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1.
mBio ; 15(7): e0087124, 2024 Jul 17.
Artículo en Inglés | MEDLINE | ID: mdl-38899869

RESUMEN

Chemosensory systems allow bacteria to respond and adapt to environmental conditions. Many bacteria contain more than one chemosensory system, but knowledge of their specific roles in regulating different functions remains scarce. Here, we address this issue by analyzing the function of the F6, F8, and alternative (non-motility) cellular functions (ACF) chemosensory systems of the model plant pathogen Pseudomonas syringae pv. tomato. In this work, we assign PsPto chemoreceptors to each chemosensory system, and we visualize for the first time the F6 and F8 chemosensory systems of PsPto using cryo-electron tomography. We confirm that chemotaxis and swimming motility are controlled by the F6 system, and we demonstrate how different components from the F8 and ACF systems also modulate swimming motility. We also determine how the kinase and response regulators from the F6 and F8 chemosensory systems do not work together in the regulation of biofilm, whereas both components from the ACF system contribute together to regulate these traits. Furthermore, we show how the F6, F8, and ACF kinases interact with the ACF response regulator WspR, supporting crosstalk among chemosensory systems. Finally, we reveal how all chemosensory systems play a role in regulating virulence. IMPORTANCE: Chemoperception through chemosensory systems is an essential feature for bacterial survival, as it allows bacterial interaction with its surrounding environment. In the case of plant pathogens, it is especially relevant to enter the host and achieve full virulence. Multiple chemosensory systems allow bacteria to display a wider plasticity in their response to external signals. Here, we perform a deep characterization of the F6, F8, and alternative (non-motility) cellular functions chemosensory systems in the model plant pathogen Pseudomonas syringae pv. tomato DC3000. These chemosensory systems regulate key virulence-related traits, like motility and biofilm formation. Furthermore, we unveil an unexpected crosstalk among these chemosensory systems at the level of the interaction between kinases and response regulators. This work shows novel results that contribute to the knowledge of chemosensory systems and their role in functions alternative to chemotaxis.


Asunto(s)
Proteínas Bacterianas , Biopelículas , Quimiotaxis , Pseudomonas syringae , Solanum lycopersicum , Pseudomonas syringae/genética , Pseudomonas syringae/patogenicidad , Pseudomonas syringae/metabolismo , Pseudomonas syringae/fisiología , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Biopelículas/crecimiento & desarrollo , Solanum lycopersicum/microbiología , Virulencia , Enfermedades de las Plantas/microbiología , Regulación Bacteriana de la Expresión Génica
2.
EMBO J ; 41(10): e109523, 2022 05 16.
Artículo en Inglés | MEDLINE | ID: mdl-35301732

RESUMEN

The process by which bacterial cells build their intricate flagellar motility apparatuses has long fascinated scientists. Our understanding of this process comes mainly from studies of purified flagella from two species, Escherichia coli and Salmonella enterica. Here, we used electron cryo-tomography (cryo-ET) to image the assembly of the flagellar motor in situ in diverse Proteobacteria: Hylemonella gracilis, Helicobacter pylori, Campylobacter jejuni, Pseudomonas aeruginosa, Pseudomonas fluorescens, and Shewanella oneidensis. Our results reveal the in situ structures of flagellar intermediates, beginning with the earliest flagellar type III secretion system core complex (fT3SScc) and MS-ring. In high-torque motors of Beta-, Gamma-, and Epsilon-proteobacteria, we discovered novel cytoplasmic rings that interact with the cytoplasmic torque ring formed by FliG. These rings, associated with the MS-ring, assemble very early and persist until the stators are recruited into their periplasmic ring; in their absence the stator ring does not assemble. By imaging mutants in Helicobacter pylori, we found that the fT3SScc proteins FliO and FliQ are required for the assembly of these novel cytoplasmic rings. Our results show that rather than a simple accretion of components, flagellar motor assembly is a dynamic process in which accessory components interact transiently to assist in building the complex nanomachine.


