A Widespread Bacterial Secretion System with Diverse Substrates.

In host-associated bacteria, surface and secreted proteins mediate acquisition of nutrients, interactions with host cells, and specificity of tissue localization. In Gram-negative bacteria, the mechanism by which many proteins cross and/or become tethered to the outer membrane remains unclear. The domain of unknown function 560 (DUF560) occurs in outer membrane proteins throughout Proteobacteria and has been implicated in host-bacterium interactions and lipoprotein surface exposure. We used sequence similarity networking to reveal three subfamilies of DUF560 homologs. One subfamily includes those DUF560 proteins experimentally characterized thus far: NilB, a host range determinant of the nematode-mutualist Xenorhabdus nematophila, and the surface lipoprotein assembly modulators Slam1 and Slam2, which facilitate lipoprotein surface exposure in Neisseria meningitidis (Y. Hooda, C. C. Lai, A. Judd, C. M. Buckwalter, et al., Nat Microbiol 1:16009, 2016, https://doi.org/10.1038/nmicrobiol.2016.9; Y. Hooda, C. C. L. Lai, T. F. Moraes, Front Cell Infect Microbiol 7:207, 2017, https://doi.org/10.3389/fcimb.2017.00207). We show that DUF560 proteins from a second subfamily facilitate secretion of soluble, nonlipidated proteins across the outer membrane. Using in silico analysis, we demonstrate that DUF560 gene complement correlates with bacterial environment at a macro level and host association at a species level. The DUF560 protein superfamily represents a newly characterized Gram-negative secretion system capable of lipoprotein surface exposure and soluble protein secretion with conserved roles in facilitating symbiosis. In light of these data, we propose that it be titled the type 11 secretion system (TXISS). IMPORTANCE The microbial constituency of a host-associated microbiome emerges from a complex physical and chemical interplay of microbial colonization factors, host surface conditions, and host immunological responses. To fill unique niches within a host, bacteria encode surface and secreted proteins that enable interactions with and responses to the host and co-occurring microbes. Bioinformatic predictions of putative bacterial colonization factor localization and function facilitate hypotheses about the potential of bacteria to engage in pathogenic, mutualistic, or commensal activities. This study uses publicly available genome sequence data alongside experimental results from Xenorhabdus nematophila to demonstrate a role for DUF560 family proteins in secretion of bacterial effectors of host interactions. Our research delineates a broadly distributed family of proteins and enables more accurate predictions of the localization of colonization factors throughout Proteobacteria.

Electron Cryotomography of Bacterial Secretion Systems.

In biology, function arises from form. For bacterial secretion systems, which often span two membranes, avidly bind to the cell wall, and contain hundreds of individual proteins, studying form is a daunting task, made possible by electron cryotomography (ECT). ECT is the highest-resolution imaging technique currently available to visualize unique objects inside cells, providing a three-dimensional view of the shapes and locations of large macromolecular complexes in their native environment. Over the past 15 years, ECT has contributed to the study of bacterial secretion systems in two main ways: by revealing intact forms for the first time and by mapping components into these forms. Here we highlight some of these contributions, revealing structural convergence in type II secretion systems, structural divergence in type III secretion systems, unexpected structures in type IV secretion systems, and unexpected mechanisms in types V and VI secretion systems. Together, they offer a glimpse into a world of fantastic forms-nanoscale rotors, needles, pumps, and dart guns-much of which remains to be explored.

Phylogenetic profiling, an untapped resource for the prediction of secreted proteins and its complementation with sequence-based classifiers in bacterial type III, IV and VI secretion systems.

In the establishment and maintenance of the interaction between pathogenic or symbiotic bacteria with a eukaryotic organism, protein substrates of specialized bacterial secretion systems called effectors play a critical role once translocated into the host cell. Proteins are also secreted to the extracellular medium by free-living bacteria or directly injected into other competing organisms to hinder or kill. In this work, we explore an approach based on the evolutionary dependence that most of the effectors maintain with their specific secretion system that analyzes the co-occurrence of any orthologous protein group and their corresponding secretion system across multiple genomes. We compared and complemented our methodology with sequence-based machine learning prediction tools for the type III, IV and VI secretion systems. Finally, we provide the predictive results for the three secretion systems in 1606 complete genomes at http://www.iib.unsam.edu.ar/orgsissec/.

Secretion Systems Used by Bacteria to Subvert Host Functions.

