Live Cell Analysis and Mathematical Modeling Identify Determinants of Attenuation of Dengue Virus 2’-O-Methylation Mutant

Dengue virus (DENV) is the most common mosquito-transmitted virus infecting ~390 million people worldwide. In spite of this high medical relevance, neither a vaccine nor antiviral therapy is currently available. DENV elicits a strong interferon (IFN) response in infected cells, but at the same time actively counteracts IFN production and signaling. Although the kinetics of activation of this innate antiviral defense and the timing of viral counteraction critically determine the magnitude of infection and thus disease, quantitative and kinetic analyses are lacking and it remains poorly understood how DENV spreads in IFN-competent cell systems. To dissect the dynamics of replication versus antiviral defense at the single cell level, we generated a fully viable reporter DENV and host cells with authentic reporters for IFN-stimulated antiviral genes. We find that IFN controls DENV infection in a kinetically determined manner that at the single cell level is highly heterogeneous and stochastic. Even at high-dose, IFN does not fully protect all cells in the culture and, therefore, viral spread occurs even in the face of antiviral protection of naïve cells by IFN. By contrast, a vaccine candidate DENV mutant, which lacks 2’-O-methylation of viral RNA is profoundly attenuated in IFN-competent cells. Through mathematical modeling of time-resolved data and validation experiments we show that the primary determinant for attenuation is the accelerated kinetics of IFN production. This rapid induction triggered by mutant DENV precedes establishment of IFN-resistance in infected cells, thus causing a massive reduction of virus production rate. In contrast, accelerated protection of naïve cells by paracrine IFN action has negligible impact. In conclusion, these results show that attenuation of the 2’-O-methylation DENV mutant is primarily determined by kinetics of autocrine IFN action on infected cells.     

In situ Localization and Isolation of Actin Filaments from Pollen Tubes of Amaryllis vittata Ait

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Mathematical Modeling of Ischemia–Reperfusion Injury and Postconditioning Therapy

Reperfusion (restoration of blood flow) after a period of ischemia (interruption of blood flow) can paradoxically place tissues at risk of further injury: so-called ischemia–reperfusion injury or IR injury. Recent studies have shown that postconditioning (intermittent periods of further ischemia applied during reperfusion) can reduce IR injury. We develop a mathematical model to describe the reperfusion and postconditioning process following an ischemic insult, treating the blood vessel as a two-dimensional channel, lined with a monolayer of endothelial cells that interact (respiration and mechanotransduction) with the blood flow. We investigate how postconditioning affects the total cell density within the endothelial layer, by varying the frequency of the pulsatile flow and the oxygen concentration at the inflow boundary. We find that, in the scenarios we consider, the pulsatile flow should be of high frequency to minimize cellular damage, while oxygen concentration at the inflow boundary should be held constant, or subject to only low-frequency variations, to maximize cell proliferation.     

Impact of Imitation Processes on the Effectiveness of?Ring Vaccination

Ring vaccination can be a highly effective control strategy for an emerging disease or in the final phase of disease eradication, as witnessed in the eradication of smallpox. However, the impact of behavioural dynamics on the effectiveness of ring vaccination has not been explored in mathematical models. Here, we analyze a series of stochastic models of voluntary ring vaccination. Contacts of an index case base vaccinating decisions on their own individual payoffs to vaccinate or not vaccinate, and they can also imitate the behaviour of other contacts of the index case. We find that including imitation changes the probability of containment through ring vaccination considerably. Imitation can cause a strong majority of contacts to choose vaccination in some cases, or to choose non-vaccination in other cases-even when the equivalent solution under perfectly rational (non-imitative) behaviour yields mixed choices. Moreover, imitation processes can result in very different outcomes in different stochastic realizations sampled from the same parameter distributions, by magnifying moderate tendencies toward one behaviour or the other: in some realizations, imitation causes a strong majority of contacts not to vaccinate, while in others, imitation promotes vaccination and reduces the number of secondary infections. Hence, the effectiveness of ring vaccination can depend significantly and unpredictably on imitation processes. Therefore, our results suggest that risk communication efforts should be initiated early in an outbreak when ring vaccination is to be applied, especially among subpopulations that are heavily influenced by peer opinions.     

Models of Electrical Activity: Calibration and Prediction Testing on?the?Same Cell

Mathematical models are increasingly important in biology, and testability is becoming a critical issue. One limitation is that one model simulation tests a parameter set representing one instance of the biological counterpart, whereas biological systems are heterogeneous in their properties and behavior, and a model often is fitted to represent an ideal average. This is also true for models of a cell’s electrical activity; even within a narrowly defined population there can be considerable variation in electrophysiological phenotype. Here, we describe a computational experimental approach for parameterizing a model of the electrical activity of a cell in real time. We combine the inexpensive parallel computational power of a programmable graphics processing unit with the flexibility of the dynamic clamp method. The approach involves 1), recording a cell’s electrical activity, 2), parameterizing a model to the recording, 3), generating predictions, and 4), testing the predictions on the same cell used for the calibration. We demonstrate the experimental feasibility of our approach using a cell line (GH4C1). These cells are electrically active, and they display tonic spiking or bursting. We use our approach to predict parameter changes that can convert one pattern to the other.     