Asunto(s)
Campylobacter jejuni , Helicobacter pylori , Proteínas Bacterianas/metabolismo , Campylobacter jejuni/metabolismo , Tomografía con Microscopio Electrónico/métodos , Escherichia coli/genética , Escherichia coli/metabolismo , Flagelos/metabolismo , Sistemas de Secreción Tipo III/metabolismo
3.
Nat Commun ; 11(1): 5763, 2020 11 13.
Artículo en Inglés | MEDLINE | ID: mdl-33188180

RESUMEN

The prokaryotic chemotaxis system is arguably the best-understood signaling pathway in biology. In all previously described species, chemoreceptors organize into a hexagonal (P6 symmetry) extended array. Here, we report an alternative symmetry (P2) of the chemotaxis apparatus that emerges from a strict linear organization of the histidine kinase CheA in Treponema denticola cells, which possesses arrays with the highest native curvature investigated thus far. Using cryo-ET, we reveal that Td chemoreceptor arrays assume an unusual arrangement of the supra-molecular protein assembly that has likely evolved to accommodate the high membrane curvature. The arrays have several atypical features, such as an extended dimerization domain of CheA and a variant CheW-CheR-like fusion protein that is critical for maintaining an ordered chemosensory apparatus. Furthermore, the previously characterized Td oxygen sensor ODP influences CheA ordering. These results suggest a greater diversity of the chemotaxis signaling system than previously thought.


Asunto(s)
Membrana Celular/metabolismo , Células Quimiorreceptoras/citología , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Membrana Celular/ultraestructura , Células Quimiorreceptoras/metabolismo , Quimiotaxis , Secuencia Conservada , Escherichia coli/citología , Eliminación de Gen , Histidina Quinasa/metabolismo , Dominios Proteicos , Homología de Secuencia de Aminoácido , Treponema/metabolismo
4.
Nat Commun ; 11(1): 5533, 2020 Oct 27.
Artículo en Inglés | MEDLINE | ID: mdl-33110082

RESUMEN

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

5.
Nat Commun ; 11(1): 5080, 2020 10 08.
Artículo en Inglés | MEDLINE | ID: mdl-33033258

RESUMEN

Natural transformation is the process by which bacteria take up genetic material from their environment and integrate it into their genome by homologous recombination. It represents one mode of horizontal gene transfer and contributes to the spread of traits like antibiotic resistance. In Vibrio cholerae, a type IVa pilus (T4aP) is thought to facilitate natural transformation by extending from the cell surface, binding to exogenous DNA, and retracting to thread this DNA through the outer membrane secretin, PilQ. Here, we use a functional tagged allele of VcPilQ purified from native V. cholerae cells to determine the cryoEM structure of the VcPilQ secretin in amphipol to ~2.7 Å. We use bioinformatics to examine the domain architecture and gene neighborhood of T4aP secretins in Proteobacteria in comparison with VcPilQ. This structure highlights differences in the architecture of the T4aP secretin from the type II and type III secretion system secretins. Based on our cryoEM structure, we design a series of mutants to reversibly regulate VcPilQ gate dynamics. These experiments support the idea of VcPilQ as a potential druggable target and provide insight into the channel that DNA likely traverses to promote the spread of antibiotic resistance via horizontal gene transfer by natural transformation.


Asunto(s)
Sistemas de Secreción Bacterianos/ultraestructura , Microscopía por Crioelectrón , Fimbrias Bacterianas/ultraestructura , Secretina/química , Vibrio cholerae/metabolismo , Vibrio cholerae/ultraestructura , Cisteína/genética , Proteínas de la Membrana/ultraestructura , Modelos Moleculares , Mutación/genética , Filogenia , Dominios Proteicos , Transformación Bacteriana
6.
Proc Natl Acad Sci U S A ; 117(16): 8941-8947, 2020 04 21.
Artículo en Inglés | MEDLINE | ID: mdl-32241888

RESUMEN

The bacterial flagellum is an amazing nanomachine. Understanding how such complex structures arose is crucial to our understanding of cellular evolution. We and others recently reported that in several Gammaproteobacterial species, a relic subcomplex comprising the decorated P and L rings persists in the outer membrane after flagellum disassembly. Imaging nine additional species with cryo-electron tomography, here, we show that this subcomplex persists after flagellum disassembly in other phyla as well. Bioinformatic analyses fail to show evidence of any recent horizontal transfers of the P- and L-ring genes, suggesting that this subcomplex and its persistence is an ancient and conserved feature of the flagellar motor. We hypothesize that one function of the P and L rings is to seal the outer membrane after motor disassembly.


