PerC manipulates metabolism and surface antigens in enteropathogenic Escherichia coli

Purpose: This study aimed to identify the genes regulated by plasmid-encoded regulator C (PerC). Whole transcriptomes of WT typical enteropathogenic E. coli (tEPEC) strains E2348/69 and coisogenic null-perC mutant JPEP22 were analyzed to identify and quantify differentially expressed genes. Methods: RNA was isolated from the WT and null-perC mutant strains and RNA integrity (RIN) was determined to be 10 out of possible 10 for all samples by Aligent Bioanalyzer. rRNA was depleted and resulting mRNA was reverse-transcribed into cDNA. Libraries were multiplexed for discrimination between libraries, barcoded for sequencing, and amplified with random hexamers for 15 PCR cycles. Transcripts were sequenced for 50 bases in single-end fashion within one lane of an Illumina Hiseq 2000 flow cell. This yielded roughly 30 million reads per sample. Results: Differential gene expression (DGE) analysis showed that 157 genes were statistically significantly regulated using a false-discovery rate (FDR) of 10% (q ≤ 0.10). Of these genes, the perC-mutant strain had far greater transcripts for the fim operon genes, fewer transcripts of nitrate reductase genes and anaerobic metabolism genes, and fewer transcripts for Hfq-dependent ncRNAs compared to WT. Conclusions: Differential transcript abundance between the perC-mutant and WT strains indicate PerC's negative regulation of the fim genes and positive regulation of anaerobic metabolism and ncRNA genes. Whole mRNA profiles of WT and coisogenic perC mutant EPEC strains were identified by isolation, mRNA enrichment, reverse-transcription, and Illumina sequencing