Space and time on the membrane: modelling Type VI secretion system dynamics as a state-dependent random walk.

The type six secretion system (T6SS) is a transmembrane protein complex that mediates bacterial cell killing. The T6SS comprises three main components (transmembrane, baseplate and sheath/tube complexes) that are sequentially assembled in order to enable an attacking cell to transport payloads into neighbouring cells. A T6SS attack disrupts the function of essential cellular components of target cells, typically resulting in their death. While the assembled T6SS adopts a fixed position in the cell membrane of the attacking cell, the location of the firing site varies between firing events. In Serratia marcescens, a post-translational regulatory network regulates the assembly and firing kinetics of the T6SS in a manner that affects the attacking cell's ability to kill target cells. Moreover, when the ability of membrane complexes to reorient is reduced, an attacking cell's competitiveness is also reduced. In this study, we will develop a mathematical model that describes both the spatial motion and assembly/disassembly of a firing T6SS. The model represents the motion of a T6SS on the cell membrane as a state-dependent random walk. Using the model, we will explore how both spatial and temporal effects can combine to give rise to different firing phenotypes. Using parameters inferred from the available literature, we show that variation in estimated diffusion coefficients is sufficient to give rise to either spatially local or global firers.

Autoregulation of Transcription and Translation: A Qualitative Analysis.

The regulation of both mRNA transcription and translation by down-stream gene products allows for a range of rich dynamical behaviours (e.g. homeostatic, oscillatory, excitability and intermittent solutions). Here, qualitative analysis is applied to an existing model of a gene regulatory network in which a protein dimer inhibits its own transcription and upregulates its own translation rate. It is demonstrated that the model possesses a unique steady state, conditions are derived under which limit cycle solutions arise and estimates are provided for the oscillator period in the limiting case of a relaxation oscillator. The analysis demonstrates that oscillations can arise only if mRNA is more stable than protein and the effect of nonlinear translation inhibition is sufficiently strong. Moreover, it is shown that the oscillation period can vary non-monotonically with transcription rate. Thus the proposed framework can provide an explanation for observed species-specific dependency of segmentation clock period on Notch signalling activity. Finally, this study facilitates the application of the proposed model to more general biological settings where post transcriptional regulation effects are likely important.

What Are the Effects of Moso Bamboo Expansion into Japanese Cedar on Arbuscular Mycorrhizal Fungi: Altering the Community Composition Rather than the Diversity.

The unbridled expansion of moso bamboo (Phyllostachys edulis) occurs throughout the world and has a series of consequences. However, the effect of bamboo expansion on arbuscular mycorrhizal fungi (AMF) is still poorly understood. We assessed the changes in the AMF community during bamboo expansion into Japanese cedar (Cryptomeria japonica) forests by analyzing AMF in three forest types-Japanese cedar (JC), bamboo-cedar mixed (BC) and moso bamboo (MB)-using 454 pyrosequencing technology. We found that the AMF community composition differed significantly among forest types. The relative abundance of Glomerales decreased from 74.0% in JC to 61.8% in BC and 42.5% in MB, whereas the relative abundance of Rhizophagus increased from 24.9% in JC to 35.9% in BC and 56.7% in MB. Further analysis showed that soil characteristics explained only 19.2% of the AMF community variation among forest types. Hence, vegetation is presumably the main driver of the alteration of the AMF community. The α diversity of AMF was similar between JC and MB, although it was higher in BC. Overall, this research sheds more light on AMF community dynamics during moso bamboo expansion. Our results highlight that the consequences of bamboo expansion in monoculture forests differ from those in mixed forests.

Auto-Regulation of Transcription and Translation: Oscillations, Excitability and Intermittency.

Several members of the Hes/Her family, conserved targets of the Notch signalling pathway, encode transcriptional repressors that dimerise, bind DNA and self-repress. Such autoinhibition of transcription can yield homeostasis and, in the presence of delays that account for processes such as transcription, splicing and transport, oscillations. Whilst previous models of autoinhibition of transcription have tended to treat processes such as translation as being unregulated (and hence linear), here we develop and explore a mathematical model that considers autoinhibition of transcription together with nonlinear regulation of translation. It is demonstrated that such a model can yield, in the absence of delays, nonlinear dynamical behaviours such as excitability, homeostasis, oscillations and intermittency. These results indicate that regulation of translation as well as transcription allows for a much richer range of behaviours than is possible with autoregulation of transcription alone. A number of experiments are suggested that would that allow for the signature of autoregulation of translation as well as transcription to be experimentally detected in a Notch signalling system.

