Staphylococcus aureus is a Gram-positive bacterial pathogen of global concern and a leading cause of bacterial infections worldwide. Asymptomatic carriage of S . aureus on the skin and in the anterior nares is common and recognized as a predisposing factor to invasive infection. Transition of S . aureus from the carriage state to that of invasive infection is often accompanied by a temperature upshift from approximately 33°C to 37°C. Such a temperature shift is known in other pathogens to influence gene expression, often resulting in increased production of factors that promote survival or virulence within the host. One mechanism by which bacteria modulate gene expression in response to temperature is by the regulatory activity of RNA-based thermosensors, cis- acting riboregulators that control translation efficiency. This study was designed to identify and characterize RNA-based thermosensors in S . aureus . Initially predicted by in silico analyses of the S . aureus USA300 genome, reporter-based gene expression analyses and site-specific mutagenesis were performed to demonstrate the presence of a functional thermosensor within the 5’ UTR of cidA , a gene implicated in biofilm formation and survival of the pathogen. The nucleic sequence composing the identified thermosensor are sufficient to confer temperature-dependent post-transcriptional regulation, and activity is predictably altered by the introduction of site-specific mutations designed to stabilize or destabilize the structure within the identified thermosensor. The identified regulator is functional in both the native bacterial host S . aureus and in the distally related species Escherichia coli , suggesting that its regulatory activity is independent of host-specific factors. Interestingly, unlike the majority of bacterial RNA-based thermosensors characterized to date, the cidA thermosensor facilitates increased target gene expression at lower temperatures. In addition to the characterization of the first RNA-based thermosensor in the significant pathogen S . aureus , it highlights the diversity of function within this important class of ribo-regulators.
RNA thermometers are cis-acting riboregulators that mediate the posttranscriptional regulation of gene expression in response to environmental temperature. Such regulation is conferred by temperature-responsive structural changes within the RNA thermometer that directly result in differential ribosomal binding to the regulated transcript. The significance of RNA thermometers in controlling bacterial physiology and pathogenesis is becoming increasingly clear. This study combines in silico, molecular genetics, and biochemical analyses to characterize both the structure and function of a newly identified RNA thermometer within the ompA transcript of Shigella dysenteriae. First identified by in silico structural predictions, genetic analyses have demonstrated that the ompA RNA thermometer is a functional riboregulator sufficient to confer posttranscriptional temperature-dependent regulation, with optimal expression observed at the host-associated temperature of 37°C. Structural studies and ribosomal binding analyses have revealed both increased exposure of the ribosomal binding site and increased ribosomal binding to the ompA transcript at permissive temperatures. The introduction of site-specific mutations predicted to alter the temperature responsiveness of the ompA RNA thermometer has predictable consequences for both the structure and function of the regulatory element. Finally, in vitro tissue culture-based analyses implicate the ompA RNA thermometer as a bona fide S. dysenteriae virulence factor in this bacterial pathogen. Given that ompA is highly conserved among Gram-negative pathogens, these studies not only provide insight into the significance of riboregulation in controlling Shigella virulence, but they also have the potential to facilitate further understanding of the physiology and/or pathogenesis of a wide range of bacterial species.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.