The LhrC sRNAs control expression of T cell-stimulating antigen TcsA in Listeria monocytogenes by decreasing tcsA mRNA stability

Abstract

The bacterial pathogen Listeria monocytogenes encodes seven homologous small regulatory RNAs, named the LhrC family of sRNAs. The LhrCs are highly induced under infection-relevant conditions and are known to inhibit the expression of multiple target mRNAs encoding virulence-associated surface proteins. In all cases studied so far, the LhrCs use their CU-rich regions for base pairing to complementary AG-rich sequences of the ribosomal binding site (RBS) of specific target mRNAs. Consequently, LhrC-mRNA interaction results in inhibition of translation followed by mRNA degradation, corresponding to the canonical model for sRNA-mediated gene regulation in bacteria. Here, we demonstrate that the LhrC sRNAs employ a different regulatory mechanism when acting to down-regulate the expression of tcsA, encoding a T cell-stimulating antigen. In this case, LhrC base pairs to an AG-rich site located well upstream of the RBS in tcsA mRNA. Using an in vitro translation assay, we found that LhrC could not prevent the ribosome from translating the tcsA messenger. Rather, the LhrC sRNAs act to decrease the half-life of tcsA mRNA in vivo. Importantly, LhrC-mediated destabilization of tcsA mRNA relies on an intact LhrC binding site near the 5´-end of the tcsA mRNA and occurs independently of translation. Based on these findings, we propose an alternative mechanism for LhrC-mediated control in L. monocytogenes that relies solely on sRNA-induced degradation of a target mRNA.

RNA-Dependent Regulation of Virulence in Pathogenic Bacteria

“…There are five copies of the lhrC gene ranging from 111 to 114 nucleotides in size (Sievers et al, 2014). Initial observations of LhrC expression during L. monocytogenes intracellular growth in macrophages and its putative role in facilitating pathogenesis have been substantiated by three reports of Kallipolitis and colleagues (Sievers et al, 2014; Dos Santos et al, 2018; Ross et al, 2019). LhrC RNAs 1-5 (LhrC1-5) were found to be upregulated during conditions of cell envelope stress such as those generated by the antibiotic cephalosporin or bile salts (Sievers et al, 2014).…”

Regulation of mRNA Stability During Bacterial Stress Responses

“…Additionally, while the stem-loop increases prfA stability (Loh et al, 2012), the binding of SreA to prfA triggers transcript degradation (Loh et al, 2009). Also in L. monocytogenes, posttranscriptional regulation of Tcsa, the T cell-stimulating antigen encoded by tcsA, was recently reported to be under the control of the sRNA LhrC in a translation-independent manner, by recruiting an undefined RNase (Ross et al, 2019). In S. aureus SarA, a histone-like protein, influences mRNA turnover of virulence factors, such as protein A (spa) and the collagen adhesion protein (cna) during exponential growth (Roberts et al, 2006;Morrison et al, 2012).…”

Small RNAs as Fundamental Players in the Transference of Information During Bacterial Infectious Diseases

“…Trans acting sRNAs bind to their targets with partial complementarity (Caldelari et al, 2013). There are different types of interaction, for instance the destabilization of the mRNA by pairing to upstream locations from RBS (Ross et al, 2019) or by interference with the 5 UTR (Rübsam et al, 2018). Binding of the sRNA can also mask the ribosomal binding site (RBS), consequently the ribosome cannot bind and the translation of the gene is attenuated (Kiekens et al, 2018).…”

“…In this organism, the seven sRNAs comprising the LhrC family act by regulating the target tcsA that encodes a virulence protein, CD4 + T cell-stimulating antigen (Sanderson et al, 1995). The regulatory mechanism of LhrCs was recently elucidated, and follow a non-canonical pathway, where sRNAs bind far upstream of the RBS, impairing the stability of the mRNA through the participation of an unknown RNase decreasing the messenger half-life (Ross et al, 2019; Table 2). During the nutritional starvation confronted by the pathogen in the host, the bacteria can make use of the available iron at the host iron-containing proteins, as the heme group in hemoglobin (Cassat and Skaar, 2013).…”