A bacterial regulatory RNA attenuates virulence, spread and human host cell phagocytosis

Abstract

Staphylococcus aureus pathogenesis is directed by regulatory proteins and RNAs. We report the case of an RNA attenuating virulence and host uptake, possibly to sustain commensalism. A S. aureus sRNA, SprC (srn_3610), reduced virulence and bacterial loads in a mouse infection model. S. aureus deleted for sprC became more virulent and increased bacterial dissemination in colonized animals. Conversely, inducing SprC expression lowered virulence and the bacterial load. Without sprC, S. aureus phagocytosis by monocytes and macrophages was higher, whereas bacteria were internalized at lower yields when SprC expression was stimulated. Without sprC, higher internalization led to a greater number of extracellular bacteria, facilitating colonization. SprC expression decreased after phagocytosis, concurring with the facilitated growth of bacteria lacking the sRNA in the presence of an oxidant. The major staphylococcal autolysin facilitates S. aureus uptake by human phagocytes. ATL proved to be negatively regulated by SprC. The SprC domains involved in pairing with atl mRNA were analyzed. The addition of ATL reduced phagocytosis of bacteria lacking sprC with no effects on wild-type bacterial uptake, implying that SprC influences phagocytosis, at least in part, by controlling ATL. Since the control of SprC on ATL was modest, other factors must contribute to atl regulation.

Insights into the regulation of small RNA expression: SarA represses the expression of two sRNAs inStaphylococcus aureus

“…Its expression sharply decreases after S. aureus phagocytosis by human macrophages (33). We therefore investigated whether a TF might be involved in controlling srn_3610_sprC expression.…”

“…(33). Overnight cultures of S. aureus were diluted to an OD 600 nm of 0.1 in fresh BHI or TSB broth, then incubated at 37°C and 160 rpm.…”

“…We recently showed that Srn_3610_SprC attenuates S. aureus virulence and spread in an animal model of infection (33). Those observations imply that its expression must decrease during infection.…”

Small Regulatory RNAs of Rickettsia conorii

“…The in vitro transcribed sRNA and mRNA were mixed in a binding buffer (Promega) and incubated at 70 °C for 5 minutes followed by 30 °C for 15 minutes. Since RNAs are known to exhibit complex structures, incubation at 70–90 °C for 1–5 minutes was found to be optimal to achieve RNA population with homogeneous fold and has been extensively followed while performing EMSA studies to identify the sRNA-mRNA interactions697071. Further incubation at 30 °C for 15 minutes was chosen for this study as Rickettsia species while present in tick vectors are exposed to temperatures ranging from 28–30 °C.…”

“…Over the past decade, bacterial post-transcriptional regulation is garnering significant interest due to discoveries related to the roles of small RNAs in modulating gene expression during growth and stress conditions in vitro and host interactions in vivo 12. Bacterial small RNAs (sRNAs) are generally 50–500 bp long and fall into three categories, namely riboswitches which are located upstream of mRNAs, cis-acting sRNAs synthesized from the complementary strand of an open reading frame (ORF), and trans-acting sRNAs transcribed from the intergenic regions with only partial complementarity to their target genes.…”