Study of elastic collisions of Myxococcus xanthus in swarms

In very low density situations where a single myxobacterial cell is isolated from direct contact with other cells, the slime capsule interaction with the substrate or slime tracks on the substrate produce a viscous drag that results in a smooth gliding motion. Viscoelastic interactions of myxobacteria cells in a low-density domain close to the edge of a swarm are studied using a combination of a cell-based three-dimensional computational model and cell-tracking experiments. The model takes into account the flexible nature of Myxococcus xanthus as well as the effects of adhesion between cells arising from the interaction of the capsular polysaccharide covering two cells in contact with each other. New image and dynamic cell curvature analysis algorithms are used to track and measure the change in cell shapes that occur as flexible cells undergo significant bending during collisions resulting in direct calibration of the model parameters. Like aspect-ratio and directional reversals, the flexibility of cells and the adhesive cell-cell and cell-substrate interactions of M. xanthus play an important role in smooth gliding and more efficient swarming.     

Self-organized and highly ordered domain structures within swarms of Myxococcus xanthus

Coordinated group movement (swarming) is a key aspect of Myxococcus xanthus' social behavior. Here we report observation of domain structures formed by multiple cells within large three-dimensional swarming groups grown on amorphous glass substrates, using the atomic force microscope (AFM). Novel analyses revealed that 90% of the wild type swarms displayed some form of preferential cell alignment. In contrast, cells with mutations in the social and adventurous motility systems displayed a distinct lack of cell alignment. Video microscopy observations of domain features of in vivo swarming M. xanthus cells were also consistent with the AFM data. The results presented here reveal that unique domain formation within swarms of wild type cells is a biologically driven process requiring the social and adventurous motility systems and is not a statistical phenomenon or thermodynamic process arising from liquid crystal behavior.     

Phosphate acquisition components of the Myxococcus xanthus Pho regulon are regulated by both phosphate availability and development

In many organisms, phosphatase expression and phosphate (P) uptake are coordinately regulated by the Pho regulon. In Myxococcus xanthus P limitation initiates multicellular development, a process associated with changes in phosphatase expression. We sought here to characterize the link between P acquisition and development in this bacterium, an organism capable of preying upon other microorganisms as a sole nutrient source. M. xanthus seems to possess no significant internal P stores, as reducing the P concentration to less than 10 μM retarded growth within one doubling time. Pyrophosphate, polyphosphate, and glyceraldehyde-3-phosphate could support growth as sole P sources, although many other P-containing biomolecules could not (including nucleic acids and phospholipids). Several Pho regulon promoters were found to be highly active during vegetative growth, and P limitation specifically induced pstSCAB, AcPA1, and pho3 promoter activity and repressed pit expression. Enhanced pstSCAB and pho3 promoter activities in a phoP4 mutant (in the presence of high and low concentrations of P) suggested that PhoP4 acts as a repressor of these genes. However, in a phoP4 background, the activities of pstSCAB remained P regulated, suggesting that there is additional regulation by a P-sensitive factor. Initiation of multicellular development caused immediate down-regulation of Pho regulon genes and caused pstSCAB and pho3 promoter activities to become P insensitive. Hence, P acquisition components of the M. xanthus Pho regulon are regulated by both P availability and development, with developmental down-regulation overriding up-regulation by P limitation. These observations suggest that when development is initiated, subsequent changes in P availability become irrelevant to the population, which presumably has sufficient intrinsic P to ensure completion of the developmental program.     

Autocides produced by Myxococcus xanthus.

Ethanol extracts of Myxococcus xanthus contained several substances, referred to as autocides, which were bactericidal to the producing strain but showed no activity against other bacteria. The autocides were produced by growing cells and remained largely cell bound throughout the growth cycle; ca. 5% of the autocidal activity was found in the supernatant fluid at the time cell lysis began. The autocides were separated by sequential-column and thin-layer chromatography into five active fractions (AM I through AM V). Each of the fractions was at least 20 times more active against M. xanthus than against the other gram-negative or gram-positive bacteria tested. AM I, AM IV, and AM V were inactive against yeasts, whereas a mixture of fractions AM II and AM III was active against Rhodotorula sp. At low concentrations, AM I reversibly inhibited the growth of M. xanthus; at higher concentrations of AM I, the cells lysed within 1 h. The lowest concentration of AM IV that showed any activity caused rapid cell death and lysis. The mode of action of the major autocide, AM V, was different from that of AM I and AM IV. During the initial 2 h of treatment, the viable count of M. xanthus cells remained constant; during the next few hours killing occurred without lysis; within 24 h lysis was complete. The autocidal activity of each of the fractions was expressed when the cells were suspended in buffer, as well as in growth medium. The possible role of autocides in developmental lysis of M. xanthus is discussed.     

