Linking specification to differentiation:From proneural genes to the regulation of ciliogenesis

Much of developmental biology is concerned with the processes by which cells become committed to particular fates in a regulated fashion, whereas cell biology addresses, among other things, the variety of differentiated forms and functions that cells can acquire. One open question is how the regulators of the former process lead to attainment of the latter. “High-level” regulators of cell fate specification include the proneural factors, which drive cells to commit as precursors in the sensory nervous system. Recent research has concentrated on the gene expression events downstream of proneural factor function. Here we summarize this research and describe our own research that has provided clear links between a proneural factor, atonal and the cell biological program of ciliogenesis, which is a central aspect of sensory neuron differentiation

Positive autoregulation of the transcription factor Pax6 in response to increased levels of either of its major isoforms, Pax6 or Pax6(5a), in cultured cells

<p>Abstract</p> <p>Background</p> <p>Pax6 is a transcription factor essential for normal development of the eyes and nervous system. It has two major isoforms, Pax6 and Pax6(5a), and the ratios between their expression levels vary within narrow limits. We tested the effects of overexpressing either one or other isoform on endogenous Pax6 expression levels in Neuro2A and NIH3T3 cells.</p> <p>Results</p> <p>We found that both isoforms caused an up-regulation of endogenous Pax6 expression in cells with (Neuro2A) or without (NIH3T3) constitutive Pax6 expression. Western blots showed that cells stably transfected with constructs expressing either Pax6 or Pax6(5a) contained raised levels of both Pax6 and Pax6(5a). Quantitative RT-PCR confirmed an increase in levels of <it>Pax6(5a) </it>mRNA in cells containing Pax6-expressing constructs and an increase in levels of <it>Pax6 </it>mRNA in cells containing Pax6(5a)-expressing constructs. The fact that the introduction of constructs expressing only one isoform increased the cellular levels of not only that isoform but also the other indicates that activation of the endogenous <it>Pax6 </it>locus occurred. The ratio between the levels of the two isoforms was maintained close to physiological values. The overexpression of either isoform in neuroblastoma (Neuro2A) cell lines also promoted morphological change and an increase in β-III-tubulin expression, indicating an increase in neurogenesis.</p> <p>Conclusion</p> <p>Our results demonstrate that Pax6 can up-regulate production of Pax6 protein from an entire intact endogenous <it>Pax6 </it>locus in its genomic environment. This adds to previous studies showing that Pax6 can up-regulate reporter expression driven by isolated <it>Pax6 </it>regulatory elements. Furthermore, our results suggest that an important function of positive feedback might be to stabilise the relative levels of Pax6 and Pax6(5a).</p

Comparison of rule- and ordinary differential equation-based dynamic model of DARPP-32 signalling network

Dynamic modelling has considerably improved our understanding of complex molecular mechanisms. Ordinary differential equations (ODEs) are the most detailed and popular approach to modelling the dynamics of molecular systems. However, their application in signalling networks, characterised by multi-state molecular complexes, can be prohibitive. Contemporary modelling methods, such as rule- based (RB) modelling, have addressed these issues. The advantages of RB modelling over ODEs have been presented and discussed in numerous reviews. In this study, we conduct a direct comparison of the time courses of a molecular system founded on the same reaction network but encoded in the two frameworks. To make such a comparison, a set of reactions that underlie an ODE model was manually encoded in the Kappa language, one of the RB implementations. A comparison of the models was performed at the level of model specification and dynamics, acquired through model simulations. In line with previous reports, we confirm that the Kappa model recapitulates the general dynamics of its ODE counterpart with minor differences. These occur when molecules have multiple sites binding the same interactor. Furthermore, activation of these molecules in the RB model is slower than in the ODE one. As reported for other molecular systems, we find that, also for the DARPP-32 reaction network, the RB representation offers a more expressive and flexible syntax that facilitates access to fine details of the model, easing model reuse. In parallel with these analyses, we report a refactored model of the DARPP-32 interaction network that can serve as a canvas for the development of more complex dynamic models to study this important molecular system

Improved prediction of hiking speeds using a data driven approach

Hikers and hillwalkers typically use the gradient in the direction of travel (walking slope) as the main variable in established methods for predicting walking time (via the walking speed) along a route. Research into fell-running has suggested further variables which may improve speed algorithms in this context; the gradient of the terrain (hill slope) and the level of terrain obstruction. Recent improvements in data availability, as well as widespread use of GPS tracking now make it possible to explore these variables in a walking speed model at a sufficient scale to test statistical significance. We tested various established models used to predict walking speed against public GPS data from almost 88,000 km of UK walking / hiking tracks. Tracks were filtered to remove breaks and non-walking sections. A new generalised linear model (GLM) was then used to predict walking speeds. Key differences between the GLM and established rules were that the GLM considered the gradient of the terrain (hill slope) irrespective of walking slope, as well as the terrain type and level of terrain obstruction in off-road travel. All of these factors were shown to be highly significant, and this is supported by a lower root-mean-square-error compared to existing functions. We also observed an increase in RMSE between the GLM and established methods as hill slope increases, further supporting the importance of this variable.Comment: 37 pages, 13 figure

Normal ventral telencephalic expression of Pax6 is required for normal development of thalamocortical axons in embryonic mice

