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.

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.

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.

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.

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.

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.

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.

Modelling Delta-Notch perturbations during zebrafish somitogenesis.

The discovery over the last 15 years of molecular clocks and gradients in the pre-somitic mesoderm of numerous vertebrate species has added significant weight to Cooke and Zeeman's 'clock and wavefront' model of somitogenesis, in which a travelling wavefront determines the spatial position of somite formation and the somitogenesis clock controls periodicity (Cooke and Zeeman, 1976). However, recent high-throughput measurements of spatiotemporal patterns of gene expression in different zebrafish mutant backgrounds allow further quantitative evaluation of the clock and wavefront hypothesis. In this study we describe how our recently proposed model, in which oscillator coupling drives the propagation of an emergent wavefront, can be used to provide mechanistic and testable explanations for the following observed phenomena in zebrafish embryos: (a) the variation in somite measurements across a number of zebrafish mutants; (b) the delayed formation of somites and the formation of 'salt and pepper' patterns of gene expression upon disruption of oscillator coupling; and (c) spatial correlations in the 'salt and pepper' patterns in Delta-Notch mutants. In light of our results, we propose a number of plausible experiments that could be used to further test the model.

Modelling hair follicle growth dynamics as an excitable medium.

The hair follicle system represents a tractable model for the study of stem cell behaviour in regenerative adult epithelial tissue. However, although there are numerous spatial scales of observation (molecular, cellular, follicle and multi follicle), it is not yet clear what mechanisms underpin the follicle growth cycle. In this study we seek to address this problem by describing how the growth dynamics of a large population of follicles can be treated as a classical excitable medium. Defining caricature interactions at the molecular scale and treating a single follicle as a functional unit, a minimal model is proposed in which the follicle growth cycle is an emergent phenomenon. Expressions are derived, in terms of parameters representing molecular regulation, for the time spent in the different functional phases of the cycle, a formalism that allows the model to be directly compared with a previous cellular automaton model and experimental measurements made at the single follicle scale. A multi follicle model is constructed and numerical simulations are used to demonstrate excellent qualitative agreement with a range of experimental observations. Notably, the excitable medium equations exhibit a wider family of solutions than the previous work and we demonstrate how parameter changes representing altered molecular regulation can explain perturbed patterns in Wnt over-expression and BMP down-regulation mouse models. Further experimental scenarios that could be used to test the fundamental premise of the model are suggested. The key conclusion from our work is that positive and negative regulatory interactions between activators and inhibitors can give rise to a range of experimentally observed phenomena at the follicle and multi follicle spatial scales and, as such, could represent a core mechanism underlying hair follicle growth.

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.

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.

Using stable isotopes to differentiate trophic feeding channels within soil food webs.

The soil is probably the most diverse habitat there is, with organisms ranging in sizes from less than 1 µm to several metres in length. However, it is increasingly evident that we know little about the interactions occurring between these organisms, the functions that they perform as individual species, or together within their different feeding guilds. These interactions between groups of organisms and physical and chemical processes shape the soil as a habitat and influence the nature of the soil food web with consequences for the above-ground vegetation and food web. Protists are known as one of the most abundant groups of bacterivores within the soil; however, they are also consumers of a number of other food sources. Even though they are responsible for a large proportion of the mineralisation of bacterial biomass and have a large impact on the C and N cycles within the soil they are regularly overlooked when investigating the complete soil food web. Recently, stable isotopes have been used to determine trophic interactions and here we describe how this technique has been used to highlight linkages between protists and the soil food web.

Protozoan pulses unveil their pivotal position within the soil food web.