Bacteriolytic enzymes produced by Myxococcus xanthus

The bacteriolytic activities in the culture fluid of Myxococcus xanthus were purified and separated into six active fractions by the use of Bio-Gel CM-2 and Bio-Gel P-60. These fractions were identified as: (i) an amidase, (ii) a glucosaminidase, (iii) a glucosaminidase and an amidase, (iv) a protease with probable amidase activity, (v) another protease with probable amidase activity, and (vi) a peptidase active on both d-alanyl-diaminopimelate and d-alanyl-lysine peptide bonds. On one occasion, another amidase was eluted from Bio-Gel CM. Preliminary studies on some characteristics of the enzymes and their production during growth are reported.     

Immunomodulation by myxospores of Myxococcus xanthus

Glycerol-induced myxospores of Myxococcus xanthus caused non-specific modulation of humoral and cellular immune responses in laboratory animals. The number of cells which formed specific haemolysins in spleens of mice immunized with sheep erythrocytes was increased when 0.5 X 10(8) myxospores were inoculated 2 d after the erythrocytes, and decreased when myxospores were injected 2 d before or at the same time as the erythrocytes. Both the IgG primary response and the secondary response to erythrocytes were decreased in rabbits after pretreatment with 2 X 10(8) myxospores per rabbit. Delayed-type hypersensitivity to sheep erythrocytes was also suppressed in mice after intraperitoneal (i.p.) injection of 0.3 X 10(8) myxospores. One day after i.p. injection of myxospores, neither an inflammatory response nor bone marrow cell depletion was observed in mice. These results support the idea that M. xanthus myxospores possess diverse immunomodulation properties apparently due to factors different from the classical LPS of Gram-negative bacteria.     

Aggregation during fruiting body formation in Myxococcus xanthus is driven by reducing cell movement

When starved, Myxococcus xanthus cells assemble themselves into aggregates of about 10(5) cells that grow into complex structures called fruiting bodies, where they later sporulate. Here we present new observations on the velocities of the cells, their orientations, and reversal rates during the early stages of fruiting body formation. Most strikingly, we find that during aggregation, cell velocities slow dramatically and cells orient themselves in parallel inside the aggregates, while later cell orientations are circumferential to the periphery. The slowing of cell velocity, rather than changes in reversal frequency, can account for the accumulation of cells into aggregates. These observations are mimicked by a continuous agent-based computational model that reproduces the early stages of fruiting body formation. We also show, both experimentally and computationally, how changes in reversal frequency controlled by the Frz system mutants affect the shape of these early fruiting bodies.     

Cell density-dependent growth of Myxococcus xanthus on casein.

When Myxococcus xanthus FB was grown on 0.2% casein it exhibited a phenomenon we call cooperative growth. That is, above 104 cells per ml, both strains that were studied exhibited increasing growth rates as a function of increasing cell numbers. Between 104 and 107 cells per ml, the mean doubling times of strains YS and TNS decreased from 15.2 to 8 h and 26 to 8.5 h, respectively. The extracellular proteinase activity of the two strains was equivalent and directly proportional to cell number. Cooperative growth was correlated with increased concentration of hydrolyzed casein in the medium, suggesting cooperative hydrolysis of casein. At low cell densities neither strain was capable of measurable growth on casein in liquid media, and we have calculated that the average concentration of hydrolyzed casein in the medium was indeed too low to support growth. At low cell densities, growth on hydrolyzed casein (Casitone) was normal and independent of cell concentration. Demonstration of cooperative growth at higher cell densities supports the suggestion that the communal behavior of myxobacteria results in more efficient feeding.     

Myxococcus xanthus twin-arginine translocation system is important for growth and development

The twin-arginine translocation (Tat) system serves to export fully folded proteins across the cytoplasmic membrane. In many bacteria, three major components, TatA, TatB and TatC, are the functionally essential constituents of the Tat system. A Myxococcus xanthus tatB–tatC deletion mutant could aggregate and form mounds, but was unable to form fruiting bodies under nutritionally limiting conditions. When tatB–tatC mutant vegetative cells were cultured with 0.5 M glycerol, the cell morphology changed to spore-like spherical cells, but the spores were not resistant to heat and sonication treatments. In contrast to the wild-type strain, the tatB–tatC mutant also showed a decreased cell growth rate and a lower maximum cell concentration. These results suggest possibility that the Tat system may contribute to export of various important proteins for development and growth for M. xanthus.     