In this review we examine the use of secretion systems by bacteria to subvert host functions. Bacteria have evolved multiple systems to interact with and overcome their eukaryotic host and other prokaryotes. Secretion systems are required for the release of several effectors through the bacterial membrane(s) into the extracellular space or directly into the cytoplasm of the host. We review the secretion systems of Gram-positive and Gram-negative bacteria and describe briefly the structural composition of the seven secretion systems that have been associated with increased virulence through subversion of host functions. Some of the effects of such systems on eukaryotic host processes have been studied extensively. We also describe the best-characterized effectors of each secretion system to give an overview of the molecular mechanisms employed by bacteria to hide from the immune system and convert eukaryotic cells into optimal ecological niches for their replication.

T6SS intraspecific competition orchestrates Vibrio cholerae genotypic diversity

Vibrio cholerae is a diverse species that inhabits a wide range of environments from copepods in brackish water to the intestines of humans. In order to remain competitive, V. cholerae uses the versatile type-VI secretion system (T6SS) to secrete anti-prokaryotic and anti-eukaryotic effectors. In addition to competing with other bacterial species, V. cholerae strains also compete with one another. Some strains are able to coexist, and are referred to as belonging to the same compatibility group. Challenged by diverse competitors in various environments, different V. choleare strains secrete different combination of effectors - presumably to best suit their niche. Interestingly, all pandemic V. cholerae strains encode the same three effectors. In addition to the diversity displayed in the encoded effectors, the regulation of V. cholerae also differs between strains. Two main layers of regulation appear to exist. One strategy connects T6SS activity with behavior that is suited to fighting eukaryotic cells, while the other is linked with natural competence - the ability of the bacterium to acquire and incorporate extracellular DNA. This relationship between bacterial killing and natural competence is potentially a source of diversification for V. cholerae as it has been shown to incorporate the DNA of cells recently killed through T6SS activity. It is through this process that we hypothesize the transfer of virulence factors, including T6SS effector modules, to happen. Switching of T6SS effectors has the potential to change the range of competitors V. cholerae can kill and to newly define which strains V. cholerae can co-exist with, two important parameters for survival in diverse environments (AU)

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Biological Functions of the Secretome of Neisseria meningitidis.

Neisseria meningitidis is a Gram-negative bacterial pathogen that normally resides as a commensal in the human nasopharynx but occasionally causes disease with high mortality and morbidity. To interact with its environment, it transports many proteins across the outer membrane to the bacterial cell surface and into the extracellular medium for which it deploys the common and well-characterized autotransporter, two-partner and type I secretion mechanisms, as well as a recently discovered pathway for the surface exposure of lipoproteins. The surface-exposed and secreted proteins serve roles in host-pathogen interactions, including adhesion to host cells and extracellular matrix proteins, evasion of nutritional immunity imposed by iron-binding proteins of the host, prevention of complement activation, neutralization of antimicrobial peptides, degradation of immunoglobulins, and permeabilization of epithelial layers. Furthermore, they have roles in interbacterial interactions, including the formation and dispersal of biofilms and the suppression of the growth of bacteria competing for the same niche. Here, we will review the protein secretion systems of N. meningitidis and focus on the functions of the secreted proteins.

Two-Partner Secretion: Combining Efficiency and Simplicity in the Secretion of Large Proteins for Bacteria-Host and Bacteria-Bacteria Interactions.

Initially identified in pathogenic Gram-negative bacteria, the two-partner secretion (TPS) pathway, also known as Type Vb secretion, mediates the translocation across the outer membrane of large effector proteins involved in interactions between these pathogens and their hosts. More recently, distinct TPS systems have been shown to secrete toxic effector domains that participate in inter-bacterial competition or cooperation. The effects of these systems are based on kin vs. non-kin molecular recognition mediated by specific immunity proteins. With these new toxin-antitoxin systems, the range of TPS effector functions has thus been extended from cytolysis, adhesion, and iron acquisition, to genome maintenance, inter-bacterial killing and inter-bacterial signaling. Basically, a TPS system is made up of two proteins, the secreted TpsA effector protein and its TpsB partner transporter, with possible additional factors such as immunity proteins for protection against cognate toxic effectors. Structural studies have indicated that TpsA proteins mainly form elongated ß helices that may be followed by specific functional domains. TpsB proteins belong to the Omp85 superfamily. Open questions remain on the mechanism of protein secretion in the absence of ATP or an electrochemical gradient across the outer membrane. The remarkable dynamics of the TpsB transporters and the progressive folding of their TpsA partners at the bacterial surface in the course of translocation are thought to be key elements driving the secretion process.