<p>Abstract</p> <p>Background</p> <p>In addition to its well-known expression in dorsal telencephalic progenitor cells, where it regulates cell proliferation and identity, the transcription factor Pax6 is expressed in some ventral telencephalic cells, including many postmitotic neurons. Its functions in these cells are unknown.</p> <p>Results</p> <p>We generated a new floxed allele of <it>Pax6 </it>and tested the consequences of a highly specific ventral telencephalic depletion of Pax6. We used the <it>Six3</it>^{<it>A1A2</it>}-<it>Cre </it>allele that drives production of Cre recombinase in a specific region of Pax6-expression close to the internal capsule, through which thalamic axons navigate to cerebral cortex. Depletion in this region caused many thalamic axons to take aberrant routes, either failing to turn normally into ventral telencephalon to form the internal capsule or exiting the developing internal capsule ventrally. We tested whether these defects might have resulted from abnormalities of two structural features proposed to guide thalamic axons into and through the developing internal capsule. First, we looked for the early pioneer axons that project from the region of the future internal capsule to the thalamus and are thought to guide thalamocortical axons to the internal capsule: we found that they are present in conditional mutants. Second, we examined the development of the corridor of Islet1-expressing cells that guides thalamic axons through ventral telencephalon and found that it was broader and less dense than normal in conditional mutants. We also examined corticofugal axons that are thought to interact with ascending thalamocortical axons, resulting in each set providing guidance to the other, and found that some are misrouted to lateral telencephalon.</p> <p>Conclusion</p> <p>These findings indicate that ventral telencephalic Pax6 is important for formation of the Islet1-expressing corridor and the thalamic and cortical axons that grow through it. We suggest that Pax6 might affect thalamic axonal growth indirectly via its effect on the corridor.</p

Deep anthropogenic topsoils in Scotland : a geoarchaeological and historical investigation into distribution, character and conservation under modern land cover

Deep anthropogenic topsoils – those augmented through long-term additions of mineral bulk among fertilising agents – retain in both their physical and chemical make-up significant indicators for cultural activity. This project researched the geographical distribution and historical context of deep anthropogenic topsoils in Scotland and the Isles, and used this information to investigate the impact of current land cover upon the cultural information they retain. In so doing, the project investigated the potential for conservation of this significant cultural resource. A review of the historical information available on agricultural and manuring practices for Scotland identified several factors likely to affect deep topsoil distribution and frequency. These were: the availability of bulk manures to Scottish farmers, the significance of the seaweed resource in determining fertiliser strategies in coastal areas, and the influence of urban settlement and associated patterns of domestic and industrial waste disposal on the location of deep topsoils. Evidence for widespread deep topsoil development was limited. The primary data source used – the First Statistical Account of Scotland – was manipulated into a spatial database in ArcView GIS, to which geographical data from the Soil Survey of Scotland and national archaeological survey databases were added. This was used to devise a survey programme aiming both to investigate the potential factors affecting soil development listed above, and to locate deep topsoil sites for analysis. Three sites were identified with deep topsoils under different cover types (woodland, arable and pasture). The urban-influenced context of two of these highlighted the significance of urban settlement to the location of Scottish deep topsoils. Analysis of pH, organic matter, and total phosphorus content showed a correlation between raised organic matter and a corresponding increase in phosphorus content in soils under permanent vegetation. By contrast, soils under arable cultivation showed no such rise. This was attributed to the action of cropping in removing modern organic inputs prior to down-profile cycling. The potential for pasture and woodland cover to affect relict soil signatures was therefore observed. Thin section analysis aimed to both provide micromorphological characterisation of the three deep topsoil sites and investigate the effect of modern land cover on micromorphological indicators. Distinctive differences in micromorphological character were observed between the rural and urban deep topsoils, with the latter showing a strong focus on carbonised fuel residues and industrial wastes. All sites showed a highly individual micromorphological character, reflective of localised fertilising systems. There was no correlation between land cover type and survival of material indictors for anthropogenic activity, with soil cultural indicators surviving well, particularly those characteristic of urban-influenced topsoils. Suggestions for preservation strategies for this potentially rare and highly localised cultural resource included the incorporation of deep anthropogenic topsoil conservation into current government policy relating to care of the rural historic environment, and the improvement of data on the resource through ongoing survey and excavation.EThOS - Electronic Theses Online ServiceHistoric Scotland : University of StirlingGBUnited Kingdo

Discussion of "Geodesic Monte Carlo on Embedded Manifolds"

Contributed discussion and rejoinder to "Geodesic Monte Carlo on Embedded Manifolds" (arXiv:1301.6064)Comment: Discussion of arXiv:1301.6064. To appear in the Scandinavian Journal of Statistics. 18 page

Regulation of the Pax6 : Pax6(5a) mRNA ratio in the developing mammalian brain

BACKGROUND: Early in mammalian brain development cell proliferation generates a population of progenitor cells whose subsequent divisions produce increasing numbers of postmitotic neurons. Pax6 affects both processes and it has been suggested that this changing role is due at least in part to changes in the relative concentrations of its two main isoforms, (i) Pax6 and (ii) Pax6(5a), created by insertion of a 42 bp exon (exon 5a) into one of the two DNA-binding domains. Crucially, however, no previous study has determined whether the ratio between Pax6 and Pax6(5a) transcripts alters during mammalian neurogenesis in vivo. RESULTS: Using RNase protection assays, we show that Pax6 transcripts are 6–10 times more prevalent than Pax6(5a) transcripts early in neurogenesis in the murine telencephalon, diencephalon and hindbrain and that the ratio later falls significantly to about 3:1 in these regions. CONCLUSION: These changes in vivo are similar in magnitude to those shown previously to alter target gene activity in vitro and might, therefore, allow the single mammalian Pax6 gene to carry out different functions at different times in mammalian brain development