-
Nature Apr 2023Endosymbiotic bacteria have evolved intricate delivery systems that enable these organisms to interface with host biology. One example, the extracellular contractile...
Endosymbiotic bacteria have evolved intricate delivery systems that enable these organisms to interface with host biology. One example, the extracellular contractile injection systems (eCISs), are syringe-like macromolecular complexes that inject protein payloads into eukaryotic cells by driving a spike through the cellular membrane. Recently, eCISs have been found to target mouse cells, raising the possibility that these systems could be harnessed for therapeutic protein delivery. However, whether eCISs can function in human cells remains unknown, and the mechanism by which these systems recognize target cells is poorly understood. Here we show that target selection by the Photorhabdus virulence cassette (PVC)-an eCIS from the entomopathogenic bacterium Photorhabdus asymbiotica-is mediated by specific recognition of a target receptor by a distal binding element of the PVC tail fibre. Furthermore, using in silico structure-guided engineering of the tail fibre, we show that PVCs can be reprogrammed to target organisms not natively targeted by these systems-including human cells and mice-with efficiencies approaching 100%. Finally, we show that PVCs can load diverse protein payloads, including Cas9, base editors and toxins, and can functionally deliver them into human cells. Our results demonstrate that PVCs are programmable protein delivery devices with possible applications in gene therapy, cancer therapy and biocontrol.
Topics: Animals; Humans; Mice; Cell Membrane; Eukaryotic Cells; Photorhabdus; CRISPR-Associated Protein 9; Toxins, Biological; Proteins; Drug Delivery Systems; Protein Transport
PubMed: 36991127
DOI: 10.1038/s41586-023-05870-7 -
Nature Dec 2019The current need for novel antibiotics is especially acute for drug-resistant Gram-negative pathogens. These microorganisms have a highly restrictive permeability...
The current need for novel antibiotics is especially acute for drug-resistant Gram-negative pathogens. These microorganisms have a highly restrictive permeability barrier, which limits the penetration of most compounds. As a result, the last class of antibiotics that acted against Gram-negative bacteria was developed in the 1960s. We reason that useful compounds can be found in bacteria that share similar requirements for antibiotics with humans, and focus on Photorhabdus symbionts of entomopathogenic nematode microbiomes. Here we report a new antibiotic that we name darobactin, which was obtained using a screen of Photorhabdus isolates. Darobactin is coded by a silent operon with little production under laboratory conditions, and is ribosomally synthesized. Darobactin has an unusual structure with two fused rings that form post-translationally. The compound is active against important Gram-negative pathogens both in vitro and in animal models of infection. Mutants that are resistant to darobactin map to BamA, an essential chaperone and translocator that folds outer membrane proteins. Our study suggests that bacterial symbionts of animals contain antibiotics that are particularly suitable for development into therapeutics.
Topics: Animals; Anti-Bacterial Agents; Bacterial Outer Membrane Proteins; Cell Line; Disease Models, Animal; Drug Discovery; Drug Resistance, Microbial; Escherichia coli Proteins; Female; Gastrointestinal Microbiome; Gram-Negative Bacteria; Humans; Mice; Microbial Sensitivity Tests; Microbial Viability; Mutation; Nematoda; Operon; Phenylpropionates; Photorhabdus; Substrate Specificity; Symbiosis
PubMed: 31747680
DOI: 10.1038/s41586-019-1791-1 -
Cell Apr 2019Contractile injection systems (CISs) are cell-puncturing nanodevices that share ancestry with contractile tail bacteriophages. Photorhabdus virulence cassette (PVC)...
Contractile injection systems (CISs) are cell-puncturing nanodevices that share ancestry with contractile tail bacteriophages. Photorhabdus virulence cassette (PVC) represents one group of extracellular CISs that are present in both bacteria and archaea. Here, we report the cryo-EM structure of an intact PVC from P. asymbiotica. This over 10-MDa device resembles a simplified T4 phage tail, containing a hexagonal baseplate complex with six fibers and a capped 117-nanometer sheath-tube trunk. One distinct feature of the PVC is the presence of three variants for both tube and sheath proteins, indicating a functional specialization of them during evolution. The terminal hexameric cap docks onto the topmost layer of the inner tube and locks the outer sheath in pre-contraction state with six stretching arms. Our results on the PVC provide a framework for understanding the general mechanism of widespread CISs and pave the way for using them as delivery tools in biological or therapeutic applications.