Protozoa are one of the most abundant groups of bacterivores within the soil and are responsible for mineralisation of bacterial biomass, having a large impact on C and N cycling. Little is known of their contribution to soil nutrient transfers or the identity of their consumers. Here, for the first time indigenous flagellates and ciliates, enriched to 83 atom% for (13)C and 10 atom% for (15)N, were introduced to soil cores from two different land managements, grassland and woodland with the same soil type, to trace the flow of protozoan C and N through the soil food web. Nematodes, Collembola, earthworms and insect larvae obtained the greatest amounts of C and N of protozoan origin, either through direct consumption or uptake of biomass post-cell death. Our results show that changes in management, affect the functioning of the soil food web and the utilisation of protozoa as a food source.

Comparing a discrete and continuum model of the intestinal crypt.

The integration of processes at different scales is a key problem in the modelling of cell populations. Owing to increased computational resources and the accumulation of data at the cellular and subcellular scales, the use of discrete, cell-level models, which are typically solved using numerical simulations, has become prominent. One of the merits of this approach is that important biological factors, such as cell heterogeneity and noise, can be easily incorporated. However, it can be difficult to efficiently draw generalizations from the simulation results, as, often, many simulation runs are required to investigate model behaviour in typically large parameter spaces. In some cases, discrete cell-level models can be coarse-grained, yielding continuum models whose analysis can lead to the development of insight into the underlying simulations. In this paper we apply such an approach to the case of a discrete model of cell dynamics in the intestinal crypt. An analysis of the resulting continuum model demonstrates that there is a limited region of parameter space within which steady-state (and hence biologically realistic) solutions exist. Continuum model predictions show good agreement with corresponding results from the underlying simulations and experimental data taken from murine intestinal crypts.

The clock and wavefront model revisited.

The currently accepted interpretation of the clock and wavefront model of somitogenesis is that a posteriorly moving molecular gradient sequentially slows the rate of clock oscillations, resulting in a spatial readout of temporal oscillations. However, while molecular components of the clocks and wavefronts have now been identified in the pre-somitic mesoderm (PSM), there is not yet conclusive evidence demonstrating that the observed molecular wavefronts act to slow clock oscillations. Here we present an alternative formulation of the clock and wavefront model in which oscillator coupling, already known to play a key role in oscillator synchronisation, plays a fundamentally important role in the slowing of oscillations along the anterior-posterior (AP) axis. Our model has three parameters which can be determined, in any given species, by the measurement of three quantities: the clock period in the posterior PSM, somite length and the length of the PSM. A travelling wavefront, which slows oscillations along the AP axis, is an emergent feature of the model. Using the model we predict: (a) the distance between moving stripes of gene expression; (b) the number of moving stripes of gene expression and (c) the oscillator period profile along the AP axis. Predictions regarding the stripe data are verified using existing zebrafish data. We simulate a range of experimental perturbations and demonstrate how the model can be used to unambiguously define a reference frame along the AP axis. Comparing data from zebrafish, chick, mouse and snake, we demonstrate that: (a) variation in patterning profiles is accounted for by a single nondimensional parameter; the ratio of coupling strengths; and (b) the period profile along the AP axis is conserved across species. Thus the model is consistent with the idea that, although the genes involved in pattern propagation in the PSM vary, there is a conserved patterning mechanism across species.

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.

Modelling spatially regulated beta-catenin dynamics and invasion in intestinal crypts.

Experimental data (e.g., genetic lineage and cell population studies) on intestinal crypts reveal that regulatory features of crypt behavior, such as control via morphogen gradients, are remarkably well conserved among numerous organisms (e.g., from mouse and rat to human) and throughout the different regions of the small and large intestines. In this article, we construct a partial differential equation model of a single colonic crypt that describes the spatial distribution of Wnt pathway proteins along the crypt axis. The novelty of our continuum model is that it is based upon assumptions that can be directly related to processes at the cellular and subcellular scales. We use the model to predict how the distributions of Wnt pathway proteins are affected by mutations. The model is then extended to investigate how mutant cell populations can invade neighboring crypts. The model simulations suggest that cell crowding caused by increased proliferation and decreased cell loss may be sufficient for a mutant cell population to colonize a neighboring healthy crypt.

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.

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.