Asunto(s)
Bacterias/genética , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas Bacterianas/genética , Flagelos/genética , Especiación Genética , Bacterias/citología , Bacterias/metabolismo , Membrana Externa Bacteriana/metabolismo , Membrana Externa Bacteriana/ultraestructura , Proteínas de la Membrana Bacteriana Externa/metabolismo , Proteínas Bacterianas/metabolismo , Biología Computacional , Microscopía por Crioelectrón , Tomografía con Microscopio Electrónico , Flagelos/metabolismo , Genes Bacterianos , Filogenia
7.
Nat Commun ; 11(1): 2041, 2020 04 27.
Artículo en Inglés | MEDLINE | ID: mdl-32341341

RESUMEN

How complex, multi-component macromolecular machines evolved remains poorly understood. Here we reveal the evolutionary origins of the chemosensory machinery that controls flagellar motility in Escherichia coli. We first identify ancestral forms still present in Vibrio cholerae, Pseudomonas aeruginosa, Shewanella oneidensis and Methylomicrobium alcaliphilum, characterizing their structures by electron cryotomography and finding evidence that they function in a stress response pathway. Using bioinformatics, we trace the evolution of the system through γ-Proteobacteria, pinpointing key evolutionary events that led to the machine now seen in E. coli. Our results suggest that two ancient chemosensory systems with different inputs and outputs (F6 and F7) existed contemporaneously, with one (F7) ultimately taking over the inputs and outputs of the other (F6), which was subsequently lost.


Asunto(s)
Sustancias Macromoleculares/química , Methylococcaceae/fisiología , Pseudomonas aeruginosa/fisiología , Shewanella/fisiología , Vibrio cholerae/fisiología , Evolución Biológica , Quimiotaxis , Biología Computacional , Tomografía con Microscopio Electrónico , Escherichia coli/fisiología , Proteínas de Escherichia coli , Flagelos/fisiología , Gammaproteobacteria/fisiología , Genoma Bacteriano , Proteínas Quimiotácticas Aceptoras de Metilo/química , Filogenia
8.
Mol Microbiol ; 114(3): 367-376, 2020 09.
Artículo en Inglés | MEDLINE | ID: mdl-32347610

RESUMEN

Vibrio cholerae, the causative agent of the acute diarrheal disease cholera, is able to thrive in diverse habitats such as natural water bodies and inside human hosts. To ensure their survival, these bacteria rely on chemosensory pathways to sense and respond to changing environmental conditions. These pathways constitute a highly sophisticated cellular control system in Bacteria and Archaea. Reflecting the complex life cycle of V. cholerae, this organism has three different chemosensory pathways that together contain over 50 proteins expressed under different environmental conditions. Only one of them is known to control motility, while the function of the other two remains to be discovered. Here, we provide an overview of the chemosensory systems in V. cholerae and the advances toward understanding their structure and function.


Asunto(s)
Proteínas Bacterianas/fisiología , Quimiotaxis , Transducción de Señal , Vibrio cholerae/fisiología , Cólera/microbiología , Humanos , Vibrio cholerae/ultraestructura , Virulencia
9.
Nucleic Acids Res ; 48(D1): D459-D464, 2020 01 08.
Artículo en Inglés | MEDLINE | ID: mdl-31754718

