4 hours post infection. The objectives in this study were first, to identify the different response between the epithelial cell lines from different gut regions; second, to study how the Salmonella serotypes used could elicit a different host response; and third, to determine the effect of the time-points on the differentially gene expression.
Project description:The GeneChip Porcine Genome Array was used to identify the transcriptional response upon either Salmonella typhimurium (ST) or Salmonella choleraesuis (SC) infection in two porcine epithelial cell lines (IPEC-J2, from jejunum and IPI-2I, from ileum) during 2 and 4 hours post infection. The objectives in this study were first, to identify the different response between the epithelial cell lines from different gut regions; second, to study how the Salmonella serotypes used could elicit a different host response; and third, to determine the effect of the time-points on the differentially gene expression.
Project description:Salmonella enterica is one of the most important foodborne pathogens that infect a variety of animals and birds. In humans, S. Typhimurium causes gastroenteritis, leading to vomiting, diarrhea, fever, and abdominal cramps. We mainly get infected with Salmonella by ingesting comminated poultry products. Therefore, developing an oral live attenuated vaccine for the poultry industry is our best bet against Salmonella infection. In this article, we investigated the potential of the next generation of Salmonella vaccines. We generated a library of potentially attenuated S. Typhimurium mutants and compared fitness to that of a commercial vaccine. We also investigated the invasion and survival potential of these mutants in chicken macrophages. Our data indicate that although these mutants had no significant growth defects, they were much sensitive to macrophage attack. Analyzing the transcriptome data from infected primary chicken macrophages, we concluded that these mutants elicit a robust immune response by activating several immunoregulatory pathways. Our data also indicates that by combining phoPQ deletion with an already existing cya-crp deletion in MeganVac1, a much stronger immune response can be generated.