Energetics of gliding motility in Mycoplasma mobile

Mycoplasma mobile glides on surfaces at up to 7 microm/s by an unknown mechanism. We studied the energetics that power gliding by using a novel, growth medium-free system. We found that cells could glide in defined media if the glass substrate is preconditioned by exposure to horse serum. The active component that potentiates gliding is sensitive to proteinase K treatment. We used the defined medium system to test the effect of various inhibitors, ionophores, and poisons on motility of M. mobile. Valinomycin, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), N,N'-dicyclohexylcarbodiimide, phenamil, amiloride, rifampin, and puromycin had no short-term effects on gliding. We also confirmed that we were able to modulate the membrane potential with valinomycin and FCCP by using a potential-sensitive dye. Shifting the pH likewise had no effect on motility. These results rule out the use of conventional ion motive forces to power gliding. Arsenate had a dramatic inhibitory effect on gliding, and both the speed and the fraction of cells moving tracked ATP levels. Sodium orthovanadate had a slight but significant inhibitory effect on gliding. Taken together, these results suggest that the motor system of M. mobile is likely an ATPase or is directly coupled to an ATPase.     

Prospects for the gliding mechanism of Mycoplasma mobile

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Living microtransporter by uni-directional gliding of Mycoplasma along microtracks

The gliding bacterium Mycoplasma mobile adheres to plastic surfaces and moves around vigorously. However, it has not been possible to control the direction of movements on plain surfaces. Here we report that, on patterned lithographic substrates, M. mobile cells are unable to climb tall walls and move along the bottom edge of the walls. This property to move persistently along walls enabled us to design patterns that control direction of movements, resulting in uni-directional circling or one-way gating between two areas. Furthermore, cells loaded with streptavidin beads following biotinylation of surface proteins moved at normal speeds. These bacteria could be useful as living microtransporters, carrying cargo around within micrometer-scale spaces.     

Transposon mutagenesis identifies genes associated with Mycoplasma pneumoniae gliding motility

The wall-less prokaryote Mycoplasma pneumoniae, a common cause of chronic respiratory tract infections in humans, is considered to be among the smallest and simplest known cells capable of self-replication, yet it has a complex architecture with a novel cytoskeleton and a differentiated terminal organelle that function in adherence, cell division, and gliding motility. Recent findings have begun to elucidate the hierarchy of protein interactions required for terminal organelle assembly, but the engineering of its gliding machinery is largely unknown. In the current study, we assessed gliding in cytadherence mutants lacking terminal organelle proteins B, C, P1, and HMW1. Furthermore, we screened over 3,500 M. pneumoniae transposon mutants individually to identify genes associated with gliding but dispensable for cytadherence. Forty-seven transformants having motility defects were characterized further, with transposon insertions mapping to 32 different open reading frames widely distributed throughout the M. pneumoniae genome; 30 of these were dispensable for cytadherence. We confirmed the clonality of selected transformants by Southern blot hybridization and PCR analysis and characterized satellite growth and gliding by microcinematography. For some mutants, satellite growth was absent or developed more slowly than that of the wild type. Others produced lawn-like growth largely devoid of typical microcolonies, while still others had a dull, asymmetrical leading edge or a filamentous appearance of colony spreading. All mutants exhibited substantially reduced gliding velocities and/or frequencies. These findings significantly expand our understanding of the complexity of M. pneumoniae gliding and the identity of possible elements of the gliding machinery, providing a foundation for a detailed analysis of the engineering and regulation of motility in this unusual prokaryote.     

