AtxA-Controlled Small RNAs of Bacillus anthracis Virulence Plasmid pXO1 Regulate Gene Expression in trans.

Ileana D Corsi,Theresa M Koehler,Soumita Dutta,Ambro Van Hoof

doi:10.3389/fmicb.2020.610036

Abstract

Small regulatory RNAs (sRNAs) are short transcripts that base-pair to mRNA targets or interact with regulatory proteins. sRNA function has been studied extensively in Gram-negative bacteria; comparatively less is known about sRNAs in Firmicutes. Here we investigate two sRNAs encoded by virulence plasmid pXO1 of Bacillus anthracis, the causative agent of anthrax. The sRNAs, named “XrrA and XrrB” (for pXO1-encoded regulatory RNA) are abundant and highly stable primary transcripts, whose expression is dependent upon AtxA, the master virulence regulator of B. anthracis. sRNA levels are highest during culture conditions that promote AtxA expression and activity, and sRNA levels are unaltered in Hfq RNA chaperone null-mutants. Comparison of the transcriptome of a virulent Ames-derived strain to the transcriptome of isogenic sRNA-null mutants revealed multiple 4.0- to >100-fold differences in gene expression. Most regulatory effects were associated with XrrA, although regulation of some transcripts suggests functional overlap between the XrrA and XrrB. Many sRNA-regulated targets were chromosome genes associated with branched-chain amino acid metabolism, proteolysis, and transmembrane transport. Finally, in a mouse model for systemic anthrax, the lungs and livers of animals infected with xrrA-null mutants had a small reduction in bacterial burden, suggesting a role for XrrA in B. anthracis pathogenesis.

Highlights

Eukaryotic and bacterial organisms have evolved mechanisms employing specialized RNA molecules to modulate gene expression
We previously investigated regulatory functions of the B. anthracis phosphotransferase system - regulated domains (PRDs)-containing virulence regulators (PCVRs), AtxA, AcpA, and AcpB, using RNA-seq to compare gene expression by the Ames parent strain, an isogenic PCVR-null mutant, and strains of the PCVR-null mutant complemented with the individual PCVRs (Raynor et al, 2018)
To quantify RNAs associated with these loci and verify PCVR-mediated regulation, we re-analyzed raw paired-end reads from Raynor et al (2018) [raw paired-end reads accessible at NCBI Gene Expression Omnibus (GEO) database (Edgar et al, 2002), accession number GSE152357] using the Galaxy web resource for bioinformatic analysis (Afgan et al, 2018) (Figure 1A)

Summary

Introduction

Eukaryotic and bacterial organisms have evolved mechanisms employing specialized RNA molecules to modulate gene expression. The microRNAs (miRNAs) of eukaryotic cells are non-coding RNAs of 21 to 24 nucleotides that serve as mRNA-targeting templates to guide the function of miRNA-associated proteins (Grishok et al, 2001; Song et al, 2003; Vaucheret et al, 2004). Upon target recognition, these proteins, collectively referred to as Argonaute proteins, induce mRNA decay or inhibit mRNA translation initiation (Pillai et al, 2004; Filipowicz et al, 2005; Wu and Xie, 2006). Deletion of hfq results in shortened half-lives of sRNAs and impaired sRNA-mediated regulation (Vytvytska et al, 1998; Sledjeski et al, 2001; Lenz et al, 2004; Deng et al, 2012; Santiago-Frangos et al, 2016)

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Results

Conclusion