RESUMEN
MreB is a bacterial actin that is important for cell shape and cell wall biosynthesis in many bacterial species. MreB also plays crucial roles in Myxococcus xanthus gliding motility, but the underlying mechanism remains unknown. Here we tracked the dynamics of single MreB particles in M. xanthus using single-particle tracking photoactivated localization microscopy. We found that a subpopulation of MreB particles moves rapidly along helical trajectories, similar to the movements of the MotAB-like gliding motors. The rapid MreB motion was stalled in the mutants that carried truncated gliding motors. Remarkably, M. xanthus MreB moves one to two orders of magnitude faster than its homologs that move along with the cell wall synthesis machinery in Bacillus subtilis and Escherichia coli, and this rapid movement was not affected by the inhibitors of cell wall biosynthesis. Our results show that in M. xanthus, MreB provides a scaffold for the gliding motors while the gliding machinery drives the movement of MreB filaments, analogous to the interdependent movements of myosin motors and actin in eukaryotic cells.
Asunto(s)
Actinas/metabolismo , Proteínas Bacterianas/metabolismo , Movimiento Celular/fisiología , Myxococcus xanthus/metabolismo , Myxococcus xanthus/fisiología , Actinas/química , Actinas/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Luminiscentes/química , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Fluorescente , Mutación , Myxococcus xanthus/química , Myxococcus xanthus/genética , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteína Fluorescente RojaRESUMEN
Bacterial gliding motility is the smooth movement of cells on solid surfaces unaided by flagella or pili. Many diverse groups of bacteria exhibit gliding, but the mechanism of gliding motility has remained a mystery since it was first observed more than a century ago. Recent studies on the motility of Myxococcus xanthus, a soil myxobacterium, suggest a likely mechanism for gliding in this organism. About forty M. xanthus genes were shown to be involved in gliding motility, and some of their protein products were labeled and localized within cells. These studies suggest that gliding motility in M. xanthus involves large multiprotein structural complexes, regulatory proteins, and cytoskeletal filaments. In this review, we summarize recent experiments that provide the basis for this emerging view of M. xanthus motility. We also discuss alternative models for gliding.
Asunto(s)
Flagelos/fisiología , Genes Bacterianos , Myxococcus xanthus/fisiología , Adenosina Trifosfato/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Flagelos/metabolismo , Adhesiones Focales/metabolismo , Adhesiones Focales/fisiología , Modelos Biológicos , Proteínas Motoras Moleculares/genética , Proteínas Motoras Moleculares/metabolismo , Movimiento , Complejos Multiproteicos/metabolismo , Myxococcus xanthus/genética , Myxococcus xanthus/metabolismo , Vías SecretorasRESUMEN
Gliding motility in Myxococcus xanthus is powered by flagella stator homologs that move in helical trajectories using proton motive force. The Frz chemosensory pathway regulates the cell polarity axis through MglA, a Ras family GTPase; however, little is known about how MglA establishes the polarity of gliding, because the gliding motors move simultaneously in opposite directions. Here we examined the localization and dynamics of MglA and gliding motors in high spatial and time resolution. We determined that MglA localizes not only at the cell poles, but also along the cell bodies, forming a decreasing concentration gradient toward the lagging cell pole. MglA directly interacts with the motor protein AglR, and the spatial distribution of AglR reversals is positively correlated with the MglA gradient. Thus, the motors moving toward lagging cell poles are less likely to reverse, generating stronger forward propulsion. MglB, the GTPase-activating protein of MglA, regulates motor reversal by maintaining the MglA gradient. Our results suggest a mechanism whereby bacteria use Ras family proteins to modulate cellular polarity.
