The small non-coding RNA RsaE influences extracellular matrix composition in Staphylococcus epidermidis biofilm communities

Abstract

RsaE is a conserved small regulatory RNA (sRNA) which was previously reported to represent a riboregulator of central carbon flow and other metabolic pathways in Staphylococcus aureus and Bacillus subtilis. Here we show that RsaE contributes to extracellular (e)DNA release and biofilm-matrix switching towards polysaccharide intercellular adhesin (PIA) production in a hypervariable Staphylococcus epidermidis isolate. Transcriptome analysis through differential RNA sequencing (dRNA-seq) in combination with confocal laser scanning microscopy (CLSM) and reporter gene fusions demonstrate that S. epidermidis protein- and PIA-biofilm matrix producers differ with respect to RsaE and metabolic gene expression. RsaE is spatiotemporally expressed within S. epidermidis PIA-mediated biofilms, and its overexpression triggers a PIA biofilm phenotype as well as eDNA release in an S. epidermidis protein biofilm matrix-producing strain background. dRNA-seq and Northern blot analyses revealed RsaE to exist as a major full-length 100-nt transcript and a minor processed species lacking approximately 20 nucleotides at the 5'-end. RsaE processing results in expansion of the mRNA target spectrum. Thus, full-length RsaE interacts with S. epidermidis antiholin-encoding lrgA mRNA, facilitating bacterial lysis and eDNA release. Processed RsaE, however, interacts with the 5'-UTR of icaR and sucCD mRNAs, encoding the icaADBC biofilm operon repressor IcaR and succinyl-CoA synthetase of the tricarboxylic acid (TCA) cycle, respectively. RsaE augments PIA-mediated biofilm matrix production, most likely through activation of icaADBC operon expression via repression of icaR as well as by TCA cycle inhibition and re-programming of staphylococcal central carbon metabolism towards PIA precursor synthesis. Additionally, RsaE supports biofilm formation by mediating the release of eDNA as stabilizing biofilm matrix component. As RsaE itself is heterogeneously expressed within biofilms, we consider this sRNA to function as a factor favoring phenotypic heterogeneity and supporting division of labor in S. epidermidis biofilm communities.

An RNA-centric global view ofClostridioides difficilereveals broad activity of Hfq in a clinically important Gram-positive bacterium

“…Premature transcription termination (PTT) within 5' untranslated regions were recently shown to be widespread events in the Gram-positive bacteria B. subtilis, Listeria monocytogenes and Enterococcus faecalis (Dar et al, 2016 (Bohn et al, 2010;Rochat et al, 2018) and also to promote biofilm formation in Staphylococcus epidermidis by supporting extracellular DNA release and the production of polysaccharideintercellular adhesin (PIA) (Schoenfelder et al, 2019). However, the function of Hfq in facilitating the interactions between sRNA regulators and their target mRNAs in Gram-positive bacteria remains elusive, since most of the known sRNAs seem to exert their regulatory activity independent of this global RNA-binding protein (Pitman and Cho, 2015).…”

A non‐coding RNA from the intercellular adhesion (ica) locus of Staphylococcus epidermidis controls polysaccharide intercellular adhesion (PIA)‐mediated biofilm formation

“…However, the promoter fusion experiments described above suggested primary transcription of icaZ rather than read‐through from the neighbouring gene. To further address this issue experimentally, we analysed an available RNA‐seq data set from ica ‐positive isolate S. epidermidis PS2 by comparing TEX (Terminator™ 5 ʹ ‐Phosphate‐Dependent Exonuclease) treated and untreated RNA samples (Schoenfelder et al , ). TEX is an exonuclease that degrades processed RNAs carrying a 5 ʹ monophosphate, while primary transcripts with a 5 ʹ triphosphate remain intact (Sharma et al , ).…”

Phenotypes of Streptococcus mutans isolates in sound and cavitated tooth surfaces and its relationship to early childhood caries in China

“…Importantly, sRNAs coordinate adaptation processes in response to environmental changes in bacteria by regulating target mRNA expression [37][38][39][40][41]. Specifically, srn225147 and srn821798 were sequenced in our previous studies [22,23].…”

“…The biofilm assay was performed according to Zhou et al, (2018) with a minor adjustment [41]. Twohundred μL of BHI broth of each isolate with 1 × 10 7 CFU/mL S. mutans and 1% sucrose were added in 96-well plates.…”