Hyperosmotic Stress Enzyme Signaling Modulates Oct4, Nanog, And Rex1 Expression And Induces Prioritized Differentiation Of Murine Embryonic Stem Cells

HYPEROSMOTIC STRESS ENZYME SIGNALING MODULATES OCT4, NANOG, AND REX1 EXPRESSION AND INDUCES PRIORITIZED DIFFERENTIATION OF MURINE EMBRYONIC STEM CELLS by JILL SLATER MAY 2013 Advisor: Daniel Rappolee, Ph.D. Major: Physiology Degree: Doctor of Philosophy Transcription factor expression and therefore lineage identity in the periimplantation embryo and its stem cells may be influenced by extracellular stresses, potentially affecting pregnancy outcome. Cellular stress forces cells to suppress some normal activities (such as protein synthesis and cell proliferation) in order to repair stress-damaged macromolecules and restore homeostasis. Therefore, any new activities that embryonic cells initiate while concurrently funding the demands of the stress response reveal the developmental priorities of these cells. Previous work showed that cultured multipotent trophoblast stem cells (TSC) initiated differentiation in response to hyperosmotic stress, favoring the development of the earliest functioning placental lineage (parietal trophoblast giant cells) while suppressing that of laterdifferentiating lineages (chorionic/syncytiotrophoblast). The studies described in this dissertation studied the stress response of the other extant lineage of the early blastocyst, cells derived from the inner cell mass, murine embryonic stem cells (mESC). Hyperosmotic stress slowed mESC accumulation due to slowing of the cell cycle, not apoptosis. PI3K signaling was responsible for cell survival 136 under stressed conditions. Stress initially triggered mESC differentiation through MEK1, JNK, and PI3K signaling, leading to proteasomal degradation of OCT4, NANOG, SOX2, and REX1 protein. Concurrent with this post-transcriptional effect was the degradation of their mRNA transcripts. As stress continued, cells adapted, cell cycle resumed, and OCT4 and NANOG mRNA and protein expression returned to near normal levels. The protein recovery was mediated by p38 and PI3K signaling, as well as by that of an unknown MEK1/2 target. REX1 expression, however, did not recover; its ongoing suppression was due to JNK signaling. mESC did not overtly differentiate during stress, but were primed to differentiate toward the extraembryonic lineages, upregulating markers of primitive endoderm and suppressing epiblast markers. The studies were continued in the peri-implantation model, embryoid bodies (EBs), in which differentiation is allowed rather than actively suppressed. Unstressed EB culture recapitulated the lineage inductions of in vivo embryos. EBs were only able to be cultured in the presence of low levels of hyperosmotic stress (10mM sorbitol); higher levels led to a failure of mESC to aggregate. Aggregation and subsequent embryoid body formation was rescued when either JNK or p38 MAPKs were inhibited during mESC culture. Low levels of osmotic stress increased the magnitude of primitive endoderm markers, Lrp2 and Dab2. Transient, sub-lethal stress delivered prior to the start of hanging drop culture was remembered by mESC, suppressing differentiation events slated to occur from 1-6d later. Mesoderm marker, Brachyury, and anterior visceral endoderm marker, Goosecoid, expression was suppressed. The timing of stress delivery was very significant in determining its outcome. Hyperosmotic stress delivered at the onset of differentiation induced a prioritized differentiation of mESC, inducing the earlier-developing primitive endoderm, and strongly suppressing later137 developing mesoderm and anterior visceral endoderm

Fostering student engagement through a real-world, collaborative project across disciplines and institutions

Ample research has identified several features of a learning experience likely to enhance student learning, including collaboration, open-ended exploration, and problem-based learning in real-life scenarios. Missing is a model of how instructors might combine these elements into a single project that works flexibly across disciplines and institutions. This article fills this gap by offering such a model and reporting on its effectiveness in fostering student engagement. It describes a project that instructors at four colleges and universities in Flint, Michigan (USA) piloted during the height of the Flint water crisis. The project asked students to apply class content to the real-world problem unfolding around them, and offered students an opportunity to collaborate with peers. We collected qualitative and quantitative data on students’ reactions to the project, and found that the project succeeded in engaging students. We offer recommendations for how instructors can create similar projects in their own classrooms