RESUMEN

Bacteria and archaea employ dedicated signal transduction systems that modulate gene expression, second-messenger turnover, quorum sensing, biofilm formation, motility, host-pathogen and beneficial interactions. The updated MiST database provides a comprehensive classification of microbial signal transduction systems. This update is a result of a substantial scaling to accommodate constantly growing microbial genomic data. More than 125 000 genomes, 516 million genes and almost 100 million unique protein sequences are currently stored in the database. For each bacterial and archaeal genome, MiST 3.0 provides a complete signal transduction profile, thus facilitating theoretical and experimental studies on signal transduction and gene regulation. New software infrastructure and distributed pipeline implemented in MiST 3.0 enable regular genome updates based on the NCBI RefSeq database. A novel MiST feature is the integration of unique profile HMMs to link complex chemosensory systems with corresponding chemoreceptors in bacterial and archaeal genomes. The data can be explored online or via RESTful API (freely available at https://mistdb.com).


Asunto(s)
Bases de Datos Genéticas , Genoma Arqueal , Genoma Bacteriano , Transducción de Señal/genética , Programas Informáticos , Regulación de la Expresión Génica Arqueal , Regulación Bacteriana de la Expresión Génica
10.
EMBO J ; 38(14): e100957, 2019 07 15.
Artículo en Inglés | MEDLINE | ID: mdl-31304634

RESUMEN

The self-assembly of cellular macromolecular machines such as the bacterial flagellar motor requires the spatio-temporal synchronization of gene expression with proper protein localization and association of dozens of protein components. In Salmonella and Escherichia coli, a sequential, outward assembly mechanism has been proposed for the flagellar motor starting from the inner membrane, with the addition of each new component stabilizing the previous one. However, very little is known about flagellar disassembly. Here, using electron cryo-tomography and sub-tomogram averaging of intact Legionella pneumophila, Pseudomonas aeruginosa, and Shewanella oneidensis cells, we study flagellar motor disassembly and assembly in situ. We first show that motor disassembly results in stable outer membrane-embedded sub-complexes. These sub-complexes consist of the periplasmic embellished P- and L-rings, and bend the membrane inward while it remains apparently sealed. Additionally, we also observe various intermediates of the assembly process including an inner-membrane sub-complex consisting of the C-ring, MS-ring, and export apparatus. Finally, we show that the L-ring is responsible for reshaping the outer membrane, a crucial step in the flagellar assembly process.


Asunto(s)
Bacterias/citología , Proteínas Bacterianas/metabolismo , Flagelos/ultraestructura , Bacterias/metabolismo , Bacterias/ultraestructura , Membrana Externa Bacteriana/metabolismo , Tomografía con Microscopio Electrónico , Escherichia coli/citología , Escherichia coli/metabolismo , Escherichia coli/ultraestructura , Flagelos/metabolismo , Legionella pneumophila/citología , Legionella pneumophila/metabolismo , Legionella pneumophila/ultraestructura , Pseudomonas aeruginosa/citología , Pseudomonas aeruginosa/metabolismo , Pseudomonas aeruginosa/ultraestructura , Shewanella/citología , Shewanella/metabolismo , Shewanella/ultraestructura
11.
PLoS One ; 14(4): e0215531, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-30986271

RESUMEN

Three-dimensional electron microscopy techniques like electron tomography provide valuable insights into cellular structures, and present significant challenges for data storage and dissemination. Here we explored a novel method to publicly release more than 11,000 such datasets, more than 30 TB in total, collected by our group. Our method, based on a peer-to-peer file sharing network built around a blockchain ledger, offers a distributed solution to data storage. In addition, we offer a user-friendly browser-based interface, https://etdb.caltech.edu, for anyone interested to explore and download our data. We discuss the relative advantages and disadvantages of this system and provide tools for other groups to mine our data and/or use the same approach to share their own imaging datasets.