CDK1 and CDK2 regulate NICD1 turnover and the periodicity of the segmentation clock.

All vertebrates share a segmented body axis. Segments form from the rostral end of the presomitic mesoderm (PSM) with a periodicity that is regulated by the segmentation clock. The segmentation clock is a molecular oscillator that exhibits dynamic clock gene expression across the PSM with a periodicity that matches somite formation. Notch signalling is crucial to this process. Altering Notch intracellular domain (NICD) stability affects both the clock period and somite size. However, the mechanism by which NICD stability is regulated in this context is unclear. We identified a highly conserved site crucial for NICD recognition by the SCF E3 ligase, which targets NICD for degradation. We demonstrate both CDK1 and CDK2 can phosphorylate NICD in the domain where this crucial residue lies and that NICD levels vary in a cell cycle-dependent manner. Inhibiting CDK1 or CDK2 activity increases NICD levels both in vitro and in vivo, leading to a delay of clock gene oscillations and an increase in somite size.

Response of ecosystem CO2 fluxes to grazing intensities - a five-year experiment in the Hulunber meadow steppe of China.

Grazing is the primary land use in the Hulunber meadow steppe. However, the quantitative effects of grazing on ecosystem carbon dioxide (CO2) fluxes in this zone remain unclear. A controlled experiment was conducted from 2010 to 2014 to study the effects of six stocking rates on CO2 flux, and the results showed that there were significant differences in CO2 fluxes by year, treatment, and month. The effects of light and intermediate grazing remained relatively constant with grazing year, whereas the effects of heavy grazing increased substantially with grazing duration. CO2 flux significantly decreased with increasing grazing intensity and duration, and it was significantly positively correlated with rainfall, soil moisture (SM), the carbon to nitrogen ratio (C/N ratio), soil available phosphorus (SAP), soil NH4+-N, soil NO3-N, aboveground biomass (AGB), coverage, height, and litter and negatively correlated with air temperature, total soil N (TN) and microbial biomass N (MBN). A correspondence analysis showed that the main factors influencing changes in CO2 emissions under grazing were AGB, height, coverage, SM, NH4+-N and NO3-N. Increased rainfall and reduced grazing resulted in greater CO2 emissions. Our study provides important information to improve our understanding of the role of livestock grazing in GHG emissions.

Temporal Ordering of Dynamic Expression Data from Detailed Spatial Expression Maps.

During somitogenesis, pairs of epithelial somites form in a progressive manner, budding off from the anterior end of the pre-somitic mesoderm (PSM) with a strict species-specific periodicity. The periodicity of the process is regulated by a molecular oscillator, known as the "segmentation clock," acting in the PSM cells. This clock drives the oscillatory patterns of gene expression across the PSM in a posterior-anterior direction. These so-called clock genes are key components of three signaling pathways: Wnt, Notch, and fibroblast growth factor (FGF). In addition, Notch signaling is essential for synchronizing intracellular oscillations in neighboring cells. We recently gained insight into how this may be mechanistically regulated. Upon ligand activation, the Notch receptor is cleaved, releasing the intracellular domain (NICD), which moves to the nucleus and regulates gene expression. NICD is highly labile, and its phosphorylation-dependent turnover acts to restrict Notch signaling. The profile of NICD production (and degradation) in the PSM is known to be oscillatory and to resemble that of a clock gene. We recently reported that both the Notch receptor and the Delta ligand, which mediate intercellular coupling, themselves exhibit dynamic expression at both the mRNA and protein levels. In this article, we describe the sensitive detection methods and detailed image analysis tools that we used, in combination with the computational modeling that we designed, to extract and overlay expression data from distinct points in the expression cycle. This allowed us to construct a spatio-temporal picture of the dynamic expression profile for the receptor, the ligand, and the Notch target clock genes throughout an oscillation cycle. Here, we describe the protocols used to generate and culture the PSM explants, as well as the procedure to stain for the mRNA or protein. We also explain how the confocal images were subsequently analyzed and temporally ordered computationally to generate ordered sequences of clock expression snapshots, hereafter defined as "kymographs," for the visualization of the spatiotemporal expression of Delta-like1 (Dll1) and Notch1 throughout the PSM.