Tau, XMAP215/Msps and Eb1 co-operate interdependently to regulate microtubule polymerisation and bundle formation in axons

The formation and maintenance of microtubules requires their polymerisation, but little is known about how this polymerisation is regulated in cells. Focussing on the essential microtubule bundles in axons of Drosophila and Xenopus neurons, we show that the plus-end scaffold Eb1, the polymerase XMAP215/Msps and the lattice-binder Tau co-operate interdependently to promote microtubule polymerisation and bundle organisation during axon development and maintenance. Eb1 and XMAP215/Msps promote each other's localisation at polymerising microtubule plus-ends. Tau outcompetes Eb1-binding along microtubule lattices, thus preventing depletion of Eb1 tip pools. The three factors genetically interact and show shared mutant phenotypes: reductions in axon growth, comet sizes, comet numbers and comet velocities, as well as prominent deterioration of parallel microtubule bundles into disorganised curled conformations. This microtubule curling is caused by Eb1 plus-end depletion which impairs spectraplakin-mediated guidance of extending microtubules into parallel bundles. Our demonstration that Eb1, XMAP215/Msps and Tau co-operate during the regulation of microtubule polymerisation and bundle organisation, offers new conceptual explanations for developmental and degenerative axon pathologies

Age, Ethnicity and Equalities: Synthesising Policy and Practice Messages from Two Recent Studies of Elder Abuse in the UK

Two recent studies of elder abuse in the UK are located in current policy contexts of adult safeguarding. After describing the studies, the discussion draws out their central messages and identifies the challenges that the studies present to recent policy debates and innovations. These relate to the need to properly integrate both wider older people’s issues and issues of racism and ethnicity within developments in adult safeguarding policy as well as social care services as the personalisation agenda advances

Hydrological controls on oviposition habitat are associated with egg-laying phenology of some caddisflies

1. Seasonal variation in resource availability can have strong effects on life histories and population densities. Emergent rocks (ERs) are an essential oviposition resource for multiple species of stream insects. The availability of ERs depends upon water depth and clast size, which vary with discharge and river geomorphology, respectively. Recruitment success for populations may depend on whether peak egg-laying periods occur at times when ER are also abundant. For multiple species that oviposit on ER, we tested whether seasonal fluctuations in ER abundance were concurrent with oviposition phenology. We also tested whether high discharge drowned ERs for sufficiently long periods to preclude egg laying, and whether this problem varied between rivers differing in channel morphology and particle size distribution.2. We obtained a continuous timeseries of water level (WL) measured every 30 min for two years at sites on three rivers in south-eastern Australia with similar hydrology but different geomorphology. A relationship between WL and ER numbers was determined empirically at each site and these relationships were used to predict ER availability over the two years. Egg masses of ten species of caddisflies were enumerated each month for a year in one river to establish oviposition phenology. 3. Abundance of ERs was inversely related to discharge in all three rivers. ERs were most abundant during autumn and scarce during spring. Site-specific geomorphology resulted in skewed or multimodal distributions of ER abundance each year. Between years, catchment-scale hydrometeorology mediated patterns of ER availability, despite the close proximity of sites. Temporal variance in ER availability was not consistently correlated with mean WL or WL variance. ER variance increased with WL variance, when WL was below a threshold equivalent to mean annual WL. Above this threshold, most ER were likely to be submerged.4. Oviposition phenology varied strongly among the ten species of caddisflies, with egg-laying ranging from in 1-2 months to year-round. Temporal variations in ER and egg mass abundance were not correlated for most species. Below a threshold minimum number of ER, egg masses were highly crowded onto the few available ER, which is evidence that ER were in short supply. For five species, high egg mass abundance was positively associated with periods of the year when the time above the threshold number of ERs was high. Unusually, two species laid most egg masses during winter and when the time above this threshold was short. Three species showed no association between egg mass abundance and time above this threshold; two of these species laid eggs year-round.5. Regional hydrometeorology controlled the availability of ERs, but between-river differences were sufficient to deliver different outcomes in the availability of oviposition sites between years and seasons. Caddisflies were rarely prevented from laying eggs but periods when ERs were in short supply created crowding, which may be associated with negative fitness effects on hatching larvae. Geomorphological controls on availability of oviposition resources may have strong implications for the coexistence of species that overlap temporally in egg-laying