Asunto(s)
Proteínas Bacterianas/fisiología , Proteínas Motoras Moleculares/fisiología , Myxococcus xanthus/fisiología , Proteínas Bacterianas/genética , Cuerpo Celular/fisiología , Polaridad Celular/fisiología , Microscopía Fluorescente , Modelos Biológicos , Proteínas Motoras Moleculares/genética , Movimiento/fisiología , Myxococcus xanthus/citología , Myxococcus xanthus/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteínas ras/genética , Proteínas ras/fisiologíaRESUMEN
For many bacteria, motility is essential for survival, growth, virulence, biofilm formation and intra/interspecies interactions. Since natural environments differ, bacteria have evolved remarkable motility systems to adapt, including swimming in aqueous media, and swarming, twitching and gliding on solid and semi-solid surfaces. Although tremendous advances have been achieved in understanding swimming and swarming motilities powered by flagella, and twitching motility powered by Type IV pili, little is known about gliding motility. Bacterial gliders are a heterogeneous group containing diverse bacteria that utilize surface motilities that do not depend on traditional flagella or pili, but are powered by mechanisms that are less well understood. Recently, advances in our understanding of the molecular machineries for several gliding bacteria revealed the roles of modified ion channels, secretion systems and unique machinery for surface movements. These novel mechanisms provide rich source materials for studying the function and evolution of complex microbial nanomachines. In this review, we summarize recent findings made on the gliding mechanisms of the myxobacteria, flavobacteria and mycoplasmas.
Asunto(s)
Movimiento Celular/fisiología , Movimiento/fisiología , Movimiento Celular/genética , Flavobacteriaceae/metabolismo , Modelos Biológicos , Mycoplasma/metabolismo , Myxococcales/metabolismo , Vías Secretoras/genética , Vías Secretoras/fisiología , Virulencia/fisiologíaRESUMEN
The Frz pathway of Myxococcus xanthus controls cell reversal frequency to support directional motility during swarming and fruiting body formation. Previously, we showed that phosphorylation of the response regulator FrzZ correlates with reversal frequencies, suggesting that this activity represents the output of the Frz pathway. Here, we tested the effect of different expression levels of FrzZ and its cognate kinase FrzE on M. xanthus motility. FrzZ overexpression caused a slight increase in phosphorylation and reversals. By contrast, FrzE overexpression abolished phosphorylation of FrzZ; this inhibition required the response regulator domain of FrzE. FrzZ phosphorylation was restored when both FrzE and FrzZ were overexpressed together. Our results show that the response regulator domain of FrzE is a negative regulator of FrzE kinase activity. This inhibition can be modulated by FrzZ, which acts as a positive regulator. Interestingly, fluorescence microscopy revealed that FrzZ and FrzE localize differently: FrzE colocalizes with the FrzCD receptor and the nucleoid, while FrzZ shows dispersed and polar localization. However, FrzZ binds tightly to the truncated variant FrzEΔ(CheY) . This indicates that the response regulator domain of FrzE is required for the interaction between FrzE and FrzZ to be transient, providing an unexpected regulatory output to the Frz pathway.
Asunto(s)
Proteínas Bacterianas/metabolismo , Myxococcus xanthus/metabolismo , Proteínas Bacterianas/genética , Quimiotaxis/fisiología , Myxococcus xanthus/genética , Fenotipo , Fosforilación , Transducción de SeñalRESUMEN
Chemosensory systems (CSS) are complex regulatory pathways capable of perceiving external signals and translating them into different cellular behaviors such as motility and development. In the δ-proteobacterium Myxococcus xanthus, chemosensing allows groups of cells to orient themselves and aggregate into specialized multicellular biofilms termed fruiting bodies. M. xanthus contains eight predicted CSS and 21 chemoreceptors. In this work, we systematically deleted genes encoding components of each CSS and chemoreceptors and determined their effects on M. xanthus social behaviors. Then, to understand how the 21 chemoreceptors are distributed among the eight CSS, we examined their phylogenetic distribution, genomic organization and subcellular localization. We found that, in vivo, receptors belonging to the same phylogenetic group colocalize and interact with CSS components of the respective phylogenetic group. Finally, we identified a large chemosensory module formed by three interconnected CSS and multiple chemoreceptors and showed that complex behaviors such as cell group motility and biofilm formation require regulatory apparatus composed of multiple interconnected Che-like systems.