Soil resilience and recovery: rapid community responses to management changes.

BACKGROUND AND AIMS: Soil degradation is a major global problem; to investigate the potential for recovery of soil biota and associated key functions, soils were monitored during the early years of conversion between permanent grassland, arable cropping and bare fallow (maintained by regular tilling). Distinct differences in soil properties had become apparent 50 years after a previous conversion. METHODS: Subplots on previously permanent grassland, arable and bare fallow soil were converted to the two alternatives, generating 9 treatments. Soil properties (soil organic carbon, mesofauna, microbial community structure and activity) were measured. RESULTS: After 2 years, mesofauna and microbial abundance increased where plants were grown on previously bare fallow soils and declined where grassland was converted to bare fallow treatment. Overall prokaryote community composition remained more similar to the previous treatments of the converted plots than to the new treatments but there were significant changes in the relative abundance of some groups and functional genes. Four years after conversion, SOC in arable and bare fallow soils converted to grassland had increased significantly. CONCLUSIONS: Conversion to permanent grassland effectively replenished C in previously degraded soil; the soil microbiome showed significant conversion-related changes; plant-driven recovery was quicker than C loss in the absence of plants.

HIV-1 capsid uncoating initiates after the first strand transfer of reverse transcription.

BACKGROUND: Correct disassembly of the HIV-1 capsid shell, called uncoating, is increasingly recognised as central for multiple steps during retroviral replication. However, the timing, localisation and mechanism of uncoating are poorly understood and progress in this area is hampered by difficulties in measuring the process. Previous work suggested that uncoating occurs soon after entry of the viral core into the cell, but recent studies report later uncoating, at or in the nucleus. Furthermore, inhibiting reverse transcription delays uncoating, linking these processes. RESULTS: Here, we have used a combined approach of experimental interrogation of viral mutants and mathematical modelling to investigate the timing of uncoating with respect to reverse transcription. By developing a minimal, testable, model and employing multiple uncoating assays to overcome the disadvantages of each single assay, we find that uncoating is not concomitant with the initiation of reverse transcription. Instead, uncoating appears to be triggered once reverse transcription reaches a certain stage, namely shortly after first strand transfer. CONCLUSIONS: Using multiple approaches, we have identified a point during reverse transcription that induces uncoating of the HIV-1 CA shell. We propose that uncoating initiates after the first strand transfer of reverse transcription.

Impact of Grassland Reseeding, Herbicide Spraying and Ploughing on Diversity and Abundance of Soil Arthropods.

In order to determine the interactive effect of reseeding, herbicide spraying and ploughing on soil fauna communities, we conducted a grassland reseeding experiment combined with pre-reseed management to examine how with the whole reseeding process affects soil faunal composition. Sampling occasions and exact treatments were as follows: (1) before chemical herbicide spray; (2) after spray but before ploughing; (3) after ploughing but before reseeding; and (4) after 1 year of recovery. Our results demonstrate that, Acari and Collembola were the two soil fauna taxa with the highest abundance and accounted for around 96% of the relative total abundance among the various managements. Herbicide application tended to increase soil invertebrate abundance. Conversely, subsequent ploughing significantly reduced soil invertebrate abundance and had an obvious negative effect on soil primary and secondary decomposers, which were mainly due to the variations of Acari (especially Oribatida) and Coleoptera group abundance. Moreover, reseeding also reduced the individual number of the groups mentioned above, and favored those predators with a larger body size and individual weight. After 1 year recovery, Collembola abundance recovered to the pre-treatment levels, while with Arthropod and Acari groups were still fluctuating.

Arbuscular mycorrhizal fungi in soil and roots respond differently to phosphorus inputs in an intensively managed calcareous agricultural soil.