Asunto(s)
Cadena de Bloques , Redes de Comunicación de Computadores , Bases de Datos Factuales , Tomografía con Microscopio Electrónico , Difusión de la Información , Programas Informáticos , Humanos , Almacenamiento y Recuperación de la Información
12.
Elife ; 82019 01 16.
Artículo en Inglés | MEDLINE | ID: mdl-30648971

RESUMEN

The bacterial flagellar motor, a cell-envelope-embedded macromolecular machine that functions as a cellular propeller, exhibits significant structural variability between species. Different torque-generating stator modules allow motors to operate in different pH, salt or viscosity levels. How such diversity evolved is unknown. Here, we use electron cryo-tomography to determine the in situ macromolecular structures of three Gammaproteobacteria motors: Legionella pneumophila, Pseudomonas aeruginosa, and Shewanella oneidensis, providing the first views of intact motors with dual stator systems. Complementing our imaging with bioinformatics analysis, we find a correlation between the motor's stator system and its structural elaboration. Motors with a single H+-driven stator have only the core periplasmic P- and L-rings; those with dual H+-driven stators have an elaborated P-ring; and motors with Na+ or Na+/H+-driven stators have both their P- and L-rings embellished. Our results suggest an evolution of structural elaboration that may have enabled pathogenic bacteria to colonize higher-viscosity environments in animal hosts.


Asunto(s)
Flagelos/metabolismo , Gammaproteobacteria/metabolismo , Proteínas Motoras Moleculares/química , Periplasma/metabolismo , Flagelos/ultraestructura , Gammaproteobacteria/ultraestructura , Periplasma/ultraestructura , Filogenia , Sodio/metabolismo
13.
Methods Mol Biol ; 1729: 373-385, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-29429105

RESUMEN

Identifying chemoreceptors in sequenced bacterial genomes, revealing their domain architecture, inferring their evolutionary relationships, and comparing them to chemoreceptors of known function become important steps in genome annotation and chemotaxis research. Here, we describe bioinformatics procedures that enable such analyses, using two closely related bacterial genomes as examples.


Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Rhizobium/metabolismo , Sinorhizobium meliloti/metabolismo , Quimiotaxis , Biología Computacional/métodos , Evolución Molecular , Genoma Bacteriano , Filogenia , Rhizobium/genética , Análisis de Secuencia de Proteína , Sinorhizobium meliloti/genética
14.
Proc Natl Acad Sci U S A ; 115(7): E1455-E1464, 2018 02 13.
Artículo en Inglés | MEDLINE | ID: mdl-29348205

RESUMEN

Cell division in many eukaryotes is driven by a ring containing actin and myosin. While much is known about the main proteins involved, the precise arrangement of actin filaments within the contractile machinery, and how force is transmitted to the membrane, remains unclear. Here we use cryosectioning and cryofocused ion beam milling to gain access to cryopreserved actomyosin rings in Schizosaccharomyces pombe for direct 3D imaging by electron cryotomography. Our results show that straight, overlapping actin filaments, running nearly parallel to each other and to the membrane, form a loose bundle of ∼150 nm in diameter that "saddles" the inward-bending membrane at the leading edge of the division septum. The filaments do not make direct contact with the membrane. Our analysis of the actin filaments reveals the variability in filament number, nearest-neighbor distances between filaments within the bundle, their distance from the membrane, and angular distribution with respect to the membrane.


Asunto(s)
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Actomiosina/metabolismo , Citocinesis/fisiología , Proteínas de Schizosaccharomyces pombe/química , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Conformación Proteica , Schizosaccharomyces/crecimiento & desarrollo
15.
Proc Natl Acad Sci U S A ; 114(48): 12809-12814, 2017 11 28.
Artículo en Inglés | MEDLINE | ID: mdl-29133402