Using physical models to study the gliding performance of extinct animals

Aerodynamic studies using physical models of fossil organisms can provide quantitative information about how performance of defined activities, such as gliding, depends on specific morphological features. Such analyses allow us to rule out hypotheses about the function of extinct organisms that are not physically plausible and to determine if and how specific morphological features and postures affect performance. The purpose of this article is to provide a practical guide for the design of dynamically scaled physical models to study the gliding of extinct animals using examples from our research on the theropod dinosaur, ?Microraptor gui, which had flight feathers on its hind limbs as well as on its forelimbs. Analysis of the aerodynamics of ?M. gui can shed light on the design of gliders with large surfaces posterior to the center of mass and provide functional information to evolutionary biologists trying to unravel the origins of flight in the dinosaurian ancestors and sister groups to birds. Measurements of lift, drag, side force, and moments in pitch, roll, and yaw on models in a wind tunnel can be used to calculate indices of gliding and parachuting performance, aerodynamic static stability, and control effectiveness in maneuvering. These indices permit the aerodynamic performance of bodies of different shape, size, stiffness, texture, and posture to be compared and thus can provide insights about the design of gliders, both biological and man-made. Our measurements of maximum lift-to-drag ratios of 2.5-3.1 for physical models of ?M. gui suggest that its gliding performance was similar to that of flying squirrels and that the various leg postures that might have been used by ?M. gui make little difference to that aspect of aerodynamic performance. We found that body orientation relative to the movement of air past the animal determines whether it is difficult or easy to maneuver.     

Mycoplasma mobile cells elongated by detergent and their pivoting movements in gliding

Mycoplasma mobile glides on solid surfaces by the repeated binding of leg structures to sialylated oligosaccharide fixed on a solid surface. To obtain information about the propulsion caused by the leg, we made elongated and stiff cells using a detergent. Within 30 min after the cells were treated with 0.1% Tween 60, the cells were elongated from 0.8 μm to 2.2 μm in length while maintaining their gliding activity. Fluorescence and electron microscopy showed that a part of the cytoskeletal structure was elongated, while the localization of proteins involved in the gliding was not modified significantly. The elongated cells glided with repeated pivoting around the cellular position of gliding machinery by 10 degrees of amplitude at a frequency of 2 to 3 times per second, suggesting that the propulsion in a line perpendicular to the cell axis can occur with different timings. The pivoting speed decreased as the cell length increased, probably from the load generated by the friction. The torque required to achieve the actual pivoting increased with the cell length without saturation, reaching 54.7 pN nm at 4.3 μm in cell length.     

Generative models versus underlying symmetries to explain biological pattern

Mathematical models play an increasingly important role in the interpretation of biological experiments. Studies often present a model that generates the observations, connecting hypothesized process to an observed pattern. Such generative models confirm the plausibility of an explanation and make testable hypotheses for further experiments. However, studies rarely consider the broad family of alternative models that match the same observed pattern. The symmetries that define the broad class of matching models are in fact the only aspects of information truly revealed by observed pattern. Commonly observed patterns derive from simple underlying symmetries. This article illustrates the problem by showing the symmetry associated with the observed rate of increase in fitness in a constant environment. That underlying symmetry reveals how each particular generative model defines a single example within the broad class of matching models. Further progress on the relation between pattern and process requires deeper consideration of the underlying symmetries.     

P65 truncation impacts P30 dynamics during Mycoplasma pneumoniae gliding

The cell wall-less prokaryote Mycoplasma pneumoniae is a major cause of community-acquired bronchitis and pneumonia in humans. Colonization is mediated largely by a differentiated terminal organelle, which is also the leading end in gliding motility. Cytadherence-associated proteins P30 and P65 appear to traffic concurrently to the distal end of developing terminal organelles. Here, truncation of P65 due to transposon insertion in the corresponding gene resulted in lower gliding velocity, reduced cytadherence, and decreased steady-state levels of several terminal organelle proteins, including P30. Utilizing fluorescent protein fusions, we followed terminal organelle development over time. New P30 foci appeared at nascent terminal organelles in P65 mutants, as in the wild type. However, with forward cell motility, P30 in the P65 mutants appeared to drag toward the trailing cell pole, where it was released, yielding a fluorescent trail to which truncated P65 colocalized. In contrast, P30 was only rarely observed at the trailing end of gliding wild-type cells. Complementation with the recombinant wild-type P65 allele by transposon delivery restored P65 levels and stabilized P30 localization to the terminal organelle.     

