RESUMO
When cells of the social amoeba Dictyostelium discoideum are starved of nutrients they start to synthesize and secrete the chemical messenger and chemoattractant cyclic adenosine monophosphate (cAMP). This signal is relayed by other cells, resulting in the establishment of periodic waves. The cells aggregate through chemotaxis toward the center of these waves. We investigated the chemotactic response of individual cells to repeated exposure to waves of cAMP generated by a microfluidic device. For fast-moving waves (short period), the chemotactic ability of the cells was found to increase upon exposure to more waves, suggesting the development of a memory over several cycles. This effect was not significant for slow-moving waves (large period). We show that the experimental results are consistent with a local excitation global inhibition-based model, extended by including a component that rises and decays slowly and that is activated by the temporal gradient of cAMP concentration. The observed enhancement in chemotaxis is relevant to populations in the wild: once sustained, periodic waves of the chemoattractant are established, it is beneficial to cells to improve their chemotactic ability in order to reach the aggregation center sooner.
Assuntos
Quimiotaxia , AMP Cíclico , Dictyostelium , Modelos Biológicos , Quimiotaxia/fisiologia , Dictyostelium/fisiologia , AMP Cíclico/metabolismo , Fatores Quimiotáticos/farmacologia , Fatores Quimiotáticos/metabolismoRESUMO
Chemotaxis, the guided motion of cells by chemical gradients, plays a crucial role in many biological processes. In the social amoeba Dictyostelium discoideum, chemotaxis is critical for the formation of cell aggregates during starvation. The cells in these aggregates generate a pulse of the chemoattractant, cyclic adenosine 3',5'-monophosphate (cAMP), every 6 min to 10 min, resulting in surrounding cells moving toward the aggregate. In addition to periodic pulses of cAMP, the cells also secrete phosphodiesterase (PDE), which degrades cAMP and prevents the accumulation of the chemoattractant. Here we show that small aggregates of Dictyostelium can disperse, with cells moving away from instead of toward the aggregate. This surprising behavior often exhibited oscillatory cycles of motion toward and away from the aggregate. Furthermore, the onset of outward cell motion was associated with a doubling of the cAMP signaling period. Computational modeling suggests that this dispersal arises from a competition between secreted cAMP and PDE, creating a cAMP gradient that is directed away from the aggregate, resulting in outward cell motion. The model was able to predict the effect of PDE inhibition as well as global addition of exogenous PDE, and these predictions were subsequently verified in experiments. These results suggest that localized degradation of a chemoattractant is a mechanism for morphogenesis.
Assuntos
Movimento Celular , Fatores Quimiotáticos/metabolismo , Dictyostelium/citologia , Agregação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Simulação por Computador , AMP Cíclico/metabolismo , Dictyostelium/efeitos dos fármacos , Fluorescência , Inibidores de Fosfodiesterase/farmacologia , Diester Fosfórico Hidrolases/metabolismo , Transdução de SinaisRESUMO
Signaling networks are at the heart of almost all biological processes. Most of these networks contain large number of components, and often either the connections between these components are not known or the rate equations that govern the dynamics of soluble signaling components are not quantified. This uncertainty in network topology and parameters can make it challenging to formulate detailed mathematical models. Boolean networks, in which all components are either on or off, have emerged as viable alternatives to detailed mathematical models that contain rate constants and other parameters. Therefore, open-source platforms of Boolean models for community use are desirable. Here, we present Boolink, a freely available graphical user interface that allows users to easily construct and analyze existing Boolean networks. Boolink can be applied to any Boolean network. We demonstrate its application using a previously published network for abscisic acid (ABA)-driven stomatal closure in Arabidopsis spp. (Arabidopsis thaliana). We also show how Boolink can be used to generate testable predictions by extending the network to include CO2 regulation of stomatal movements. Predictions of the model were experimentally tested, and the model was iteratively modified based on experiments showing that ABA effectively closes Arabidopsis stomata at near-zero CO2 concentrations (1.5-ppm CO2). Thus, Boolink enables public generation and the use of existing Boolean models, including the prior developed ABA signaling model with added CO2 signaling components.
Assuntos
Algoritmos , Fenômenos Bioquímicos , Dióxido de Carbono/metabolismo , Estômatos de Plantas/fisiologia , Transdução de Sinais/fisiologia , Interpretação Estatística de Dados , Modelos TeóricosRESUMO
Bacterial chemotaxis is a biased movement of bacteria toward the beneficial chemical gradient or away from a toxic chemical gradient. This movement is achieved by sensing a chemical gradient by chemoreceptors. In most of the chemotaxis studies, Escherichia coli has been used as a model organism. E. coli have about 4-6 flagella on their surfaces, and the motility is achieved by rotating the flagella. Each flagellum has reversible flagellar motors at its base, which rotate the flagella in counterclockwise and clockwise directions to achieve "run" and "tumble." The chemotaxis of bacteria is regulated by a network of interacting proteins. The sensory signal is processed and transmitted to the flagellar motor by cytoplasmic proteins. Bacterial chemotaxis plays an important role in many biological processes such as biofilm formation, quorum sensing, bacterial pathogenesis, and host infection. Bacterial chemotaxis can be applied for bioremediation, horizontal gene transfer, drug delivery, or maybe some other industry in near future. This review contains an overview of bacterial chemotaxis, recent findings of the physiological importance of bacterial chemotaxis in other biological processes, and the application of bacterial chemotaxis.