RESUMEN

In contrast to Escherichia coli, a model organism for chemotaxis that has 5 chemoreceptors and a single chemosensory pathway, Pseudomonas aeruginosa PAO1 has a much more complex chemosensory network, which consists of 26 chemoreceptors feeding into four chemosensory pathways. While several chemoreceptors were rigorously linked to specific pathways in a series of experimental studies, for most of them this information is not available. Thus, we addressed the problem computationally. Protein-protein interaction network prediction, coexpression data mining, and phylogenetic profiling all produced incomplete and uncertain assignments of chemoreceptors to pathways. However, comparative sequence analysis specifically targeting chemoreceptor regions involved in pathway interactions revealed conserved sequence patterns that enabled us to unambiguously link all 26 chemoreceptors to four pathways. Placing computational evidence in the context of experimental data allowed us to conclude that three chemosensory pathways in P. aeruginosa utilize one chemoreceptor per pathway, whereas the fourth pathway, which is the main system controlling chemotaxis, utilizes the other 23 chemoreceptors. Our results show that while only a very few amino acid positions in receptors, kinases, and adaptors determine their pathway specificity, assigning receptors to pathways computationally is possible. This requires substantial knowledge about interacting partners on a molecular level and focusing comparative sequence analysis on the pathway-specific regions. This general principle should be applicable to resolving many other receptor-pathway interactions.


Asunto(s)
Proteínas Bacterianas/genética , Quimiotaxis/genética , Regulación Bacteriana de la Expresión Génica , Pseudomonas aeruginosa/genética , Receptores de Superficie Celular/genética , Transducción de Señal , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/clasificación , Proteínas Bacterianas/metabolismo , Sitios de Unión , Factores Quimiotácticos/química , Factores Quimiotácticos/metabolismo , Biología Computacional/métodos , Minería de Datos/estadística & datos numéricos , Redes Reguladoras de Genes , Ligandos , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Mapeo de Interacción de Proteínas , Pseudomonas aeruginosa/metabolismo , Receptores de Superficie Celular/química , Receptores de Superficie Celular/clasificación , Receptores de Superficie Celular/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido
16.
EMBO Rep ; 18(7): 1090-1099, 2017 07.
Artículo en Inglés | MEDLINE | ID: mdl-28487352

RESUMEN

The type VI secretion system (T6SS) is a versatile molecular weapon used by many bacteria against eukaryotic hosts or prokaryotic competitors. It consists of a cytoplasmic bacteriophage tail-like structure anchored in the bacterial cell envelope via a cytoplasmic baseplate and a periplasmic membrane complex. Rapid contraction of the sheath in the bacteriophage tail-like structure propels an inner tube/spike complex through the target cell envelope to deliver effectors. While structures of purified contracted sheath and purified membrane complex have been solved, because sheaths contract upon cell lysis and purification, no structure is available for the extended sheath. Structural information about the baseplate is also lacking. Here, we use electron cryotomography to directly visualize intact T6SS structures inside Myxococcus xanthus cells. Using sub-tomogram averaging, we resolve the structure of the extended sheath and membrane-associated components including the baseplate. Moreover, we identify novel extracellular bacteriophage tail fiber-like antennae. These results provide new structural insights into how the extended sheath prevents premature disassembly and how this sophisticated machine may recognize targets.


Asunto(s)
Myxococcus xanthus/ultraestructura , Sistemas de Secreción Tipo VI/ultraestructura , Bacteriófagos/ultraestructura , Microscopía por Crioelectrón/instrumentación , Microscopía por Crioelectrón/métodos , Microscopía Intravital/instrumentación , Microscopía Intravital/métodos , Estructura Molecular , Myxococcus xanthus/química , Myxococcus xanthus/citología , Unión Proteica , Multimerización de Proteína , Sistemas de Secreción Tipo VI/química
17.
Science ; 356(6333): 82-85, 2017 04 07.
Artículo en Inglés | MEDLINE | ID: mdl-28386012

RESUMEN

The discovery of giant viruses blurred the sharp division between viruses and cellular life. Giant virus genomes encode proteins considered as signatures of cellular organisms, particularly translation system components, prompting hypotheses that these viruses derived from a fourth domain of cellular life. Here we report the discovery of a group of giant viruses (Klosneuviruses) in metagenomic data. Compared with other giant viruses, the Klosneuviruses encode an expanded translation machinery, including aminoacyl transfer RNA synthetases with specificities for all 20 amino acids. Notwithstanding the prevalence of translation system components, comprehensive phylogenomic analysis of these genes indicates that Klosneuviruses did not evolve from a cellular ancestor but rather are derived from a much smaller virus through extensive gain of host genes.