Asunto(s)
Quimiotaxis/genética , Regulación Bacteriana de la Expresión Génica , Myxococcus xanthus/genética , Transducción de Señal/genética , Biopelículas/crecimiento & desarrollo , Movimiento Celular/genética , Movimiento , Myxococcus xanthus/química , Myxococcus xanthus/crecimiento & desarrollo , FilogeniaRESUMEN
Many bacterial species use gliding motility in natural habitats because external flagella function poorly on hard surfaces. However, the mechanism(s) of gliding remain elusive because surface motility structures are not apparent. Here, we characterized the dynamics of the Myxococcus xanthus gliding motor protein AglR, a homolog of the Escherichia coli flagella stator protein MotA. We observed that AglR decorated a helical structure, and the AglR helices rotated when cells were suspended in liquid or when cells moved on agar surfaces. With photoactivatable localization microscopy, we found that single molecules of AglR, unlike MotA/MotB, can move laterally within the membrane in helical trajectories. AglR slowed down transiently at gliding surfaces, accumulating in clusters. Our work shows that the untethered gliding motors of M. xanthus, by moving within the membrane, can transform helical motion into linear driving forces that push against the surface.
Asunto(s)
Proteínas Bacterianas/metabolismo , Membrana Celular/metabolismo , Modelos Biológicos , Myxococcus xanthus/fisiología , Rayos Láser , Microscopía Fluorescente/métodos , Simulación de Dinámica Molecular , Movimiento/fisiología , Especificidad de la EspecieRESUMEN
The life cycle of Myxococcus xanthus includes co-ordinated group movement and fruiting body formation, and requires directed motility and controlled cell reversals. Reversals are achieved by inverting cell polarity and re-organizing many motility proteins. The Frz chemosensory pathway regulates the frequency of cell reversals. While it has been established that phosphotransfer from the kinase FrzE to the response regulator FrzZ is required, it is unknown how phosphorylated FrzZ, the putative output of the pathway, targets the cell polarity axis. In this study, we used Phos-tag SDS-PAGE to determine the cellular level of phospho-FrzZ under different growth conditions and in Frz signalling mutants. We detected consistent FrzZ phosphorylation, albeit with a short half-life, in cells grown on plates, but not from liquid culture. The available pool of phospho-FrzZ correlated with reversal frequencies, with higher levels found in hyper-reversing mutants. Phosphorylation was not detected in hypo-reversing mutants. Fluorescence microscopy revealed that FrzZ is recruited to the leading cell pole upon phosphorylation and switches to the opposite pole during reversals. These results are consistent with the hypothesis that the Frz pathway modulates reversal frequency through a localized response regulator that targets cell polarity regulators at the leading cell pole.
Asunto(s)
Proteínas Bacterianas/metabolismo , Polaridad Celular , Locomoción , Myxococcus xanthus/fisiología , Procesamiento Proteico-Postraduccional , Transducción de Señal , Electroforesis en Gel de Poliacrilamida , Perfilación de la Expresión Génica , Microscopía Fluorescente , Modelos Biológicos , FosforilaciónRESUMEN
Gliding motility in the bacterium Myxococcus xanthus uses two motility engines: S-motility powered by type-IV pili and A-motility powered by uncharacterized motor proteins and focal adhesion complexes. In this paper, we identified MreB, an actin-like protein, and MglA, a small GTPase of the Ras superfamily, as essential for both motility systems. A22, an inhibitor of MreB cytoskeleton assembly, reversibly inhibited S- and A-motility, causing rapid dispersal of S- and A-motility protein clusters, FrzS and AglZ. This suggests that the MreB cytoskeleton is involved in directing the positioning of these proteins. We also found that a DeltamglA motility mutant showed defective localization of AglZ and FrzS clusters. Interestingly, MglA-YFP localization mimicked both FrzS and AglZ patterns and was perturbed by A22 treatment, consistent with results indicating that both MglA and MreB bind to motility complexes. We propose that MglA and the MreB cytoskeleton act together in a pathway to localize motility proteins such as AglZ and FrzS to assemble the A-motility machineries. Interestingly, M. xanthus motility systems, like eukaryotic systems, use an actin-like protein and a small GTPase spatial regulator.