Assuntos
Bactérias/metabolismo , Fenômenos Fisiológicos Bacterianos , Células Quimiorreceptoras/metabolismo , Quimiotaxia , Flagelos/fisiologia , Bactérias/genética , Proteínas de Bactérias/genética , Proteínas de MembranaRESUMO
Adhesive cell-substrate interactions are crucial for cell motility and are responsible for the necessary traction that propels cells. These interactions can also change the shape of the cell, analogous to liquid droplet wetting on adhesive substrates. To address how these shape changes affect cell migration and cell speed we model motility using deformable, 2D cross-sections of cells in which adhesion and frictional forces between cell and substrate can be varied separately. Our simulations show that increasing the adhesion results in increased spreading of cells and larger cell speeds. We propose an analytical model which shows that the cell speed is inversely proportional to an effective height of the cell and that increasing this height results in increased internal shear stress. The numerical and analytical results are confirmed in experiments on motile eukaryotic cells.
Assuntos
Adesão Celular , Movimento Celular , Molhabilidade , Dictyostelium/citologia , Modelos BiológicosRESUMO
Negative chemotaxis refers to the motion of microorganisms away from regions with high concentrations of chemorepellents. In this study, we set controlled gradients of NiCl2, a chemorepellent, in microchannels to quantify the motion of Escherichia coli over a broad range of concentrations. The experimental technique measured the motion of the bacteria in space and time and further related the motion to the local concentration profile of the repellent. Results show that the swimming speed of bacteria increases with an increasing concentration of repellent, which in turn enhances the drift velocity. The contribution of the increased swimming speed to the total drift velocity was in the range of 20 to 40%, with the remaining contribution coming from the modulation of the tumble frequency. A simple model that incorporates receptor dynamics, including adaptation, intracellular signaling, and swimming speed variation, was able to qualitatively capture the observed trend in drift velocity.
Assuntos
Quimiotaxia , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Locomoção , Níquel/metabolismo , Modelos BiológicosRESUMO
It is well known that Escherichia coli achieves chemotaxis by modulating the bias of the flagellar motor. Recent experiments have shown that the bacteria vary their swimming speeds as well in presence of attractants. However, this increase in the swimming speed in response to the attractants has not been correlated with the increase in the flagellar motor speed. Using flickering dark-field microscopy, we measure the head-rotation speed of a large population of cells to correlate it with the flagellar motor speed. Experiments performed with wild-type and trg-deletion mutant strains suggest that the cells are capable of modulating the flagellar motor speed via mere sensing of a ligand. The motor speed can be further correlated with the swimming speed of the cells and was found to be linear. These results suggest the existence of a hitherto unknown intra-cellular pathway that modulates the flagellar motor speed in response to sensing of chemicals, thereby making chemotaxis more efficient than previously known.
Assuntos
Quimiotaxia/fisiologia , Proteínas de Escherichia coli/genética , Escherichia coli/fisiologia , Flagelos/fisiologia , Proteínas de Membrana/genética , Rotação , Deleção de Sequência/genéticaRESUMO
It is well known that Escherichia coli executes chemotactic motion in response to chemical cues by modulating the flagellar motor bias alone. However, previous studies have reported the possibility of variation in run speed in the presence of attractants although it is unclear whether bacteria can deliberately modulate their swimming speeds in response to environmental cues or if the motor speeds are hardwired. By studying the detailed motion of cells in a uniform concentration of glucose and its non-metabolizable analogue, we show that changing concentrations may be accompanied by variation in the swimming speed. For a fixed run duration, cells exposed to the attractants achieved a higher peak-swimming speed after a tumble compared with that in plain motility buffer. Our experiments using the mutant strain lacking the Trg sensor show no change in swimming speed with varying concentrations of the non-metabolizable analogue, suggesting that sensing may play a role in the observed variation of swimming speed.
Assuntos
Quimiotaxia/efeitos dos fármacos , Desoxiglucose/farmacologia , Escherichia coli K12/efeitos dos fármacos , Escherichia coli K12/fisiologia , Glucose/farmacologia , Fatores Quimiotáticos/farmacologia , TempoRESUMO
Cells of the social amoeba Dictyostelium discoideum migrate to a source of periodic traveling waves of chemoattractant as part of a self-organized aggregation process. An important part of this process is cellular memory, which enables cells to respond to the front of the wave and ignore the downward gradient in the back of the wave. During this aggregation, the background concentration of the chemoattractant gradually rises. In our microfluidic experiments, we exogenously applied periodic waves of chemoattractant with various background levels. We find that increasing background does not make detection of the wave more difficult, as would be naively expected. Instead, we see that the chemotactic efficiency significantly increases for intermediate values of the background concentration but decreases to almost zero for large values in a switch-like manner. These results are consistent with a computational model that contains a bistable memory module, along with a nonadaptive component. Within this model, an intermediate background level helps preserve directed migration by keeping the memory activated, but when the background level is higher, the directional stimulus from the wave is no longer sufficient to activate the bistable memory, suppressing directed migration. These results suggest that raising levels of chemoattractant background may facilitate the self-organized aggregation in Dictyostelium colonies.
