Transcriptome analysis using RNA-sequencing (RNA-seq) has now become the standard approach to determine the transcriptional output of an organism. Various modifications to this technology have been developed over the years, usually aiming to improve the annotation of transcript borders, or to identify novel classes of RNAs, such as small regulatory RNAs (sRNAs) and antisense transcripts. RNA-seq has also led to the identification of dozens of new sRNAs in the major human pathogen, Vibrio cholerae. Several of these sRNAs function in the context of a cell-to-cell communication process, called quorum sensing (QS). QS is key for pathogenicity and biofilm formation of V. cholerae and the sRNAs involved typically act by base pairing with multiple target mRNAs to control gene expression at the posttranscriptional level. In this chapter, we describe the use of RNA-seq technologies for the discovery and characterization of regulatory RNAs in V. cholerae and discuss their relevance to QS and collective functions, such as biofilm formation. We further outline possible methods for the identification and validation of sRNA target genes, which can provide crucial information as to the physiological roles of an sRNA.
Keywords: Hfq; Quorum sensing; RNA-sequencing; Vibrio cholerae; sRNAs.
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