Domain analysis of protein P30 in Mycoplasma pneumoniae cytadherence and gliding motility

The cell wall-less prokaryote Mycoplasma pneumoniae causes bronchitis and atypical pneumonia in humans. Mycoplasma attachment and gliding motility are required for colonization of the respiratory epithelium and are mediated largely by a differentiated terminal organelle. P30 is a membrane protein at the distal end of the terminal organelle and is required for cytadherence and gliding motility, but little is known about the functional role of its specific domains. In the current study, domain deletion and substitution derivatives of P30 were engineered and introduced into a P30 null mutant by transposon delivery to assess their ability to rescue P30 function. Domain deletions involving the extracellular region of P30 severely impacted protein stability and adherence and gliding function, as well as the capacity to stabilize terminal organelle protein P65. Amino acid substitutions in the transmembrane domain revealed specific residues uniquely required for P30 stability and function, perhaps to establish correct topography in the membrane for effective alignment with binding partners. Deletions within the predicted cytoplasmic domain did not affect P30 localization or its capacity to stabilize P65 but markedly impaired gliding motility and cytadherence. The larger of two cytoplasmic domain deletions also appeared to remove the P30 signal peptide processing site, suggesting a larger leader peptide than expected. We propose that the P30 cytoplasmic domain may be required to link P30 to the terminal organelle core, to enable the P30 extracellular domain to achieve a functional conformation, or perhaps both.     

Motivation models fail to explain oviposition behaviour in the diamondback moth

Abstract. It is widely accepted that previous experience and internal physiological factors (such as egg‐load) affect host‐plant discrimination during oviposition by phytophagous insects. However, there is some debate as to how these factors interact in a mechanistic sense to control acceptance. The role of learning and host deprivation in host acceptance by adult diamondback moths (Plutella xylostella L.) was investigated. In the first experiment, we tested whether experience of a lower ranked host cabbage increased acceptance of a higher ranked host as predicted by a hierarchy threshold model. Moths trained on cabbage were over three times more likely to accept cabbage during testing than untrained moths. There was no effect of cabbage training on acceptance of cress, indicating that the effect of training was species‐specific. In a second experiment, designed to test the prediction of motivation models that insects become less discriminating when deprived of oviposition opportunities, depriving females of host plants for 2 nights significantly increased female egg‐load (×2.3). Host deprivation did not decrease discrimination between the preferred host cress and cabbage. Cabbage and cress plants were equally likely to have been accepted by nondeprived moths after 1 night of exposure, yet moths deprived of hosts for 2 nights strongly preferred cress when tested during the first 20 min of the scotophase. During this 20‐min period, previous host deprivation increased acceptance of host plants generally but did not decrease discrimination between hosts. These data contradict the expectation that there is an inverse relationship between host species discrimination and the failure of an insect to find hosts as found in existing oviposition acceptance models. As an alternative, the Incremental Acceptance Model of host acceptance behaviour is presented, in which responsiveness to a host is a function of the recent encounter rate with host‐specific stimuli, and the oviposition reflex is regulated by nonspecific cues such as egg‐load.     

The use of structural dynamic models to explain successes and failures of biomanipulation

A development of a structural dynamic model, i.e. a model with current change of the most important parameters according to a goal function, is presented with the aim to explain the structural changes observed in lakes, when the nutrient concentration is increased or decreased. This type of models may be important in lake management as it may be possible qualitatively to predict the success or failure of biomanipulation. The answer to the crucial question: àt which phosphorus level will the success of biomanipulation be most probable?' will probably require the development of model which takes into account site specific processes and properties, i.e., a more complicated model. As goal function is proposed the thermodynamic function, exergy, which is defined as the work content of the system (model) compared with the system at thermodynamic equilibrium. It is shown that the structural dynamic modelling approach has been able to explain the shift from large to small zooplankton species at a certain level of phosphorus concentration, accompanied by a shifts from a dominance of zooplankton, and predatory fish to a system dominated by planktivorous fish and phytoplankton. The shift in zooplankton species cannot be explained by application of catastrophe theoretical models, which have been used to explain the hysteresis reaction. The results show that the shift should be expected at approximately 0.12 mg P l-1 and that a typical hysteresis reaction occurs at this concentration in accordance with the expectations. These results are consistent with many observations but should be interpreted with great caution, as the model is simple and general and don't account for a number of processes which may influence the results significantly in specific lake studies. The structural dynamic approach has previously been used in ten case studies with good agreement with the observations, but more case studies are needed before a general recommendation of the use of this type of models can be given. The results from this study point toward to apply this type of models for lake management where biomanipulation is involved, although it should be recommended to improve the presented general model with introduction of site specific properties for a considered lake study.     