Topics: Bacteriophage T4; Cell Membrane; Cryoelectron Microscopy; Models, Molecular; Photorhabdus; Protein Conformation; Type VI Secretion Systems
PubMed: 30905475
DOI: 10.1016/j.cell.2019.02.020 -
Journal of Molecular Biology Nov 2019Phenotypic heterogeneity in bacterial cell populations allows genetically identical organisms to different behavior under similar environmental conditions. The... (Review)
Review
Phenotypic heterogeneity in bacterial cell populations allows genetically identical organisms to different behavior under similar environmental conditions. The Gram-negative bacterium Photorhabdus luminescens is an excellent organism to study phenotypic heterogeneity since their life cycle involves a symbiotic interaction with soil nematodes as well as a pathogenic association with insect larvae. Phenotypic heterogeneity is highly distinct in P. luminescens. The bacteria exist in two phenotypic forms that differ in various morphologic and phenotypic traits and are therefore distinguished as primary (1°) and secondary (2°) cells. The 1 cells are bioluminescent, pigmented, produce several secondary metabolites and exo-enzymes, and support nematode growth and development. The 2° cells lack all these 1°-specific phenotypes. The entomopathogenic nematodes carry 1° cells in their upper gut and release them into an insect's body after slipping inside. During insect infection, up to the half number of 1° cells undergo phenotypic switching and convert to 2° cells. Since the 2° cells are not able to live in nematode symbiosis any more, they cannot re-associate with their symbiosis partners after the infection and remain in the soil. Phenotypic switching in P. luminescens has to be tightly regulated since a high switching frequency would lead to a complete break-down of the nematode-bacteria life cycle. Here, we present the main regulatory mechanisms known to-date that are important for phenotypic switching in P. luminescens cell populations and discuss the biological reason as well as the fate of the 2° cells in the soil.
Topics: Bacterial Physiological Phenomena; Bacterial Proteins; Biological Variation, Population; Gene Expression Regulation, Bacterial; Life Cycle Stages; Phenotype; Photorhabdus; Pigmentation; Quorum Sensing; Symbiosis
PubMed: 31022406
DOI: 10.1016/j.jmb.2019.04.015 -
Ecology and Evolution Feb 2024Understanding how parasites evolved is crucial to understand the host and parasite interaction. The evolution of entomopathogenesis in rhabditid nematodes has... (Review)
Review
Understanding how parasites evolved is crucial to understand the host and parasite interaction. The evolution of entomopathogenesis in rhabditid nematodes has traditionally been thought to have occurred twice within the phylum Nematoda: in Steinernematidae and Heterorhabditidae families, which are associated with the entomopathogenic bacteria and , respectively. However, nematodes from other families that are associated with entomopathogenic bacteria have not been considered to meet the criteria for "entomopathogenic nematodes." The evolution of parasitism in nematodes suggests that ecological and evolutionary properties shared by families in the order Rhabditida favor the convergent evolution of the entomopathogenic trait in lineages with diverse lifestyles, such as saprotrophs, phoretic, and necromenic nematodes. For this reason, this paper proposes expanding the term "entomopathogenic nematode" considering the diverse modes of this attribute within Rhabditida. Despite studies are required to test the authenticity of the entomopathogenic trait in the reported species, they are valuable links that represent the early stages of specialized lineages to entomopathogenic lifestyle. An ecological and evolutionary exploration of these nematodes has the potential to deepen our comprehension of the evolution of entomopathogenesis as a convergent trait spanning across the Nematoda.
PubMed: 38352205
DOI: 10.1002/ece3.10966 -
Cellular Microbiology Dec 2015Mono-glycosylation of host proteins is a common mechanism by which bacterial protein toxins manipulate cellular functions of eukaryotic target host cells. Prototypic for... (Review)
Review
Mono-glycosylation of host proteins is a common mechanism by which bacterial protein toxins manipulate cellular functions of eukaryotic target host cells. Prototypic for this group of glycosyltransferase toxins are Clostridium difficile toxins A and B, which modify guanine nucleotide-binding proteins of the Rho family. However, toxin-induced glycosylation is not restricted to the Clostridia. Various types of bacterial pathogens including Escherichia coli, Yersinia, Photorhabdus and Legionella species produce glycosyltransferase toxins. Recent studies discovered novel unexpected variations in host protein targets and amino acid acceptors of toxin-catalysed glycosylation. These findings open new perspectives in toxin as well as in carbohydrate research.
Topics: Bacterial Toxins; Eukaryotic Cells; Glycosylation; Glycosyltransferases; Gram-Negative Bacteria; Gram-Positive Bacteria; Host-Pathogen Interactions; Virulence
PubMed: 26445410
DOI: 10.1111/cmi.12533 -
Microbiology (Reading, England) Apr 2020Different model systems have, over the years, contributed to our current understanding of the molecular mechanisms underpinning the various types of interaction between... (Review)
Review
Different model systems have, over the years, contributed to our current understanding of the molecular mechanisms underpinning the various types of interaction between bacteria and their animal hosts. The genus comprises Gram-negative insect pathogenic bacteria that are normally found as symbionts that colonize the gut of the infective juvenile stage of soil-dwelling nematodes from the family . The nematodes infect susceptible insects and release the bacteria into the insect haemolymph where the bacteria grow, resulting in the death of the insect. At this stage the nematodes feed on the bacterial biomass and, following several rounds of reproduction, the nematodes develop into infective juveniles that leave the insect cadaver in search of new hosts. Therefore has three distinct and obligate roles to play during this life-cycle: (1) must kill the insect host; (2) must be capable of supporting nematode growth and development; and (3) must be able to colonize the gut of the next generation of infective juveniles before they leave the insect cadaver. In this review I will discuss how genetic analysis has identified key genes involved in mediating, and regulating, the interaction between and each of its invertebrate hosts. These studies have resulted in the characterization of several new families of toxins and a novel inter-kingdom signalling molecule and have also uncovered an important role for phase variation in the regulation of these different roles.
