Dataset Information

Salmonella Skp mutant proteomics

ABSTRACT: Global discovery proteomic investigation of Salmonella typhimurium (control strain, Antibiotic resistant, or AcrB mutant) was performed in order to identify cellular processes affected by AcrB.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Bacteria Salmonella Typhimurium (strain 14028s / Sgsc 2262)

SUBMITTER: Alon Savidor

LAB HEAD: Yechiel Shai

PROVIDER: PXD014818 | Pride | 2020-05-27

REPOSITORIES: Pride

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Publications

Loss of the Periplasmic Chaperone Skp and Mutations in the Efflux Pump AcrAB-TolC Play a Role in Acquired Resistance to Antimicrobial Peptides in Salmonella typhimurium.

Kapach Gal G Nuri Reut R Schmidt Christiane C Danin Adi A Ferrera Shir S Savidor Alon A Gerlach Roman G RG Shai Yechiel Y

Frontiers in microbiology 20200310

Bacterial resistance to antibiotics is a major concern worldwide, leading to an extensive search for alternative drugs. Promising candidates are antimicrobial peptides (AMPs), innate immunity molecules, shown to be highly efficient against multidrug resistant bacteria. Therefore, it is essential to study bacterial resistance mechanisms against them. For that purpose, we used experimental evolution, and isolated a Salmonella enterica serovar typhimurium-resistant line to the AMP 4DK ...[more]

PMID: 32210923

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Project description:Here we focus on identifying conserved elements that are required for Chaserr function, a lncRNA that we recently characterized as being essential for mouse viability (https://doi.org/10.1038/s41467-019-13075-8). Chaserr is highly conserved in different species, and is located in close proximity (<2kb) to the transcription start site of CHD2. It acts in concert with the CHD2 protein to maintain proper Chd2 expression levels. Using the LncLOOM framework we identified AUGG enriched conserved elements which are located in the last exon of mouse Chaserr. In order to identify proteins that potentially bind to these conserved regions, we used in vitro transcription to generate the following biotinylated RNAs; along with their antisense controls: 1) The WT sequence, containing the first 322 nt of the last exon of Chaserr (bases 764–1086 of the NR_037601 isoform). An antisense control fragment was also prepared, labelled as AWT 2) The mutant conserved (MC) sequence, that contained AUGG→UACC mutations in four conserved motifs that were found using LncLOOM. An antisense control fragment was also prepared, labelled as AMC 3) The muatant all (MA) sequence in which all seven of the AUGG sites in the last exon were mutated to UACC. An antisense control fragment was also prepared, labelled as AMA 4) We also included a No Probe Control, which did not contain an RNA fragment. The fragments were incubated with N2a cell lysates followed by RNA-pulldown and Mass Spec analysis. The experiment was performed in triplicate. A total of 938 proteins were found to associate with the WT sequence: 74 of these were enriched ≥3-fold compared to the antisense sequence.We identified 9 proteins that had ≥2-fold higher recovery when using the WT sequence compared to both mutants. Included in these proteins was DHX36, which showed the highest enrichment compared to the mutated sequences. We validated the association of DHX36 with Chaserr through RNA immunoprecipitation experiments.

Dataset Information

Salmonella Skp mutant proteomics

Publications

Loss of the Periplasmic Chaperone Skp and Mutations in the Efflux Pump AcrAB-TolC Play a Role in Acquired Resistance to Antimicrobial Peptides in <i>Salmonella typhimurium</i>.

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