Asunto(s)
Adhesión Bacteriana , Proteínas Bacterianas/metabolismo , Proteínas Motoras Moleculares/metabolismo , Myxococcus xanthus/citología , Proteínas Bacterianas/análisis , Proteínas Bacterianas/genética , Polaridad Celular , Citoesqueleto/metabolismo , Proteínas Motoras Moleculares/análisis , Mutación , Myxococcus xanthus/metabolismo , Tiourea/análogos & derivados , Tiourea/farmacologíaRESUMEN
Myxococcus xanthus is a Gram-negative bacterium that glides over surfaces without the aid of flagella. Two motility systems are used for locomotion: social-motility, powered by the retraction of type IV pili, and adventurous (A)-motility, powered by unknown mechanism(s). We have shown that AgmU, an A-motility protein, is part of a multiprotein complex that spans the inner membrane and periplasm of M. xanthus. In this paper, we present evidence that periplasmic AgmU decorates a looped continuous helix that rotates clockwise as cells glide forward, reversing its rotation when cells reverse polarity. Inhibitor studies showed that the AgmU helix rotation is driven by proton motive force (PMF) and depends on actin-like MreB cytoskeletal filaments. The AgmU motility complex was found to interact with MotAB homologs. Our data are consistent with a mechanochemical model in which PMF-driven motors, similar to bacterial flagella stator complexes, run along an endless looped helical track, driving rotation of the track; deformation of the cell surface by the AgmU-associated proteins creates pressure waves in the slime, pushing cells forward.
Asunto(s)
Citoesqueleto/metabolismo , Fimbrias Bacterianas/metabolismo , Modelos Biológicos , Myxococcus xanthus/metabolismo , Proteínas Periplasmáticas/metabolismo , Fuerza Protón-Motriz/fisiología , Citoesqueleto/genética , Fimbrias Bacterianas/genética , Myxococcus xanthus/citología , Myxococcus xanthus/genética , Proteínas Periplasmáticas/genéticaRESUMEN
Directional motility in the gliding bacterium Myxococcus xanthus requires controlled cell reversals mediated by the Frz chemosensory system. FrzCD, a cytoplasmic chemoreceptor, does not form membrane-bound polar clusters typical for most bacteria, but rather cytoplasmic clusters that appear helically arranged and span the cell length. The distribution of FrzCD in living cells was found to be dynamic: FrzCD was localized in clusters that continuously changed their size, number, and position. The number of FrzCD clusters was correlated with cellular reversal frequency: fewer clusters were observed in hypo-reversing mutants and additional clusters were observed in hyper-reversing mutants. When moving cells made side-to-side contacts, FrzCD clusters in adjacent cells showed transient alignments. These events were frequently followed by one of the interacting cells reversing. These observations suggest that FrzCD detects signals from a cell contact-sensitive signaling system and then re-localizes as it directs reversals to distributed motility engines.
Asunto(s)
Proteínas Bacterianas/metabolismo , Citoplasma/metabolismo , Myxococcus xanthus/citología , Myxococcus xanthus/metabolismo , Receptores de Superficie Celular/metabolismo , Proteínas Bacterianas/química , Proteínas Fluorescentes Verdes/metabolismo , Estructura Secundaria de Proteína , Transporte de Proteínas , Proteínas Recombinantes de Fusión/metabolismoRESUMEN
Myxococcus xanthus moves by gliding motility powered by Type IV pili (S-motility) and a second motility system, A-motility, whose mechanism remains elusive despite the identification of approximately 40 A-motility genes. In this study, we used biochemistry and cell biology analyses to identify multi-protein complexes associated with A-motility. Previously, we showed that the N-terminal domain of FrzCD, the receptor for the frizzy chemosensory pathway, interacts with two A-motility proteins, AglZ and AgmU. Here we characterized AgmU, a protein that localized to both the periplasm and cytoplasm. On firm surfaces, AgmU-mCherry colocalized with AglZ as distributed clusters that remained fixed with respect to the substratum as cells moved forward. Cluster formation was favoured by hard surfaces where A-motility is favoured. In contrast, AgmU-mCherry clusters were not observed on soft agar surfaces or when cells were in large groups, conditions that favour S-motility. Using glutathione-S-transferase affinity chromatography, AgmU was found to interact either directly or indirectly with multiple A-motility proteins including AglZ, AglT, AgmK, AgmX, AglW and CglB. These proteins, important for the correct localization of AgmU and AglZ, appear to be organized as a motility complex, spanning the cytoplasm, inner membrane and the periplasm. Identification of this complex may be important for uncovering the mechanism of A-motility.