Understanding the diversity and community structure of arbuscular mycorrhizal fungi (AMF) is important for potentially optimizing their role in mining phosphorus (P) in agricultural ecosystems. Here, we conduct a comprehensive study to investigate the vertical distribution of AMF in a calcareous field and their temporal structure in maize-roots with fertilizer P application over a three-year period. The results showed that soil available-P response to P fertilization but maize yields did not. Phosphorus fertilization had no-significant effect on richness of AMF except at greater soil-depths. High P-supply reduced root colonization while optimum-P tended to increase colonization and fungal richness on all sampling occasions. Crop phenology might override P-supply in determining the community composition of active root inhabiting fungi. Significant differences in the community structure of soil AMF were observed between the controls and P treatments in surface soil and the community shift was attributable mainly to available-P, N/P and pH. Vertical distribution was related mainly to soil electrical conductivity and Na content. Our results indicate that the structure of AMF community assemblages is correlated with P fertilization, soil depth and crop phenology. Importantly, phosphorus management must be integrated with other agricultural-practices to ensure the sustainability of agricultural production in salinized soils.

DNA double-strand break repair: a theoretical framework and its application.

DNA double-strand breaks (DSBs) are formed as a result of genotoxic insults, such as exogenous ionizing radiation, and are among the most serious types of DNA damage. One of the earliest molecular responses following DSB formation is the phosphorylation of the histone H2AX, giving rise to γH2AX. Many copies of γH2AX are generated at DSBs and can be detected in vitro as foci using well-established immuno-histochemical methods. It has previously been shown that anti-γH2AX antibodies, modified by the addition of the cell-penetrating peptide TAT and a fluorescent or radionuclide label, can be used to visualize and quantify DSBs in vivo. Moreover, when labelled with a high amount of the short-range, Auger electron-emitting radioisotope, (111)In, the amount of DNA damage within a cell can be increased, leading to cell death. In this report, we develop a mathematical model that describes how molecular processes at individual sites of DNA damage give rise to quantifiable foci. Equations that describe stochastic mean behaviours at individual DSB sites are derived and parametrized using population-scale, time-series measurements from two different cancer cell lines. The model is used to examine two case studies in which the introduction of an antibody (anti-γH2AX-TAT) that targets a key component in the DSB repair pathway influences system behaviour. We investigate: (i) how the interaction between anti-γH2AX-TAT and γH2AX effects the kinetics of H2AX phosphorylation and DSB repair and (ii) model behaviour when the anti-γH2AX antibody is labelled with Auger electron-emitting (111)In and can thus instigate additional DNA damage. This work supports the conclusion that DSB kinetics are largely unaffected by the introduction of the anti-γH2AX antibody, a result that has been validated experimentally, and hence the hypothesis that the use of anti-γH2AX antibody to quantify DSBs does not violate the image tracer principle. Moreover, it provides a novel model of DNA damage accumulation in the presence of Auger electron-emitting (111)In that is supported qualitatively by the available experimental data.

MRI texture analysis of subchondral bone at the tibial plateau.

OBJECTIVES: To determine the feasibility of MRI texture analysis as a method of quantifying subchondral bone architecture in knee osteoarthritis (OA). METHODS: Asymptomatic subjects aged 20-30 (group 1, n = 10), symptomatic patients aged 40-50 (group 2, n = 10) and patients scheduled for knee replacement aged 55-85 (group 3, n = 10) underwent high spatial resolution T1-weighted coronal 3T knee MRI. Regions of interest were created in the medial (MT) and lateral (LT) tibial subchondral bone from which 20 texture parameters were calculated. T2 mapping of the tibial cartilage was performed in groups 1 and 2. Mean parameter values were compared between groups using ANOVA. Linear discriminant analysis (LDA) was used to evaluate the ability of texture analysis to classify subjects correctly. RESULTS: Significant differences in 18/20 and 12/20 subchondral bone texture parameters were demonstrated between groups at the MT and LT respectively. There was no significant difference in mean MT or LT cartilage T2 values between group 1 and group 2. LDA demonstrated subject classification accuracy of 97 % (95 % CI 91-100 %). CONCLUSION: MRI texture analysis of tibial subchondral bone may allow detection of alteration in subchondral bone architecture in OA. This has potential applications in understanding OA pathogenesis and assessing response to treatment. KEY POINTS: • Improved techniques to monitor OA disease progression and treatment response are desirable • Subchondral bone (SB) may play significant role in the development of OA • MRI texture analysis is a method of quantifying changes in SB architecture • Pilot study showed that this technique is feasible and reliable • Significant differences in SB texture were demonstrated between individuals with/without OA.

