Atomic Structure of Type VI Contractile Sheath from Pseudomonas aeruginosa

Pseudomonas aeruginosa has three type VI secretion systems (T6SSs), H1-, H2-, and H3-T6SS, each belonging to a distinct group. The two T6SS components, TssB/VipA and TssC/VipB, assemble to form tubules that conserve structural/functional homology with tail sheaths of contractile bacteriophages and pyocins. Here, we used cryoelectron microscopy to solve the structure of the H1-T6SS P. aeruginosa TssB1C1 sheath at 3.3 Å resolution. Our structure allowed us to resolve some features of the T6SS sheath that were not resolved in the Vibrio cholerae VipAB and Francisella tularensis IglAB structures. Comparison with sheath structures from other contractile machines, including T4 phage and R-type pyocins, provides a better understanding of how these systems have conserved similar functions/mechanisms despite evolution. We used the P. aeruginosa R2 pyocin as a structural template to build an atomic model of the TssB1C1 sheath in its extended conformation, allowing us to propose a coiled-spring-like mechanism for T6SS sheath contraction

TssA forms a gp6-like ring attached to the type VI secretion sheath

The type VI secretion system (T6SS) is a supra-molecular bacterial complex that resembles phage tails. It is a killing machine which fires toxins into target cells upon contraction of its TssBC sheath. Here, we show that TssA1 is a T6SS component forming dodecameric ring structures whose dimensions match those of the TssBC sheath and which can accommodate the inner Hcp tube. The TssA1 ring complex binds the T6SS sheath and impacts its behaviour in vivo. In the phage, the first disc of the gp18 sheath sits on a baseplate wherein gp6 is a dodecameric ring. We found remarkable sequence and structural similarities between TssA1 and gp6 C-termini, and propose that TssA1 could be a baseplate component of the T6SS. Furthermore, we identified similarities between TssK1 and gp8, the former interacting with TssA1 while the latter is found in the outer radius of the gp6 ring. These observations, combined with similarities between TssF and gp6Nterminus or TssG and gp53, lead us to propose a comparative model between the phage baseplate and the T6SS

Atomic Structure of Type VI Contractile Sheath from Pseudomonas aeruginosa

Pseudomonas aeruginosa has three type VI secretion systems (T6SSs), H1-, H2-, and H3-T6SS, each belonging to a distinct group. The two T6SS components, TssB/VipA and TssC/VipB, assemble to form tubules that conserve structural/functional homology with tail sheaths of contractile bacteriophages and pyocins. Here, we used cryoelectron microscopy to solve the structure of the H1-T6SS P. aeruginosa TssB1C1 sheath at 3.3 Å resolution. Our structure allowed us to resolve some features of the T6SS sheath that were not resolved in the Vibrio cholerae VipAB and Francisella tularensis IglAB structures. Comparison with sheath structures from other contractile machines, including T4 phage and R-type pyocins, provides a better understanding of how these systems have conserved similar functions/mechanisms despite evolution. We used the P. aeruginosa R2 pyocin as a structural template to build an atomic model of the TssB1C1 sheath in its extended conformation, allowing us to propose a coiled-spring-like mechanism for T6SS sheath contraction

Systematic Deletion of Homeobox Genes in Podospora anserina Uncovers Their Roles in Shaping the Fruiting Body

Higher fungi, which comprise ascomycetes and basidiomycetes, play major roles in the biosphere. Their evolutionary success may be due to the extended dikaryotic stage of their life cycle, which is the basis for their scientific name: the Dikarya. Dikaryosis is maintained by similar structures, the clamp in basidiomycetes and the crozier in ascomycetes. Homeodomain transcription factors are required for clamp formation in all basidiomycetes studied. We identified all the homeobox genes in the filamentous ascomycete fungus Podospora anserina and constructed deletion mutants for each of these genes and for a number of gene combinations. Croziers developed normally in these mutants, including those with up to six deleted homeogenes. However, some mutants had defects in maturation of the fruiting body, an effect that could be rescued by providing wild-type maternal hyphae. Analysis of mutants deficient in multiple homeogenes revealed interactions between the genes, suggesting that they operate as a complex network. Similar to their role in animals and plants, homeodomain transcription factors in ascomycetes are involved in shaping multicellular structures