Asunto(s)
Proteínas Bacterianas/fisiología , Locomoción , Myxococcus xanthus/fisiología , Proteínas Bacterianas/análisis , Citoplasma/química , Modelos Biológicos , Modelos Químicos , Myxococcus xanthus/química , Periplasma/química , Unión Proteica , Mapeo de Interacción de ProteínasRESUMEN
Myxococcus xanthus moves by gliding motility powered by type IV pili (S-motility) and distributed motor complexes (A-motility). The Frz chemosensory pathway controls reversals for both motility systems. However, it is unclear how the Frz pathway can communicate with these different systems. In this article, we show that FrzCD, the Frz pathway receptor, interacts with AglZ, a protein associated with A-motility. Affinity chromatography and cross-linking experiments showed that the FrzCD-AglZ interaction occurs between the uncharacterized N-terminal region of FrzCD and the N-terminal pseudo-receiver domain of AglZ. Fluorescence microscopy showed AglZ-mCherry and FrzCD-GFP localized in clusters that occupy different positions in cells. To study the role of the Frz system in the regulation of A-motility, we constructed aglZ frzCD double mutants and aglZ frzCD pilA triple mutants. To our surprise, these mutants, predicted to show no A-motility (A-S+) or no motility at all (A-S-), respectively, showed restored A-motility. These results indicate that AglZ modulates a FrzCD activity that inhibits A-motility. We hypothesize that AglZ-FrzCD interactions are favoured when cells are isolated and moving by A-motility and inhibited when S-motility predominates and A-motility is reduced.
Asunto(s)
Proteínas Bacterianas/metabolismo , Myxococcus xanthus/citología , Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión Génica , Movimiento , Mutagénesis , Myxococcus xanthus/genética , Myxococcus xanthus/metabolismo , Dominios y Motivos de Interacción de ProteínasRESUMEN
Within the same human gastrointestinal tract, substantial differences in the bacterial species that inhabit oral cavity and intestinal tract have been noted. Previous research primarily attributed the differences to the influences of host environments and nutritional availabilities ("host habitat" effect). Our recent study indicated that, other than the host habitat effect, an existing microbial community could impose a selective pressure on incoming foreign bacterial species independent of host-mediated selection ("community selection" effect). In this study, we employed in vitro microbial floras representing microorganisms that inhabit the oral cavities and intestinal tract of mice in combination with Escherichia coli as a model intestinal bacterium and demonstrated that E. coli displays a striking community preference. It thrived when introduced into the intestinal microbial community and survived poorly in the microbial flora of foreign origin (oral community). A more detailed examination of this phenomenon showed that the oral community produced oxygen-free radicals in the presence of wild-type E. coli while mutants deficient in lipopolysaccharides (LPS) did not trigger significant production of these cell-damaging agents. Furthermore, mutants of E. coli defective in the oxidative stress response experienced a more drastic reduction in viability when cocultivated with the oral flora, while the exogenous addition of the antioxidant vitamin C was able to rescue it. We concluded that the oral-derived microbial community senses the E. coli LPS and kills the bacterium with oxygen-free radicals. This study reveals a new mechanism of community invasion resistance employed by established microflora to defend their domains.
Asunto(s)
Antibiosis , Escherichia coli/fisiología , Intestinos/microbiología , Boca/microbiología , Animales , Técnicas de Cocultivo , Escherichia coli/crecimiento & desarrollo , Peróxido de Hidrógeno/metabolismo , Lipopolisacáridos/biosíntesis , Ratones , Estrés OxidativoRESUMEN
The gastrointestinal (GI) tract is home to trillions of microbes. Within the same GI tract, substantial differences in the bacterial species that inhabit the oral cavity and intestinal tract have been noted. While the influence of host environments and nutritional availability in shaping different microbial communities is widely accepted, we hypothesize that the existing microbial flora also plays a role in selecting the bacterial species that are being integrated into the community. In this study, we used cultivable microbial communities isolated from different parts of the GI tract of mice (oral cavity and intestines) as a model system to examine this hypothesis. Microbes from these two areas were harvested and cultured using the same nutritional conditions, which led to two distinct microbial communities, each with about 20 different species as revealed by PCR-based denaturing gradient gel electrophoresis analysis. In vitro community competition assays showed that the two microbial floras exhibited antagonistic interactions toward each other. More interestingly, all the original isolates tested and their closely related species displayed striking community preferences: They persisted when introduced into the bacterial community of the same origin, while their viable count declined more than three orders of magnitude after 4 days of coincubation with the microbial flora of foreign origin. These results suggest that an existing microbial community might impose a selective pressure on incoming foreign bacterial species independent of host selection. The observed inter-flora interactions could contribute to the protective effect of established microbial communities against the integration of foreign bacteria to maintain the stability of the existing communities.