Integration of molecular and physiological models to explain time of anthesis in wheat

Background and Aims

A model to predict anthesis time of a wheat plant from environmental and genetic information requires integration of current concepts in physiological and molecular biology. This paper describes the structure of an integrated model and quantifies its response mechanisms.

Methods

Literature was reviewed to formulate the components of the model. Detailed re-analysis of physiological observations are utilized from a previous publication by the second two authors. In this approach measurements of leaf number and leaf and primordia appearance of near isogenic lines of spring and winter wheat grown for different durations in different temperature and photoperiod conditions are used to quantify mechanisms and parameters to predict time of anthesis.

Key Results

The model predicts the time of anthesis from the length of sequential phases: 1, embryo development; 2, dormant; 3, imbibed/emerging; 4, vegetative; 5, early reproductive; 6, pseudo-stem extension; and 7, ear development. Phase 4 ends with vernalization saturation (VS), Phase 5 with terminal spikelet (TS) and Phase 6 with flag leaf ligule appearance (FL). The durations of Phases 4 and 5 are linked to the expression of Vrn genes and are calculated in relation to change in Haun stage (HS) to account for the effects of temperature per se. Vrn1 must be expressed to sufficient levels for VS to occur. Vrn1 expression occurs at a base rate of 0·08/HS in winter ‘Batten’ and 0·17/HS in spring ‘Batten’ during Phases 1, 3 and 4. Low temperatures promote expression of Vrn1 and accelerate progress toward VS. Our hypothesis is that a repressor, Vrn4, must first be downregulated for this to occur. Rates of Vrn4 downregulation and Vrn1 upregulation have the same exponential response to temperature, but Vrn4 is quickly upregulated again at high temperatures, meaning short exposure to low temperature has no impact on the time of VS. VS occurs when Vrn1 reaches a relative expression of 0·76 and Vrn3 expression begins. However, Vrn2 represses Vrn3 expression so Vrn1 must be further upregulated to repress Vrn2 and enable Vrn3 expression. As a result, the target for Vrn1 to trigger VS was 0·76 in 8-h photoperiods (Pp) and increased at 0·026/HS under 16-h Pp as levels of Vrn2 increased. This provides a mechanism to model short-day vernalization. Vrn3 is expressed in Phase 5 (following VS), and apparent rates of Vrn3 expression increased from 0·15/HS at 8-h Pp to 0·33/HS at 16-h Pp. The final number of leaves is calculated as a function of the HS at which TS occurred (TS^HS): 2·86 + 1·1 × TS^HS. The duration of Phase 6 is then dependent on the number of leaves left to emerge and how quickly they emerge.

Conclusions

The analysis integrates molecular biology and crop physiology concepts into a model framework that links different developmental genes to quantitative predictions of wheat anthesis time in different field situations.     

The influence of various substances on the gliding motility of Mycoplasma mobile 163K

Non-toxic concentrations of various substances were tested for their influence on the gliding motility of Mycoplasma mobile 163K. A significant inhibitory effect on motility was observed with agents acting on nucleic acid synthesis (mitomycin), protein synthesis (puromycin, chloramphenicol), energy metabolism (p-chloromercuribenzoate, iodoacetate) and with compounds reacting with the cytoplasmic membrane or contractile elements (albumin, cholesterol, EDTA, 2-propanol, procain, CaCl2, MgCl2, colchicin and KI). The surface-active compounds Triton X-100, Tego and SDS increased the gliding velocity significantly in some concentrations and incubation periods. The results suggest that the motility of M. mobile depends on a functional cytoplasmic membrane and that cytoskeletal elements are involved in the gliding mechanism.     