Bio-engineering of bacterial microcompartments: a mini review

Bacterial microcompartments (BMCs) are protein-bound prokaryotic organelles, discovered in cyanobacteria more than 60 years ago. Functionally similar to eukaryotic cellular organelles, BMCs compartment metabolic activities in the cytoplasm, foremost to increase local enzyme concentration and prevent toxic intermediates from damaging the cytosolic content. Advanced knowledge of the functional and structural properties of multiple types of BMCs, particularly over the last 10 years, have highlighted design principles of microcompartments. This has prompted new research into their potential to function as programmable synthetic nano-bioreactors and novel bio-materials with biotechnological and medical applications. Moreover, due to the involvement of microcompartments in bacterial pathogenesis and human health, BMCs have begun to gain attention as potential novel drug targets. This mini-review gives an overview of important synthetic biology developments in the bioengineering of BMCs and a perspective on future directions in the field

TssA forms a gp6-like ring attached to the type VI secretion sheath

The type VI secretion system (T6SS) is a supra-molecular bacterial complex that resembles phage tails. It is a killing machine which fires toxins into target cells upon contraction of its TssBC sheath. Here, we show that TssA1 is a T6SS component forming dodecameric ring structures whose dimensions match those of the TssBC sheath and which can accommodate the inner Hcp tube. The TssA1 ring complex binds the T6SS sheath and impacts its behaviour in vivo. In the phage, the first disc of the gp18 sheath sits on a baseplate wherein gp6 is a dodecameric ring. We found remarkable sequence and structural similarities between TssA1 and gp6 C-termini, and propose that TssA1 could be a baseplate component of the T6SS. Furthermore, we identified similarities between TssK1 and gp8, the former interacting with TssA1 while the latter is found in the outer radius of the gp6 ring. These observations, combined with similarities between TssF and gp6N-terminus or TssG and gp53, lead us to propose a comparative model between the phage baseplate and the T6SS

Changes in mitochondrial glutathione levels and protein thiol oxidation in ∆yfh1 yeast cells and the lymphoblasts of patients with Friedreich's ataxia.

International audienceFriedreich's ataxia (FRDA) is a neurodegenerative disease caused by low levels of the mitochondrial protein frataxin. The main phenotypic features of frataxin-deficient human and yeast cells include iron accumulation in mitochondria, iron-sulfur cluster defects and high sensitivity to oxidative stress. Frataxin deficiency is also associated with severe impairment of glutathione homeostasis and changes in glutathione-dependent antioxidant defenses. The potential biological consequences of oxidative stress and changes in glutathione levels associated with frataxin deficiency include the oxidation of susceptible protein thiols and reversible binding of glutathione to the SH of proteins by S-glutathionylation. In this study, we isolated mitochondria from frataxin-deficient ∆yfh1 yeast cells and lymphoblasts of FRDA patients, and show evidence for a severe mitochondrial glutathione-dependent oxidative stress, with a low GSH/GSSG ratio, and thiol modifications of key mitochondrial enzymes. Both yeast and human frataxin-deficient cells had abnormally high levels of mitochondrial proteins binding an anti-glutathione antibody. Moreover, proteomics and immunodetection experiments provided evidence of thiol oxidation in α-ketoglutarate dehydrogenase (KGDH) or subunits of respiratory chain complexes III and IV. We also found dramatic changes in GSH/GSSG ratio and thiol modifications on aconitase and KGDH in the lymphoblasts of FRDA patients. Our data for yeast cells also confirm the existence of a signaling and/or regulatory process involving both iron and glutathione

Proline antagonizes GABA-induced quenching of quorum-sensing in Agrobacterium tumefaciens

Plants accumulate free L-proline (Pro) in response to abiotic stresses (drought and salinity) and presence of bacterial pathogens, including the tumor-inducing bacterium Agrobacterium tumefaciens. However, the function of Pro accumulation in host-pathogen interaction is still unclear. Here, we demonstrated that Pro antagonizes plant GABA-defense in the A. tumefaciens C58-induced tumor by interfering with the import of GABA and consequently the GABA-induced degradation of the bacterial quorum-sensing signal, 3-oxo-octanoylhomoserine lactone. We identified a bacterial receptor Atu2422, which is implicated in the uptake of GABA and Pro, suggesting that Pro acts as a natural antagonist of GABA-signaling. The Atu2422 amino acid sequence contains a Venus flytrap domain that is required for trapping GABA in human GABAB receptors. A constructed atu2422 mutant was more virulent than the wild type bacterium; moreover, transgenic plants with a low level of Pro exhibited less severe tumor symptoms than did their wild-type parents, revealing a crucial role for Venus flytrap GABA-receptor and relative abundance of GABA and Pro in host-pathogen interaction