Phylogenomics Reveals a DiverseRickettsialesType IV Secretion System

Abstract

3With an obligate intracellular lifestyle, Alphaproteobacteria of the order Rickettsiales have inextricably coevolved with their various eukaryotic hosts, resulting in small, reductive genomes and strict dependency on host resources. Unsurprisingly, large portions of Rickettsiales genomes encode proteins involved in transport and secretion. One particular transporter that has garnered recent attention from researchers is the type IV secretion system (T4SS). Homologous to the well-studied archetypal vir T4SS of Agrobacterium tumefaciens, the Rickettsiales vir homolog (rvh) T4SS is characterized primarily by duplication of several of its genes and scattered genomic distribution of all components in several conserved islets. Phylogeny estimation suggests a single event of ancestral acquirement of the rvh T4SS, likely from a nonalphaproteobacterial origin. Bioinformatics analysis of over 30 Rickettsiales genome sequences illustrates a conserved core rvh scaffold (lacking only a virB5 homolog), with lineage-specific diversification of several components (rvhB1, rvhB2, and rvhB9b), likely a result of modifications to cell envelope structure. This coevolution of the rvh T4SS and cell envelope morphology is probably driven by adaptations to various host cells, identifying the transporter as an important target for vaccine development. Despite the genetic intractability of Rickettsiales, recent advancements have been made in the characterization of several components of the rvh T4SS, as well as its putative regulators and substrates. While current data favor a role in effector translocation, functions in DNA uptake and release and/or conjugation cannot at present be ruled out, especially considering that a mechanism for plasmid transfer in Rickettsia spp. has yet to be proposed.Type IV secretion systems (T4SSs) are macromolecular complexes that transport protein, DNA, and nucleoprotein across the bacterial cell envelope in both Gram-negative and Gram-positive species, as well as some wall-less bacteria and archaea (1, 32). Functioning in naked DNA uptake and release (60), conjugation (80), and the propagation of genomic islands (69), T4SSs are prominent factors in bacterial diversification and are responsible for the horizontal spread of antimicrobial resistance and virulence genes. T4SSs are also used by some species to deliver effector molecules (DNA and/or protein) into eukaryotic host cells (28), a process that facilitates infection and subsequent pathogenesis. It is assumed that all varieties of T4SSs form a channel that spans the cell envelope and culminates in a surface-exposed structure, such as a pilus (Fig. 1A). Despite this conserved architecture, genetic diversity in a multitude of features, including gene composition and organization, underlies the hundreds of T4SSs identified through genome sequencing. Recently, T4SSs have been classified into four groups: F, P, I, and GI (70). F-T4SSs and P-T4SSs (previously known as type IVA) are widespread systems represented by the archetypes encoded by the F plasmid o...

A Rickettsia Genome Overrun by Mobile Genetic Elements Provides Insight into the Acquisition of Genes Characteristic of an Obligate Intracellular Lifestyle

“…Aside from rickA (discussed above), a few exceptions are noteworthy. Several genes encoding components of the rvh P-T4SS, which is highly conserved in Rickettasiales (48), are split or truncated relative to their counterparts in other genomes. Of the 18 rvh T4SS genes conserved in Rickettsia genomes (46), eight are problematic in REIS (rvhB1, rvhB6a, rvhB6b, rvhB6c, rvhB6e, rvhB9b, rvhB10, and rvhD4).…”

“…However, prior to this report, only the genomes of R. bellii and R. massiliae were known to encode full F-T4SSs likely involved in the spread of rickettsial plasmids. Thus, rickettsial genomes harboring plasmids, yet without a complete F-T4SS, either are incapable of transferring plasmids or use an alternative mechanism for conjugation, such as transduction or the conserved Rickettsiales vir homolog (rvh) P-like T4SS (46,48). The analysis of common protein families across all four pREIS plasmids, as well as five additional rickettsial plasmids (pRF of R. felis, pRM of R. monacensis, pRMA of R. massiliae, pRPR of R. peacockii, and pRFA of R. africae), identified 40 genes present at least once on multiple rickettsial plasmids (see Table S6 in the supplemental material).…”

Chimeric Coupling Proteins Mediate Transfer of Heterologous Type IV Effectors through the Escherichia coli pKM101-Encoded Conjugation Machine

“…We expanded our studies by testing the functionality of chimeric T4CPs composed of TraJ's NTD joined to the soluble domains of VirD4 Ap and VirD4 Wp from A. phagocytophilum and W. pipientis, respectively. Rickettsial species carry T4SS gene clusters, and a growing body of evidence indicates that type IV secretion is a general requirement for intracellular survival and proliferation (10,43,65,66). Identification of T4SS effectors, however, has been hampered by the inability to grow rickettsial species axenically and by lack of robust genetic systems.…”

Loss of Immunization-Induced Epitope-Specific CD4 T-Cell Response following Anaplasma marginale Infection Requires Presence of the T-Cell Epitope on the Pathogen and Is Not Associated with an Increase in Lymphocytes Expressing Known Regulatory Cell Phenotypes

“…A. marginale, an intracellular rickettsial pathogen, lacks lipopolysaccharide (LPS) and peptidoglycan (31,46) and does not appear to evoke a pathological inflammatory response during acute infection. Through gene conversion, A. marginale undergoes extensive and continual antigenic variation in surface MSP2 and MSP3 throughout persistent infection (3), enabling it to avoid preexisting antibody response to the dominant MSP2 variant (5).…”