Modeling T Cell Proliferation and Death in Vitro Based on Labeling Data: Generalizations of the Smith–Martin Cell Cycle Model

The fluorescent dye carboxyfluorescein diacetate succinimidyl ester (CFSE) classifies proliferating cell populations into groups according to the number of divisions each cell has undergone (i.e., its division class). The pulse labeling of cells with radioactive thymidine provides a means to determine the distribution of times of entry into the first cell division. We derive in analytic form the number of cells in each division class as a function of time based on the distribution of times to the first division. Choosing the distribution of time to the first division to fit thymidine labeling data for T cells stimulated in vitro under different concentrations of IL-2, we fit CFSE data to determine the dependence of T cell kinetic parameters on the concentration of IL-2. As the concentration of IL-2 increases, the average cell cycle time is shortened, the death rate of cells is decreased, and a higher fraction of cells is recruited into division. We also find that if the average cell cycle time increases with division class then the qualify of our fit to the data improves.     

Comparative Study on Interaction of DNaseⅠ with Pollen Actin and Rabbit Muscle Actin

Actin purified from maize pollen grains like actin from other sources could considerably inhibit the activity of DNase Ⅰ . A linear relationship existed between inhibition and the concentration of actin. However, DNase Ⅰ was less inhibited by pollen actin than by rabbit muscle actin under the same conditions. The values of Kapp of inhibition were 1.25 μg/mL for pollen actin and 0.75 μg/mL for rabbit muscle actin. DNase Ⅰdepolymerized both pollen and rabbit muscle actin filaments. But the rate of depolymerization of pollen F-actin was higher than that of rabbit muscle F-actin under the same conditions.     

Impact of the ‘tubulin economy’ on the formation and function of the microtubule cytoskeleton

The microtubule cytoskeleton is assembled from a finite pool of α,β-tubulin, the size of which is controlled by an autoregulation mechanism. Cells also tightly regulate the architecture and dynamic behavior of microtubule arrays. Here, we discuss progress in our understanding of how tubulin autoregulation is achieved and highlight work showing that tubulin, in its unassembled state, is relevant for regulating the formation and organization of microtubules. Emerging evidence suggests that tubulin regulates microtubule-associated proteins and kinesin motors that are critical for microtubule nucleation, dynamics, and function. These relationships create feedback loops that connect the tubulin assembly cycle to the organization and dynamics of microtubule networks. We term this concept the ‘tubulin economy’, which emphasizes the idea that tubulin is a resource that can be deployed for the immediate purpose of creating polymers, or alternatively as a signaling molecule that has more far-reaching consequences for the organization of microtubule arrays.     

A Mathematical Model on Water Redistribution Mechanism of?the?Seismonastic Movement of Mimosa Pudica

A theoretical model based on the water redistribution mechanism is proposed to predict the volumetric strain of motor cells in Mimosa pudica during the seismonastic movement. The model describes the water and ion movements following the opening of ion channels triggered by stimulation. The cellular strain is related to the angular velocity of the plant movement, and both their predictions are in good agreement with experimental data, thus validating the water redistribution mechanism. The results reveal that an increase in ion diffusivity across the cell membrane of <15-fold is sufficient to produce the observed seismonastic movement.     

Actin and Microtubules in Cell Motility: Which One is in Control?

The cytoskeleton is composed of three distinct elements: actin microfilaments, microtubules and intermediate filaments. The actin cytoskeleton is thought to provide protrusive and contractile forces, and microtubules to form a polarized network allowing organelle and protein movement throughout the cell. Intermediate filaments are generally considered the most rigid component, responsible for the maintenance of the overall cell shape. Cytoskeletal elements must be coordinately regulated for the cell to fulfill complex cellular functions, as diverse as cell migration, cell adhesion and cell division. Coordination between cytoskeletal elements is achieved by signaling pathways, involving common regulators such as the Rho guanosine-5'-triphosphatases (GTPases). Furthermore, evidence is now accumulating that cytoskeletal elements participate in regulating each other. As a consequence, although their functions seem well defined, they are in fact overlapping, with actin playing a role in membrane trafficking and microtubules being involved in the control of protrusive and contractile forces. This cytoskeletal crosstalk is both direct and mediated by signaling molecules. Cell motility is a well-studied example where the interplay between actin and microtubules appears bidirectional. This leads us to wonder which, if any, cytoskeletal element leads the way.     

Impact of gut-associated bifidobacteria and their phages on health: two sides of the same coin?

Applied Microbiology and Biotechnology - Bifidobacteria are among the first microbial colonisers of the human infant gut post-partum. Their early appearance and dominance in the human infant gut...     