Quantitative analysis of tibial subchondral bone: Texture analysis outperforms conventional trabecular microarchitecture analysis.

BACKGROUND: The aim of this study was to compare two different methods of quantitative assessment of tibial subchondral bone in osteoarthritis (OA): statistical texture analysis (sTA) and trabecular microarchitecture analysis (tMA). METHODS: Asymptomatic controls aged 20-30 (n = 10), patients aged 40-50 with chronic knee pain but without established OA (n = 10) and patients aged 55-85 with advanced OA scheduled for knee replacement (n = 10) underwent knee MR imaging at 3 Tesla with a three-dimensional gradient echo sequence to allow sTA and tMA. tMA and sTA features were calculated using region of interest creation in the medial (MT) and lateral (LT) tibial subchondral bone. Features were compared between groups using one-way analysis of variance. The two most discriminating tMA and sTA features were used to construct exploratory discriminant functions to assess the ability of the two methods to classify participants. RESULTS: No tMA features were significantly different between groups at either MT or LT. 17/20 and 11/20 sTA features were significantly different between groups at the MT/LT, respectively (P < 0.001). Discriminant functions created using tMA features classified 12/30 participants correctly (40% accuracy; 95% confidence interval [CI], 22-58%) based on MT data and 9/30 correctly (30%,; 95% CI, 14-46) based on LT data. Discriminant functions using sTA features classified 16/30 participants correctly (53%; 95% CI, 35-71) based on MT data and 14/30 correctly (47%; 95% CI, 29-65) based on LT data. CONCLUSION: sTA features showed more significant differences between the three study groups and improved classification accuracy compared with tMA features.

A balance of positive and negative regulators determines the pace of the segmentation clock.

Somitogenesis is regulated by a molecular oscillator that drives dynamic gene expression within the pre-somitic mesoderm. Previous mathematical models of the somitogenesis clock that invoke the mechanism of delayed negative feedback predict that its oscillation period depends on the sum of delays inherent to negative-feedback loops and inhibitor half-lives. We develop a mathematical model that explores the possibility that positive feedback also plays a role in determining the period of clock oscillations. The model predicts that increasing the half-life of the positive regulator, Notch intracellular domain (NICD), can lead to elevated NICD levels and an increase in the oscillation period. To test this hypothesis, we investigate a phenotype induced by various small molecule inhibitors in which the clock is slowed. We observe elevated levels and a prolonged half-life of NICD. Reducing NICD production rescues these effects. These data provide the first indication that tight control of the turnover of positive as well as negative regulators of the clock determines its periodicity.

Breast cancer screening and diagnosis in the 21st century within the U.K.

In the UK, breast cancer represents the most commonly diagnosed cancer in females and carries with it a significant morbidity and mortality. An established national screening programme is in place to identify those with the disease at an early stage in order to optimise treatment and prognosis. This article provides an overview for the non-breast specialist clinician whose practice regularly deals with women who may be the subject of the screening programme. It outlines the diagnostic methods employed in the screening programme, the controversies that have surrounded it and the evolving technologies that will improve detection and diagnostic accuracy.

Organ-level quorum sensing directs regeneration in hair stem cell populations.

Coordinated organ behavior is crucial for an effective response to environmental stimuli. By studying regeneration of hair follicles in response to patterned hair plucking, we demonstrate that organ-level quorum sensing allows coordinated responses to skin injury. Plucking hair at different densities leads to a regeneration of up to five times more neighboring, unplucked resting hairs, indicating activation of a collective decision-making process. Through data modeling, the range of the quorum signal was estimated to be on the order of 1 mm, greater than expected for a diffusible molecular cue. Molecular and genetic analysis uncovered a two-step mechanism, where release of CCL2 from injured hairs leads to recruitment of TNF-α-secreting macrophages, which accumulate and signal to both plucked and unplucked follicles. By coupling immune response with regeneration, this mechanism allows skin to respond predictively to distress, disregarding mild injury, while meeting stronger injury with full-scale cooperative activation of stem cells.

Spatiotemporal oscillations of Notch1, Dll1 and NICD are coordinated across the mouse PSM.

