Type IVB Secretion Systems of Legionella and Other Gram-Negative Bacteria

Abstract

Type IV secretion systems (T4SSs) play a central role in the pathogenicity of many important pathogens, including Agrobacterium tumefaciens, Helicobacter pylori, and Legionella pneumophila. The T4SSs are related to bacterial conjugation systems, and are classified into two subgroups, type IVA (T4ASS) and type IVB (T4BSS). The T4BSS, which is closely related to conjugation systems of IncI plasmids, was originally found in human pathogen L. pneumophila; pathogenesis by L. pneumophila infection requires functional Dot/Icm T4BSS. A zoonotic pathogen, Coxiella burnetii, and an arthropod pathogen, Rickettsiella grylli - both of which carry T4BSSs highly similar to the Legionella Dot/Icm system - are evolutionarily closely related and comprise a monophyletic group. A growing body of bacterial genomic information now suggests that T4BSSs are not limited to Legionella and related bacteria and IncI plasmids. Here, we review the current knowledge on T4BSS apparatus and component proteins, gained mainly from studies on L. pneumophila Dot/Icm T4BSS. Recent structural studies, along with previous findings, suggest that the Dot/Icm T4BSS contains components with primary or higher-order structures similar to those in other types of secretion systems - types II, III, IVA, and VI, thus highlighting the mosaic nature of T4BSS architecture.

Keywords: Coxiella; Dot/Icm; Legionella; Rickettsiella; conjugation; protein secretion; type IV secretion; type IVB secretion.

Figure 1

Genetic organizations of selected T4BSSs. Genetic organizations of T4BSSs from the following bacteria or plasmids are illustrated. Legionella pneumophila strain Philadelphia 1 (GenBank accession no. NC_002942); Legionella longbeachae NSW150 (NC_013861); Rickettsiella grylli (NZ_AAQJ02000001); Coxiella burnetii RSA 493 (NC_002971); Marinobacter aquaeolei VT8 pMAQU01 (NC_008738); Xanthomonas campestris pv. vesicatoria str. 85-10 pXCV183 (NC_007507); Achromobacter xylosoxidans A8 pA82 (NC_014642); Yersinia pseudotuberculosis IP 31758 153 kbp plasmid (NC_009705); Burkholderia vietnamiensis G4 pBVIE03 (NC_009229); Methylobacterium extorquens AM1 megaplasmid (NC_012811); Beijerinckia indica subsp. indica ATCC 9039 pBIND01 (NC_012811); Gluconobacter oxydans 621H pGOX1 (NC_006672); Burkholderia vietnamiensis G4 pBVIE04 (NC_009228), and IncI plasmid R64 (NC_005014). ORFs designated as “0163” are conserved in several T4BSSs but not in Legionella, Coxiella, Rickettsiella, or R64. ORFs designated as “TPase” are putative transposase derivatives. Notably, B. vietnamiensis G4 harbors multiple plasmids that carry distinct T4BSSs.

Figure 2

A phylogenetic tree of DotG/IcmE_862–1046. Proteins that have regions homologous to DotG/IcmE_862–1046 were selected by multiple rounds of PSIBLAST (Altschul et al., 1997) using the non-redundant protein database (nr), as of November 30, 2010. Legionella proteins homologous to Ti plasmid VirB10, RP4 plasmid TrbI and F plasmid TraB were incorporated in the analysis as outgroups. The evolutionary history was inferred using the Neighbor-Joining method (Saitou and Nei, 1987). The bootstrap consensus tree inferred from 500 replicates is taken to represent the evolutionary history of the taxa analyzed (Felsenstein, 1985). The percentages of replicate trees in which the associated taxa clustered together in the bootstrap test are shown next to the branches. The evolutionary distances were computed using the Poisson correction method (Zuckerkandl and Pauling, 1965) and are in the units of the number of amino acid substitutions per site. Evolutionary analyses were conducted in MEGA4 (Tamura et al., 2007).

Figure 3

Legionella pneumophila Dot/Icm T4BSS. The putative core complex containing DotC, DotD, DotH, DotG, and DotF was suggested by Vincent et al. (2006b). A possible scenario of its assembly is as follows: (1) outer membrane lipoproteins DotC and DotD recruit intrinsic periplasmic protein DotH to the outer membrane, thus forming a DotC–DotD–DotH outer membrane complex; (2) the C-terminal domain of DotG participates in the outer membrane complex, resulting in a complex spanning both inner and outer membranes; and (3) DotF participates in the core complex by binding to DotG and/or the DotC–DotD–DotH complex. Subcellular localization of Dot/Icm proteins are depicted based on lines of experimental evidence (Roy and Isberg, ; Zuckman et al., ; Coers et al., ; Matthews and Roy, ; Sexton et al., ,; Vincent et al., 2006b), or prediction from amino acid sequences (DotE, DotJ, DotV, and IcmT).

Figure 4

Core complex of pKM101 T4ASS. (A) Comparison of electron micrographic structures of Vibrio cholera secretin GspD (type II secretin; EMDB accession EMD-1763; Reichow et al., 2010), type III injectisome isolated from ΔinvJ Salmonella typhimurium (type III needle base; EMD-1224; Marlovits et al., 2006), and T4ASS core complex of pKM101 conjugal plasmid (type IVA core; EMD-5031; Fronzes et al., 2009). (B) Top and side views of pKM101 outer membrane complex (PDB accession 3JQO; Chandran et al., 2009). One of each protomer in the complex is shown in color: VirB7 (blue), VirB9 (green), and VirB10 (red). (C) VirB7 takes an extended form in the complex. Figures are generated using PyMol (Schrodinger, 2010) and resources deposited to indicated databases.

Figure 5

Comparison of the C-terminal domain of DotD with secretin periplasmic subdomains. (A) Domain organizations of L. pneumophila DotD (Nakano et al., 2010), EPEC secretin EscC (Spreter et al., 2009), and ETEC secretin GspD (Korotkov et al., 2009). (B) DotD (green, PDB accession 3ADY) superimposed onto the N0 domain of ETEC secretin GspD (blue, PDB 3EZJ). (C) DotD (green) superimposed onto the T3S domain of EPEC secretin EscC (light blue, PDB 3GR5). (D) A model of T4BSS core complex. DotD may form a periplasmic ring, like the N0 domain of type II secretin.