Small regulatory RNA and Legionella pneumophila

Review Article published: 06 May 2011 doi: 10.3389/fmicb.2011.00098 Small regulatory RNA and Legionella pneumophila Sébastien P. Faucher1* and Howard A. Shuman 2 1 2 Complex Traits Group, Department of Microbiology, McGill University, Montreal, QC, Canada Department of Microbiology, Cummings Life Sciences Center, University of Chicago, Chicago, IL, USA Edited by: Carmen Buchrieser, Pasteur Institute, France Reviewed by: Michele S. Swanson, University of Michigan, USA Jörg Vogel, University of Würzburg, Germany Gisela Storz, National Institutes of Health, USA *Correspondance: Sébastien P. Faucher, Complex Traits Group, Department of Microbiology, McGill University, 3649 Sir-WilliamOsler Promenade, Montreal, QC, Canada H3G 0B1. e-mail: Legionella pneumophila is a gram-negative bacterial species that is ubiquitous in almost any aqueous environment. It is the agent of Legionnaires’ disease, an acute and often under-reported form of pneumonia. In mammals, L. pneumophila replicates inside macrophages within a modified vacuole. Many protein regulators have been identified that control virulence-related properties, including RpoS, LetA/LetS, and PmrA/PmrB. In the past few years, the importance of regulation of virulence factors by small regulatory RNA (sRNAs) has been increasingly appreciated. This is also the case in L. pneumophila where three sRNAs (RsmY, RsmZ, and 6S RNA) were recently shown to be important determinants of virulence regulation and 79 actively transcribed sRNAs were identified. In this review we describe current knowledge about sRNAs and their regulatory properties and how this relates to the known regulatory systems of L. pneumophila. We also provide a model for sRNA-mediated control of gene expression that serves as a framework for understanding the regulation of virulence-related properties of L. pneumophila. Keywords: CsrA, RsmY, RsmZ, 6S RNA, cyclic di-GMP, CRISPR Introduction Legionella pneumophila is the causative agent of Legionnaires’ disease, an acute form of pneumonia (Fraser et al., 1977). It is a common, but often underestimated, cause of community-acquired and nosocomial pneumonia. The case-fatality rate of Legionellosis ranges between 10 and 40% and may approach 50% in nosocomial outbreaks, particularly among individuals with compromised health status (Benin et al., 2002). In Germany, where pneumonia causes are systematically investigated, Legionella is a leading cause of community-acquired pneumonia (von Baum et al., 2008). Legionella pneumophila is commonly found in almost all natural and engineered water systems where it replicates in a variety of phagocytic protozoa, including Hartmannella vermiformis. Transmission mechanisms are still unclear, but a clear association was found between local watershed hydrology and Legionellosis risk in Toronto (Ng et al., 2008), which indicates that environmental factors are key players in transmission to humans. In people, infection is thought to occur by inhalation of contaminated water droplets. Once in the lungs, L. pneumophila infects and replicates inside alveolar macrophages. To successfully infect and grow inside host cells, L. pneumophila circumvents normal endocytic trafficking pathways and inhibits phagosome acidification and fusion with lysosomes to establish a permissive replication niche called the Legionella containing vacuole (LCV) (Franco et al., 2009). The LCV is characterized by recruitment of early secretory vesicles, mitochondria, and membrane vesicles derived from the Golgi and endoplasmic reticulum (Roy and Tilney, 2002; Molofsky and Swanson, 2004; Shin and Roy, 2008). Central to the formation of the LCV and intracellular growth is the Icm/Dot type IVB secretion system, which translocates approximately 200 diverse effector proteins to the cytosol and LCV membrane (Segal and Shuman, 1998; Segal et al., 1998; Vogel et al., 1998; Cazalet et al., 2004; Chien et al., 2004; de Felipe et al., 2005; Burstein et al., 2009; Hubber and Roy, 2010; Faucher et al., 2011; Huang et al., 2011; Zhu et al., 2011). www.frontiersin.org Because L. pneumophila has evolved in a variety of niches, including aquatic environments, biofilms as well as within diverse hosts, different stress response pathways and virulence pathways must be correctly regulated. Although little is known about gene regulation in natural or engineered aquatic environments, several two-component systems are known to be involved in the regulation of stress response pathways and virulence factors required during host cells infection. These include PmrA/PmrB (Zusman et al., 2007), CpxR/CpxA (Altman and Segal, 2008) and LetA/LetS (Hammer et al., 2002). In addition, the sigma factor RpoS (σS) has been shown to regulate a number of known virulence factors including many Icm/Dot effectors (Hovel-Miner et al., 2009) and is required for intracellular multiplication in ameba and primary macrophages but not in macrophage-like cell lines, probably because of their reduced antimicrobial capacity (Hales and Shuman, 1999; AbuZant et al., 2006). There is an increased awareness of the role of small regulatory RNAs (sRNAs) in the regulation of virulence factors and other processes in bacterial pathogens (Papenfort and Vogel, 2010). sRNAs are short (40–500 nt) RNA molecules that typically do not encode proteins and mainly perform regulatory functions. They can originate from either primary transcripts, meaning the sRNA is transcribed from its own promoter and its transcription stops at a Rho-independent terminator, or from the processing of larger transcripts. The vast majority of sRNAs are post-transcriptional regulators that can either inhibit or enhance mRNA translation of the target mRNAs (Waters and Storz, 2009). Other sRNAs regulate gene expression by binding to and interfering with regulatory proteins and have global effects on gene expression. Riboswitches and untranslated regions (UTR) are not sRNA per se, being an intrinsic part of the mRNA, but they are often found by the methodologies used to identify small RNA molecules. May 2011 | Volume 2 | Article 98 | 1 Faucher and Shuman Putative sRNA molecules expressed by L. pneumophila were identified by both a bioinformatic approach as well as by deep RNAsequencing from growth in broth and inside A. castellanii (Faucher et al., 2010; Weissenmayer et al., 2011). In addition, a number of sRNAs have been implicated in the regulation of virulence factors of L. pneumophila, including the CsrB homologs RsmY and RsmZ (Rasis and Segal, 2009a; Sahr et al., 2009) and the RNA polymerase (RNAP) regulator 6S RNA (Faucher et al., 2010). This review aims to describe the current knowledge about sRNAs in general and provide a global perspective of the involvement of sRNA regulation systems in the behavior of L. pneumophila. Base-pairing sRNAs The most common type of regulatory sRNA are base-pairing sRNAs. They are short, highly structured RNA molecules that are complementary to some degree to their (...truncated)