Asunto(s)
Aminoacil-ARNt Sintetasas/genética , Genoma Viral , Virus Gigantes/clasificación , Virus Gigantes/genética , Biosíntesis de Proteínas/genética , Aminoacil-ARNt Sintetasas/química , Austria , Virus Gigantes/aislamiento & purificación , Metagenómica , Filogenia , Aguas Residuales/virología
18.
Nat Microbiol ; 2: 16269, 2017 Feb 06.
Artículo en Inglés | MEDLINE | ID: mdl-28165453

RESUMEN

Type IV pili (T4P) are filamentous appendages found on many Bacteria and Archaea. They are helical fibres of pilin proteins assembled by a multi-component macromolecular machine we call the basal body. Based on pilin features, T4P are classified into type IVa pili (T4aP) and type IVb pili (T4bP)1,2. T4aP are more widespread and are involved in cell motility3, DNA transfer4, host predation5 and electron transfer6. T4bP are less prevalent and are mainly found in enteropathogenic bacteria, where they play key roles in host colonization7. Following similar work on T4aP machines8,9, here we use electron cryotomography10 to reveal the three-dimensional in situ structure of a T4bP machine in its piliated and non-piliated states. The specific machine we analyse is the Vibrio cholerae toxin-coregulated pilus machine (TCPM). Although only about half of the components of the TCPM show sequence homology to components of the previously analysed Myxococcus xanthus T4aP machine (T4aPM), we find that their structures are nevertheless remarkably similar. Based on homologies with components of the M. xanthus T4aPM and additional reconstructions of TCPM mutants in which the non-homologous proteins are individually deleted, we propose locations for all eight TCPM components within the complex. Non-homologous proteins in the T4aPM and TCPM are found to form similar structures, suggesting new hypotheses for their functions and evolutionary histories.


Asunto(s)
Proteínas Fimbrias/química , Fimbrias Bacterianas/química , Fimbrias Bacterianas/ultraestructura , Vibrio cholerae/ultraestructura , Adhesión Bacteriana , Proteínas Bacterianas/análisis , Proteínas Bacterianas/química , Toxina del Cólera/metabolismo , Tomografía con Microscopio Electrónico/métodos , Proteínas Fimbrias/análisis , Fimbrias Bacterianas/genética , Modelos Moleculares , Mutación , Myxococcus xanthus/química , Myxococcus xanthus/ultraestructura , Vibrio cholerae/química
19.
Proc Natl Acad Sci U S A ; 113(37): 10412-7, 2016 09 13.
Artículo en Inglés | MEDLINE | ID: mdl-27573843

RESUMEN

Nearly all motile bacterial cells use a highly sensitive and adaptable sensory system to detect changes in nutrient concentrations in the environment and guide their movements toward attractants and away from repellents. The best-studied bacterial chemoreceptor arrays are membrane-bound. Many motile bacteria contain one or more additional, sometimes purely cytoplasmic, chemoreceptor systems. Vibrio cholerae contains three chemotaxis clusters (I, II, and III). Here, using electron cryotomography, we explore V. cholerae's cytoplasmic chemoreceptor array and establish that it is formed by proteins from cluster I. We further identify a chemoreceptor with an unusual domain architecture, DosM, which is essential for formation of the cytoplasmic arrays. DosM contains two signaling domains and spans the two-layered cytoplasmic arrays. Finally, we present evidence suggesting that this type of receptor is important for the structural stability of the cytoplasmic array.


Asunto(s)
Proteínas Bacterianas/metabolismo , Células Quimiorreceptoras/metabolismo , Quimiotaxis/genética , Citoplasma/metabolismo , Vibrio cholerae/metabolismo , Proteínas Bacterianas/química , Células Quimiorreceptoras/química , Microscopía por Crioelectrón , Citoplasma/química , Dominios Proteicos , Tomografía , Vibrio cholerae/genética , Vibrio cholerae/patogenicidad
20.
Nat Rev Microbiol ; 14(9): 600, 2016 09.
Artículo en Inglés | MEDLINE | ID: mdl-27452232
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