Evidence for Acquisition ofLegionellaType IV Secretion Substrates via Interdomain Horizontal Gene Transfer

Abstract

Intracellular pathogens exploit host cell functions to create a replication niche inside eukaryotic cells. The causative agent of Legionnaires' disease, the ␥-proteobacterium Legionella pneumophila, resides and replicates within a modified vacuole of protozoan and mammalian cells. L. pneumophila translocates effector proteins into host cells through the Icm-Dot complex, a specialized type IVB secretion system that is required for intracellular growth. To find out if some effector proteins may have been acquired through interdomain horizontal gene transfer (HGT), we performed a bioinformatic screen that searched for eukaryotic motifs in all open reading frames of the L. pneumophila Philadelphia-1 genome. We found 44 uncharacterized genes with many distinct eukaryotic motifs. Most of these genes contain G؉C biases compared to other L. pneumophila genes, supporting the theory that they were acquired through HGT. Furthermore, we found that several of them are expressed and up-regulated in stationary phase in an RpoS-dependent manner. In addition, at least seven of these gene products are translocated into host cells via the Icm-Dot complex, confirming their role in the intracellular environment. Reminiscent of the case with most Icm-Dot substrates, most of the strains containing mutations in these genes grew comparably to the parent strain intracellularly. Our findings suggest that in L. pneumophila, interdomain HGT may have been a major mechanism for the acquisition of determinants of infection.The ␥-proteobacterium Legionella pneumophila is an opportunistic human pathogen that multiplies within alveolar macrophages and causes the nosocomial and community-acquired pneumonia known as Legionnaires' disease (18,25,48). Human disease occurs when aerosolized L. pneumophila is inhaled from man-made or natural freshwater reservoirs harboring the bacteria. L. pneumophila poses a significant worldwide public health problem, particularly for individuals with compromised immune systems (19,38,40). Eradication of the pathogen from freshwater, industrial settings has proven difficult, since L. pneumophila thrives in environments that exclude antibacterial agents, such as biofilms and the intracellular compartments of protozoa (8,46,47).In order to create a replicative niche inside eukaryotic cells suitable for replication, L. pneumophila is believed to modulate host cell functions by the delivery of effector proteins through a type IVB secretion system known as the Icm-Dot complex (52, 59). Effector proteins presumably regulate several pathways in the host, including up-regulation of phagocytosis (23), delay in phagosome-lysosome fusion (24), recruitment of ARF1 to the phagosome (43), acquisition of endoplasmic reticulum-derived vesicles (29), and nonlytic egress from the host cell (11).Several laboratories found Icm-Dot substrates through genetic screens and bioinformatic approaches (3,7,11,13,36,43,44,57). Most of the known effector proteins are not individually required for intracellular multiplication, since knocking out th...

Genome Dynamics in Legionella: The Basis of Versatility and Adaptation to Intracellular Replication

“…Interestingly, many of these potential virulence factors never had been identified in a prokaryotic genome before the discovery in L. pneumophila, suggesting that L. pneumophila may have developed specific mechanisms to cross talk with its eukaryotic hosts (Bruggemann et al 2006). The presence of a high number of eukaryoticlike proteins in the genus Legionella was further confirmed when analyzing different L. pneumophila and L. longbeachae genomes (de Felipe et al 2005;Cazalet et al 2010;Kozak et al 2010;Schroeder et al 2010;Gomez-Valero et al 2011b). Thus, the Legionella genomes reflect the interaction of this bacterium with amoeba, which is probably the driving force in the evolution of Legionella pathogenicity.…”

“…Gomez-Valero and C. Buchrieser since then. In 2004, more than 60 of these proteins were described in the genomes of strains Paris and Lens (Cazalet et al 2004) and in 2005, in strain Philadelphia (de Felipe et al 2005). Later, the analysis of the distribution of these genes among more than 200 L. pneumophila strains using DNA arrays showed that the conservation of certain eukaryotic-like genes within the species L. pneumophila was high, indicating strong environmental selection pressures for their preservation (Cazalet et al 2008).…”

“…The main secretion system of Legionella, essential for intracellular growth, is the type IVB Dot/Icm secretion system (Marra and Shuman 1992;Berger and Isberg 1993). Using many dif-ferent methods it has been shown that at least 275 L. pneumophila proteins are secreted by this system (Campodonico et al 2005;de Felipe et al 2005;Shohdy et al 2005;de Felipe et al 2008;Burstein et al 2009;Heidtman et al 2009;Zhu et al 2011), which represents nearly 10% of the protein-coding genes predicted in the L. pneumophila genomes. At least 75 of these are eukaryotic-like proteins or carry eukaryotic domains and, for certain of these, it has also been shown experimentally that they interfere with host-cell signaling pathways ( Table 2).…”

The Response Regulator CpxR Directly Regulates Expression of SeveralLegionella pneumophila icm/dotComponents as Well as New Translocated Substrates

“…The L. pneumophila Icm/Dot type IV secretion system is composed of about 23 complex components, three accessory proteins, and a large number of IDTS (for reviews, see references 42 and 49), and many additional proteins have been suggested to be putative IDTS (6,12,29,61). This very large number of components must be coordinated at the level of gene expression in order to result in a successful infection and intracellular multiplication.…”

Bringing host-cell takeover by pathogenic bacteria to center stage

“…It seems that horizontal gene transfer from eukaryotes was a major mechanism in this co-evolution. 5 Two types of SVFs are currently in the spotlight ( Fig. 2): accessory factors of Rab GTPases and ankyrin repeat-containing proteins, Anks.…”