During somitogenesis, epithelial somites form from the pre-somitic mesoderm (PSM) in a periodic manner. This periodicity is regulated by a molecular oscillator, known as the 'segmentation clock', that is characterised by an oscillatory pattern of gene expression that sweeps the PSM in a caudal-rostral direction. Key components of the segmentation clock are intracellular components of the Notch, Wnt and FGF pathways, and it is widely accepted that intracellular negative-feedback loops regulate oscillatory gene expression. However, an open question in the field is how intracellular oscillations are coordinated, in the form of spatiotemporal waves of expression, across the PSM. In this study, we provide a potential mechanism for this process. We show at the mRNA level that the Notch1 receptor and Delta-like 1 (Dll1) ligand vary dynamically across the PSM of both chick and mouse. Remarkably, we also demonstrate similar dynamics at the protein level; hence, the pathway components that mediate intercellular coupling themselves exhibit oscillatory dynamics. Moreover, we quantify the dynamic expression patterns of Dll1 and Notch1, and show they are highly correlated with the expression patterns of two known clock components [Lfng mRNA and the activated form of the Notch receptor (cleaved Notch intracellular domain, NICD)]. Lastly, we show that Notch1 is a target of Notch signalling, whereas Dll1 is Wnt regulated. Regulation of Dll1 and Notch1 expression thus links the activity of Wnt and Notch, the two main signalling pathways driving the clock.

Spatiotemporal changes in arbuscular mycorrhizal fungal communities under different nitrogen inputs over a 5-year period in intensive agricultural ecosystems on the North China Plain.

Appropriate nitrogen (N) management is important to minimize N losses from intensively managed agricultural ecosystems. Understanding the community structure of arbuscular mycorrhizal fungi (AMF) in response to N management can be of great ecological significance, particularly with the recent emphasis on the role of AMF in N cycling. A comprehensive study of both the vertical distribution of AMF in the soil profile and the temporal changes in community structure in maize roots was conducted over a 5-year period at a field site on the North China Plain. The N treatments consisted of zero N, conventional farming practice, and optimum N based on an in-season soil Nmin test. Terminal restriction fragment length polymorphism and clone sequencing were used to analyse the AMF community. Optimum N mitigated the decline in richness of AMF in the conventional N treatment in the surface soil. Diverse and species-rich AMF communities occurred deep in the soil profile. A significant difference in AMF community structure was observed between the control and fertilizer N treatments but not between the two N application strategies. AMF communities deeper in the soil profile were subsets of those richer communities in the surface soil and the loss of AMF taxa was mostly due to the absence of rare taxa. Soil pH and Nmin contents were major soil properties affecting the soil AMF communities among the N treatments while vertical distribution was influenced mainly by soil electrical conductivity. Crop phenology had a stronger influence than N treatment on the temporal shifts in AMF communities in maize roots. Our results provide evidence for the importance of N management in maintaining AMF diversity. Changes in soil chemical properties due to N fertilization, in particular declining soil pH, should be integrated in N management strategies to reduce the negative impacts on AMF communities induced by N fertilization. Excessive N inputs induced significant changes in soil physicochemical properties, especially soil acidification, and may have negative impacts on AMF communities.

Regenerative hair waves in aging mice and extra-follicular modulators follistatin, dkk1, and sfrp4.

Hair cycling is modulated by factors both intrinsic and extrinsic to hair follicles. Cycling defects lead to conditions such as aging-associated alopecia. Recently, we demonstrated that mouse skin exhibits regenerative hair waves, reflecting a coordinated regenerative behavior in follicle populations. Here, we use this model to explore the regenerative behavior of aging mouse skin. Old mice (>18 months) tracked over several months show that with progressing age, hair waves slow down, wave propagation becomes restricted, and hair cycle domains fragment into smaller domains. Transplanting aged donor mouse skin to a young host can restore donor cycling within a 3 mm range of the interface, suggesting that changes are due to extracellular factors. Therefore, hair stem cells in aged skin can be reactivated. Molecular studies show that extra-follicular modulators Bmp2, Dkk1, and Sfrp4 increase in early anagen. Further, we identify follistatin as an extra-follicular modulator, which is highly expressed in late telogen and early anagen. Indeed, follistatin induces hair wave propagation and its level decreases in aging mice. We present an excitable medium model to simulate the cycling behavior in aging mice and illustrate how the interorgan macroenvironment can regulate the aging process by integrating both "activator" and "inhibitor" signals.