Asunto(s)
Antibiosis , Bacterias/crecimiento & desarrollo , Tracto Gastrointestinal/microbiología , Boca/microbiología , Animales , ADN Bacteriano/genética , Electroforesis en Gel de Gradiente Desnaturalizante , Masculino , Ratones , Ratones Endogámicos C57BL , Análisis de Secuencia de ADNRESUMEN
The Frz chemosensory system is a two-component signal transduction pathway that controls cell reversals and directional movements for the two motility systems in Myxococcus xanthus. To trigger cell reversals, FrzE, a hybrid CheA-CheY fusion protein, autophosphorylates the kinase domain at His-49, and phosphoryl groups are transferred to aspartate residues (Asp-52 and Asp-220) in the two receiver domains of FrzZ, a dual CheY-like protein that serves as the pathway output. The role of the receiver domain of FrzE was unknown. In this paper, we characterize the FrzE protein in vitro and show that the receiver domain of FrzE negatively regulates the autophosphorylation activity of the kinase domain of FrzE. Unexpectedly, it does not appear to play a direct role in phospho-relay as in most other histidine kinase receiver domain hybrid systems. The regulatory role of the FrzE receiver domain suggests that it may interact with or be phosphorylated by an unknown protein. We also show the dynamics of motility system-specific marker proteins in FrzE mutants as cells move forward and reverse. Our studies indicate that the two motility systems are functionally co-ordinated and that any system-specific branching of the pathway most likely occurs downstream of FrzE.
Asunto(s)
Proteínas Bacterianas/metabolismo , Movimiento Celular , Myxococcus xanthus/fisiología , Transducción de Señal , Proteínas Bacterianas/genética , Quimiotaxis/genética , Quimiotaxis/fisiología , Regulación Bacteriana de la Expresión Génica , Histidina Quinasa , Proteínas de la Membrana , Proteínas Quimiotácticas Aceptoras de Metilo , Myxococcus xanthus/genética , Myxococcus xanthus/crecimiento & desarrollo , Myxococcus xanthus/metabolismo , Proteínas Quinasas/fisiología , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismoRESUMEN
Myxococcus xanthus is a gliding bacterium with a complex life cycle that includes swarming, predation and fruiting body formation. Directed movements in M. xanthus are regulated by the Frz chemosensory system, which controls cell reversals. The Frz pathway requires the activity of FrzCD, a cytoplasmic methyl-accepting chemotaxis protein, and FrzF, a methyltransferase (CheR) containing an additional domain with three tetra trico-peptide repeats (TPRs). To investigate the role of the TPRs in FrzCD methylation, we used full-length FrzF and FrzF lacking its TPRs (FrzF(CheR)) to methylate FrzCD in vitro. FrzF methylated FrzCD on a single residue, E182, while FrzF(CheR) methylated FrzCD on three residues, E168, E175 and E182, indicating that the TPRs regulate site-specific methylation. E168 and E182 were predicted consensus methylation sites, but E175 is methylated on an HE pair. To determine the roles of these sites in vivo, we substituted each methylatable glutamate with either an aspartate or an alanine residue and determined the impact of the point mutants on single cell reversals, swarming and fruiting body formation. Single, double and triple methylation site mutants revealed that each site played a unique role in M. xanthus behaviour and that the pattern of receptor methylation determined receptor activity. This work also shows that methylation can both activate and inactivate the receptor.