Straight-chain fatty acids are dispensable in the myxobacterium Myxococcus xanthus for vegetative growth and fruiting body formation

Inactivation of the MXAN_0853 gene blocked the production in Myxococcus xanthus of straight-chain fatty acids which otherwise represent 30% of total fatty acids. Despite this drastic change in the fatty acid profile, no change in phenotype could be observed, which contrasts with previous interpretations of the role of straight-chain fatty acids in the organism's development.     

Effect of temperature on the growth of Myxococcus xanthus.

The cardinal growth characteristics of Myxococcus xanthus were examined from 14 to 40 degree C, and the examinations indicated that the organism is mesophilic in character. The maximum growth rate (0,3 doublings per h) was between 34 and 36 degree C and the temperature characteristic (micron) is 17,000 cal/mol (71,162 J/mol).     

A RelA-dependent two-tiered regulated proteolysis cascade controls synthesis of a contact-dependent intercellular signal in Myxococcus xanthus

Proteolytic cleavage of precursor proteins to generate intercellular signals is a common mechanism in all cells. In Myxococcus xanthus the contact-dependent intercellular C-signal is a 17 kDa protein (p17) generated by proteolytic cleavage of the 25 kDa csgA protein (p25), and is essential for starvation-induced fruiting body formation. p25 accumulates in the outer membrane and PopC, the protease that cleaves p25, in the cytoplasm of vegetative cells. PopC is secreted in response to starvation, therefore, restricting p25 cleavage to starving cells. We focused on identifying proteins critical for PopC secretion in response to starvation. PopC secretion depends on the (p)ppGpp synthase RelA and the stringent response, and is regulated post-translationally. PopD, which is encoded in an operon with PopC, forms a soluble complex with PopC and inhibits PopC secretion whereas the integral membrane AAA+ protease FtsH(D) is required for PopC secretion. Biochemical and genetic evidence suggest that in response to starvation, RelA is activated and induces the degradation of PopD thereby releasing pre-formed PopC for secretion and that FtsH(D) is important for PopD degradation. Hence, regulated PopC secretion depends on regulated proteolysis. Accordingly, p17 synthesis depends on a proteolytic cascade including FtsH(D) -dependent degradation of PopD and PopC-dependent cleavage of p25.     

ADP-ribosylation by the extracellular fibrils of Myxococcus xanthus

The isolated, extracellular fibrils of the myxobacterium, Myxococcus xanthus , are capable of carrying out ADP-ribosylation. The substrate for the ADP-ribosylation is reactive with monoclonal antibody 2105, which has been shown to be directed specifically against the integral fibril proteins. The extracellular fibrils thus contain both the ADP-ribosyl transferase and the substrate for the ribosylation. This process may play a role in the contact-mediated cell–cell interactions that are an important part of the social behaviour of M. xanthus .     

Morphogenesis in Myxococcus xanthus and Myxococcus virescens (Myxobacterales)

1. Myxococcus xanthus B and M. virescens V2 were compared with a view to establishing the control of their morphogenetic cycles. Both organisms are typical myxococci and on solid media with low concentrations of nutrient they form fruiting bodies, within which vegetative cells convert to myxospores. Ultrathin sections of vegetative M. virescens resembled those of M. xanthus and contained prominent heavily stained bodies, presumed to be polyphosphate granules. Shadowed preparations showed fimbriae associated with M. xanthus but not with M. virescens. 2. M. xanthus B converted to myxospores in liquid medium in response to certain alcohols. M. virescens V2 produced phase-refractile spheres, which were not viable and had an unusual ultrastructure. 3. The distributions of fruiting bodies on solid media containing 0.02% Casitone were recorded for the two species and were compared with a Poisson distribution. Cells responded to differences in cell density in a manner suggestive of a response to a chemotactic attractant. Cells growing vegetatively and also cells forming fruiting bodies produced 3',5'-cyclic adenosine monophosphate (cAMP) as measured by the incorporation of exogeneous [3H] adenosine into cAMP. 4. The significance of these findings for theories of fruiting body formation are discussed.