Assuntos
Fatores Quimiotáticos/farmacologia , Quimiotaxia/efeitos dos fármacos , AMP Cíclico/metabolismo , Dictyostelium/citologia , Dictyostelium/efeitos dos fármacos , Dictyostelium/metabolismo , Relação Dose-Resposta a Droga , Modelos BiológicosRESUMO
Cell-substrate adhesion of the social amoeba Dictyostelium discoideum, a model organism often used for the study of chemotaxis, is non-specific and does not involve focal adhesion complexes. Therefore, micropatterned substrates where adherent Dictyostelium cells are constrained to designated microscopic regions are difficult to make. Here we present a micropatterning technique for Dictyostelium cells that relies on coating the substrate with an â¼1µm thick layer of polyethylene glycol (PEG) gel. We show that, when plated on a substrate with narrow parallel stripes of PEG-gel and glass, Dictyostelium cells nearly exclusive adhere to and migrate along the glass stripes, thus providing a model system to study one-dimensional migration of amoeboid cells. Surprisingly, we find substantial differences in the adhesion to PEG-gel and glass stripes between vegetative and developed cells and between two different axenic laboratory strains of Dictyostelium, AX2 and AX4. Even more surprisingly, we find that the distribution of Dictyostelium cells between PEG-gel and glass stripes is significantly affected by the expression of several fluorescent protein markers of the cytoskeleton. We carry out atomic force microscopy based single cell force spectroscopy measurements that confirm that the force of adhesion to PEG-gel substrate can be significantly different between vegetative and developed cells, AX2 and AX4 cells, and cells with and without fluorescent markers. Thus, the choice of parental background, the degree of development, and the expression of fluorescent protein markers can all have a profound effect on cell-substrate adhesion and should be considered when comparing migration of cells and when designing micropatterned substrates.
Assuntos
Movimento Celular , Dictyostelium/citologia , Corantes Fluorescentes/metabolismo , Microtecnologia/métodos , Polietilenoglicóis/farmacologia , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Dictyostelium/efeitos dos fármacos , Géis/farmacologia , Análise EspectralRESUMO
Studies on chemotaxis of Escherichia coli have shown that modulation of tumble frequency causes a net drift up the gradient of attractants. Recently, it has been demonstrated that the bacteria is also capable of varying its runs speed in uniform concentration of attractant. In this study, we investigate the role of swimming speed on the chemotactic migration of bacteria. To this end, cells are exposed to gradients of a non-metabolizable analogue of glucose which are sensed via the Trg sensor. When exposed to a gradient, the cells modulate their tumble duration, which is accompanied with variation in swimming speed leading to drift velocities that are much higher than those achieved through the modulation of the tumble duration alone. We use an existing intra-cellular model developed for the Tar receptor and incorporate the variation of the swimming speed along with modulation of tumble frequency to predict drift velocities close to the measured values. The main implication of our study is that E. coli not only modulates the tumble frequency, but may also vary the swimming speed to affect chemotaxis and thereby efficiently sample its nutritionally rich environment.
RESUMO
Osmotic Shock is known to negatively affect growth rate along with an extended lag phase. The reduction in growth rate can be characterized as burden due to the osmotic stress. Studies have shown that production of unnecessary protein also burdens cellular growth. This has been demonstrated by growing Escherichia coli on glycerol in the presence of Isopropyl-ß-D-1-thiogalactopyranoside (IPTG) to induce ß-galactosidase synthesis which does not offer any benefit towards growth. The trade off between osmotic stress and burden on growth due to unnecessary gene expression has not been enumerated. The influence of osmotic stress on ß-galactosidase synthesis and activity is not clearly understood. Here, we study the effect of salt concentration on ß-galactosidase activity and burden on growth due to unnecessary gene expression in E.coli. We characterize the burden on growth in presence of varying concentrations of salt in the presence of IPTG using three strains, namely wild type, ∆lacI and ∆lacIlacZ mutant strains. We demonstrate that the salt concentrations, sensitively inhibits enzyme synthesis thereby influencing the burden on growth. In a wild type strain, addition of lactose into the medium demonstrated growth benefit at low salt concentration but not at higher concentrations. The extent of burden due to osmotic shock was higher in a lactose M9 medium than in a glycerol M9 medium. A linear relationship was observed between enzyme activity and burden on growth in various media types studied.