Asunto(s)
Proteínas Bacterianas/metabolismo , Metiltransferasas/química , Myxococcus xanthus/química , Myxococcus xanthus/metabolismo , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Proteínas Bacterianas/genética , Metilación , Metiltransferasas/genética , Metiltransferasas/metabolismo , Datos de Secuencia Molecular , Myxococcus xanthus/genética , Mutación Puntual , Estructura Terciaria de Proteína , Alineación de SecuenciaRESUMEN
Myxococcus xanthus is a gliding bacterium that contains two motility systems: S-motility, powered by polar type IV pili, and A-motility, powered by uncharacterized motors and adhesion complexes. The localization and coordination of the two motility engines is essential for directed motility as cells move forward and reverse. During cell reversals, the polarity and localization of motility proteins are rapidly inverted, rendering this system a fascinating example of dynamic protein localization.
Asunto(s)
Proteínas Bacterianas/metabolismo , Polaridad Celular/fisiología , Regulación Bacteriana de la Expresión Génica , Movimiento , Myxococcus xanthus/fisiología , Proteínas Bacterianas/genética , Myxococcus xanthus/genética , Myxococcus xanthus/crecimiento & desarrollo , Myxococcus xanthus/metabolismo , Transducción de SeñalRESUMEN
Myxococcus xanthus undergoes a complex starvation-induced developmental program that results in cells forming multicellular fruiting bodies by aggregating into mounds and then differentiating into spores. This developmental program requires at least 72 h and is mediated by a temporal cascade of gene regulators in response to intra- and extracellular signals. espA mutants, encoding an orphan hybrid histidine kinase, alter the timing of this developmental program, greatly accelerating developmental progression. Here, we characterized EspA and demonstrated that it autophosphorylates in vitro on the conserved histidine residue and then transfers the phosphoryl group to the conserved aspartate residue in the associated receiver domain. The conserved histidine and aspartate residues were both required for EspA function in vivo. Analysis of developmental gene expression and protein accumulation in espA mutants indicated that the expression of the A-signal-dependent spi gene was not affected but that the MrpC transcriptional regulator accumulated earlier, resulting in earlier expression of its target, the FruA transcriptional regulator. Early expression of FruA correlated with acceleration of both the aggregation and sporulation branches of the developmental program, as monitored by early methylation of the FrzCD chemosensory receptor and early expression of the sporulation-specific dev and Mxan_3227 (Omega7536) genes. These results show that EspA plays a key role in the timing of expression of genes necessary for progression of cells through the developmental program.
Asunto(s)
Proteínas Bacterianas/metabolismo , Myxococcus xanthus/enzimología , Proteínas Quinasas/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/fisiología , Regulación Bacteriana de la Expresión Génica , Histidina Quinasa , Immunoblotting , Modelos Biológicos , Myxococcus xanthus/genética , Myxococcus xanthus/metabolismo , Fenotipo , Fosforilación , Reacción en Cadena de la Polimerasa , Proteínas Quinasas/genética , Transducción de Señal/genética , Transducción de Señal/fisiologíaRESUMEN
The myxobacteria are a family of soil bacteria that form biofilms of complex architecture, aligned multilayered swarms or fruiting body structures that are simple or branched aggregates containing myxospores. Here, we examined the structural role of matrix exopolysaccharide (EPS) in the organization of these surface-dwelling bacterial cells. Using time-lapse light and fluorescence microscopy, as well as transmission electron microscopy and focused ion beam/scanning electron microscopy (FIB/SEM) electron microscopy, we found that Myxococcus xanthus cell organization in biofilms is dependent on the formation of EPS microchannels. Cells are highly organized within the three-dimensional structure of EPS microchannels that are required for cell alignment and advancement on surfaces. Mutants lacking EPS showed a lack of cell orientation and poor colony migration. Purified, cell-free EPS retains a channel-like structure, and can complement EPS- mutant motility defects. In addition, EPS provides the cooperative structure for fruiting body formation in both the simple mounds of M. xanthus and the complex, tree-like structures of Chondromyces crocatus. We furthermore investigated the possibility that EPS impacts community structure as a shared resource facilitating cooperative migration among closely related isolates of M. xanthus.