Topics: Animals; Bacterial Toxins; Gastrointestinal Tract; Host Microbial Interactions; Insecta; Life Cycle Stages; Photorhabdus; Rhabditoidea; Signal Transduction; Symbiosis
PubMed: 32209172
DOI: 10.1099/mic.0.000907 -
Current Topics in Microbiology and... 2017There is a complex interplay between the regulation of flagellar motility and the expression of virulence factors in many bacterial pathogens. Here, we review the... (Review)
Review
There is a complex interplay between the regulation of flagellar motility and the expression of virulence factors in many bacterial pathogens. Here, we review the literature on the direct and indirect roles of flagellar motility in mediating the tripartite interaction between entomopathogenic bacteria (Photorhabdus and Xenorhabdus), their nematode hosts, and their insect targets. First, we describe the swimming and swarming motility of insect pathogenic bacteria and its impact on insect colonization. Then, we describe the coupling between the expression of flagellar and virulence genes and the dynamic of expression of the flagellar regulon during invertebrate infection. We show that the flagellar type 3 secretion system (T3SS) is also an export apparatus for virulence proteins in X. nematophila. Finally, we demonstrate that phenotypic variation, a common property of the bacterial symbionts of nematodes, also alters flagellar motility in Photorhabdus and Xenorhabdus. Finally, the so-called phenotypic heterogeneity phenomenon in the flagellar gene expression network will be also discussed. As the main molecular studies were performed in X. nematophila, future perspectives for the study of the interplay between flagellum and invertebrate interactions in Photorhabdus will be discussed.
Topics: Flagella; Photorhabdus; Symbiosis; Virulence; Xenorhabdus
PubMed: 28091933
DOI: 10.1007/82_2016_53 -
Science Advances Apr 2022Extracellular contractile injection systems (eCISs) are widespread bacterial nanomachines that resemble T4 phage tail. As a typical eCIS, virulence cassette (PVC) was...
Extracellular contractile injection systems (eCISs) are widespread bacterial nanomachines that resemble T4 phage tail. As a typical eCIS, virulence cassette (PVC) was proposed to inject toxins into eukaryotic cells by puncturing the cell membrane from outside. This makes it an ideal tool for protein delivery in biomedical research. However, how to manipulate this nanocomplex as a molecular syringe is still undetermined. Here, we identify that one group of N-terminal signal peptide (SP) sequences are crucial for the effector loading into the inner tube of PVC complex. By application of genetic operation, cryo-electron microscopy, in vitro translocation assays, and animal experiments, we show that, under the guidance of the SP, numerous prokaryotic and eukaryotic proteins can be loaded into PVC to exert their functions across cell membranes. We therefore might customize PVC as a potent protein delivery nanosyringe for biotherapy by selecting cargo proteins in a broad spectrum, regardless of their species, sizes, and charges.
Topics: Animals; Cryoelectron Microscopy; Photorhabdus; Polyvinyl Chloride; Protein Sorting Signals; Virulence
PubMed: 35486720
DOI: 10.1126/sciadv.abm2343 -
Applied Microbiology and Biotechnology Jun 2022Insects and fungal pathogens pose constant problems to public health and agriculture, especially in resource-limited parts of the world; and the use of chemical... (Review)
Review
Insects and fungal pathogens pose constant problems to public health and agriculture, especially in resource-limited parts of the world; and the use of chemical pesticides continues to be the main methods for the control of these organisms. Photorhabdus spp. and Xenorhabdus spp., (Fam; Morganellaceae), enteric symbionts of Steinernema, and Heterorhabditis nematodes are naturally found in soil on all continents, except Antarctic, and on many islands throughout the world. These bacteria produce diverse secondary metabolites that have important biological and ecological functions. Secondary metabolites include non-ribosomal peptides, polyketides, and/or hybrid natural products that are synthesized using polyketide synthetase (PRS), non-ribosomal peptide synthetase (NRPS), or similar enzymes and are sources of new pesticide/drug compounds and/or can serve as lead molecules for the design and synthesize of new alternatives that could replace current ones. This review addresses the effects of these bacterial symbionts on insect pests, fungal phytopathogens, and animal pathogens and discusses the substances, mechanisms, and impacts on agriculture and public health. KEY POINTS: • Insects and fungi are a constant menace to agricultural and public health. • Chemical-based control results in resistance development. • Photorhabdus and Xenorhabdus are compelling sources of biopesticides.
Topics: Animals; Biological Products; Insecta; Nematoda; Photorhabdus; Rhabditida; Symbiosis; Xenorhabdus
PubMed: 35723692
DOI: 10.1007/s00253-022-12023-9