Processing is required for a fully functional protein P30 in Mycoplasma pneumoniae gliding and cytadherence

The cell wall-less prokaryote Mycoplasma pneumoniae causes bronchitis and atypical pneumonia in humans. Mycoplasma attachment to the host respiratory epithelium is required for colonization and mediated largely by a differentiated terminal organelle. P30 is an integral membrane protein located at the distal end of the terminal organelle. The P30 null mutant II-3 is unable to attach to host cells and nonmotile and has a branched cellular morphology compared to the wild type, indicating an important role for P30 in M. pneumoniae biology. P30 is predicted to have an N-terminal signal sequence, but the presence of such a motif has not been confirmed experimentally. In the current study we analyzed P30 derivatives having epitope tags engineered at various locations to demonstrate that posttranslational processing occurred in P30. Several potential cleavage sites predicted in silico were examined, and a processing-defective mutant was created to explore P30 maturation further. Our results suggested that signal peptide cleavage occurs between residues 52 and 53 to yield mature P30. The processing-defective mutant exhibited reduced gliding velocity and cytadherence, indicating that processing is required for fully functional maturation of P30. We speculate that P30 processing may trigger a conformational change in the extracellular domain or expose a binding site on the cytoplasmic domain to allow interaction with a binding partner as a part of functional maturation.     

Involvement of P1 adhesin in gliding motility of Mycoplasma pneumoniae as revealed by the inhibitory effects of antibody under optimized gliding conditions

To examine the participation of P1 adhesin in gliding of Mycoplasma pneumoniae, we examined the effects of an anti-P1 monoclonal antibody on individual gliding mycoplasmas. The antibody reduced the gliding speed and removed the gliding cells from the glass over time in a concentration-dependent manner but had only a slight effect on nongliding cells, suggesting that the conformational changes of P1 adhesin and its displacement are involved in the gliding mechanism.     

Computational models explain the oligosaccharide specificity of cyanovirin-N

The prokaryotic lectin cyanovirin-N (CV-N) is a potent inhibitor of HIV envelope-mediated cell entry, and thus is a leading candidate among a new class of potential anti-HIV microbicides. The activity of CV-N is a result of interactions with the D1 arm of high-mannose oligosaccharides on the viral glycoprotein gp120. Here, we present computationally refined models of CV-N recognition of the di- and trisaccharides that represent the terminal three sugars of the D1 arm by each CV-N binding site. These models complement existing structural data, both from NMR spectroscopy and X-ray crystallography. When used with a molecular dynamics/continuum electrostatic (MD/PBSA) approach to compute binding free energies, these models explain the relative affinity of each site for the two saccharides. This work presents the first validation of the application of continuum electrostatic models to carbohydrate-protein association. Taken as a whole, the results both provide models of CV-N sugar recognition and demonstrate the utility of these computational methods for the study of carbohydrate-binding proteins.     

Simple MaxEnt models explain food web degree distributions

Degree distributions are widely used to characterize networks, including food webs, and play a vital role in models of food web structure. To date, there have been no mechanistic or statistical explanations for the form of food web degree distributions. Here, I introduce models for food web degree distributions based on the principle of maximum entropy (MaxEnt) and show that the distributions of the number of consumers and resources in 23 (45%) and 35 (69%) of 51 food webs are not significantly different at a 95% confidence level from the MaxEnt distribution. These findings offer a new null model for the most probable degree distributions in food webs and other networks. They suggest that there is relatively little pressure favoring generalist or specialist consumption strategies but that biological drivers or methodological bias may force the consumer distribution away from the MaxEnt form.     

Movement on the cell surface of the gliding bacterium,Mycoplasma mobile,is limited to its head-like structure

Mycoplasma mobile cells glide on solid surfaces such as glass with a fast and continuous motion in the direction of the membrane protrusion (head-like structure) at one cell pole. To examine its cell-surface movement, a latex bead was attached to a cell and behavior in gliding was monitored. The bead was carried without movement relative to the cell body, suggesting that the cell does not roll around the cell axis and the surface movement is limited to a small area. A small percentage of cells showed an elongated head-like structure in an old batch culture. The head-like structure moved forward, sometimes leaving the cell body in one position, resulting in a stretching of this head-like structure. These results indicate that the head-like structure drags the cell body, leading us to conclude that the force for gliding is generated at the head-like structure.