Modeling of the effects of IL-17 and TNF-α on endothelial cells and thrombus growth

Rheumatoid and psoriatic arthritis are chronic inflammatory diseases, with massive increase of cardiovascular events (CVE), and contribution of the cytokines TNF-α and IL-17. Chronic inflammation inside the joint membrane or synovium results from the activation of fibroblasts/synoviocytes, and leads to the release of cytokines from monocytes (Tumor Necrosis Factor or TNF) and from T lymphocytes (Interleukin-17 or IL-17). At the systemic level, the very same cytokines affect endothelial cells and vessel wall. We have previously shown [1], [2] that IL-17 and TNF-α, specifically when combined, increase procoagulation, decrease anticoagulation and increase platelet aggregation, leading to thrombosis. These results are the basis for the models of interactions between IL-17 and TNF, and genes expressed by activated endothelial cells. This work is devoted to mathematical modeling and numerical simulations of blood coagulation and clot growth under the influence of IL-17 and TNF-α. We show that they can provoke thrombosis, leading to the complete or partial occlusion of blood vessels. The regimes of blood coagulation and conditions of occlusion are investigated in numerical simulations and in approximate analytical models. The results of mathematical modeling allow us to predict thrombosis development for an individual patient.     

Mathematical Modeling of the Dynamics of Plant Mineral Nutrition in the Fertilizer–Soil–Plant System

A numerical simulation of the spatial–temporal dynamics of a multi-parameter system has been developed. The components of this system are plant biomass, the mobile and stationary forms of mineral nutrition elements, rhizosphere microorganisms, and environmental parameters (temperature, humidity, and acidity). Parametric identification and verification of the adequacy of the model were carried out based on the experimental data on the growth of Krasnoufimskaya-100 spring wheat on peat lowland soil. The results are represented by temporal distributions of biomass from agricultural crops and the findings on the contents of the main nutrition elements within the plant (nitrogen, phosphorus, and potassium). An agronomic assessment and interpretation of the results are given.

     

Light-Dependent Chloroplast Reorientation in Mougeotia and Mesotaenium: Riased by Pigment-Regulated Plasmalemma Anchorage Sites to Actin Filaments?

Conjugatophycean green algae, such as Mougeotia and Mesotaenium, are presumably the most ancient organisms to show phytochrome-mediated photomodulatory processes, i.e. chloroplast reorientational movements. Experiments have provided striking evidence for a dichroic mode of light absorption by the phytochrome molecules located at the periphery of the cylindrical cell; in addition, the transition moment of the chromophoric group of phytochrome has been shown to change by a fixed angle upon conversion of P_r to P_fr and vice versa. Consequently, a hypothesis has been put forward involving a tetrapolar phytochrome gradient at the plasmalemma. This presumed pigment pattern precisely controls chloroplast reorientation in the low-irradiance response. Intriguingly, a blue-light absorbing pigment is expressed in Mougeotia as well, which also mediates low-irradiance response via a presumed tetrapolar gradient, apparently independent of the phytochrome. Two hypotheses for the controlling mechanism of chloroplast reorientation have been put forward:

• a) Coupling of the influx of calcium through the plasmalemma to the tetrapolar gradient of the sensor pigment proper, resulting in a tetrapolar gradient of calcium in the cytoplasm. This is the “reorientation via calcium” hypothesis.
• b) Coupling of actin anchorage sites on the plasmalemma to the tetrapolar gradient of the sensor pigment proper, resulting in a tetrapolar gradient of actin anchorage sites. Cytoplasmic calcium, released from internal stores or taken up through the plasmalemma, triggers actomyosin interaction. This is the “reorientation via anchorage sites” hypothesis.

Consistent with the latter hypothesis, photoregulation by two steps seems to be indicated, (i) cytoplasmic initiation of actomyosin interaction, (ii) the graded formation of plasmalemma anchorage sites for actin filaments.     

Restoration of Seminatural Grasslands: What is the Impact on Ants?

The number of species‐rich seminatural grasslands in Northern Europe has decreased significantly due to the abandonment of traditional land use practices. To preserve these habitats, an increasing number of abandoned and overgrown grasslands have been restored by cutting down trees and shrubs and reintroducing grazing. These practices are considered a useful tool to recover the species richness of vascular plants, but their impact on other taxa is hardly known. Here we studied ants as one important group of grassland insects. We investigated (1) the effects of restoration of nongrazed and afforested seminatural grasslands, compared to continuously managed reference sites; and (2) the modulating impacts of habitat characteristics and time elapsed since restoration. We found a total of 27 ant species, 11 of these were characteristic of open habitats and seven characteristic of forests. Neither species richness per site nor the number of open‐habitat species, nor the number of forest species differed between restored and reference sites. Yet, within the restored sites, the total species richness and the number of open‐habitat species was positively related to the time since restoration and the percentage of bare rock. High frequencies of most open‐habitat species were associated with low vegetation, older restored sites, and reference sites. Most forest species showed their highest frequencies in tree‐ and shrub‐dominated habitat. We conclude that restoration efforts have been successful in terms of retrieving species richness. A regular and moderate grazing regime subsequent to the restoration is suggested in order to support a high abundance of open‐habitat species.