Multi-scale Characterization and Prediction of Coupled Subsurface Biogeochemical-Hydrological Processes

To advance solutions needed for remediation of DOE contaminated sites, approaches are needed that can elucidate and predict reactions associated with coupled biological, geochemical, and hydrological processes over a variety of spatial scales and in heterogeneous environments. Our previous laboratory experimental experiments, which were conducted under controlled and homogeneous conditions, suggest that geophysical methods have the potential for elucidating system transformations that often occur during remediation. Examples include tracking the onset and aggregation of precipitates associated with sulfate reduction using seismic and complex resistivity methods (Williams et al., 2005; Ntarlagiannis et al., 2005) as well as estimating the volume of evolved gas associated with denitrification using radar velocity. These exciting studies illustrated that geophysical responses correlated with biogeochemical changes, but also that multiple factors could impact the geophysical signature and thus a better understanding as well as integration tools were needed to advance the techniques to the point where they can be used to provide quantitative estimates of system transformations

Spatial arrangement or amount? Spatially variable oviposition habitat can determine aquatic insect egg abundance

Both the amount and spatial arrangement (configurational heterogeneity) of resources can affect population abundance and community diversity via influence on the growth, survival, reproduction, recruitment and movement of species. However, in most cases, it is difficult to separate the effects of resource amount from arrangement because these two attributes are often naturally correlated. In this study, we examined the configurational heterogeneity of resources (oviposition habitat—emergent rocks, ER) within rivers and decoupled the effects of resource amount from those due to the spatial arrangement on oviposition by eight species of aquatic insects (seven caddisflies and one mayfly). To capture the configurational heterogeneity of resources in 28 sites (riffles) across multiple streams in Australia and Scotland, we calculated fractal dimensions (DB) using the box-counting technique. We then used simulated riffles to explore how numbers of ER, edginess (the proportion of ER along river margins) and patchiness (clustering of emergent rocks in the middle) separately and together affected the values of DB using asymptotic regression models. Finally, we used multiple regression to test whether the numbers of egg masses laid in natural riffles of each of the eight species were explained by the number of ER, fractal dimension or both. The distributions of ER in natural riffles were scale-independent, self-repeating patterns (i.e. they were fractal), and values of DB varied significantly among riffles. Variations in fractal dimensions among simulated riffles were significantly related to the number of ER, edginess and patchiness. However, in natural riffles, only the number of ER and patchiness affected DB. Egg mass abundances were related to the fractal dimensions of ER distributions in riffles in three species and to the number of ER in five species. The fractal dimensions of riffles are unlikely to be driven by large-scale processes but instead may result from within-riffle variability that influences rock movement, arrangement and emergence. Increased oviposition by aquatic insects in riffles with greater numbers of ER suggests that these species may be limited by the amount of oviposition resources. Of the species responding to fractal dimension, two species favoured tightly clustered ER whereas the third favoured ER in much looser clusters. It is feasible that aquatic insects can detect ER clusters from the air by responding to changes in the reflectivity of water (albedo) caused by turbulence around ER. That fractals can capture configurational heterogeneity of resources in streams suggests that this technique is useful to test for general ecological patterns across diverse stream systems. Whether configurational heterogeneity influences adults directly or indirectly, these results showed that the amount and arrangement of suitable oviposition habitat plays a role in determining egg mass densities, with potential consequences for larval densities. These patterns may have important community-level and functional consequences and hence spatial arrangements should be considered when humans manipulate these resources in rivers

Tau, XMAP215/Msps and Eb1 co-operate interdependently to regulate microtubule polymerisation and bundle formation in axons

AbstractThe formation and maintenance of microtubules requires their polymerisation, but little is known about how this polymerisation is regulated in cells. Focussing on the essential microtubule bundles in axons of Drosophila and Xenopus neurons, we show that the plus-end scaffold Eb1, the polymerase XMAP215/Msps and the lattice-binder Tau co-operate interdependently to promote microtubule polymerisation and bundle organisation during axon development and maintenance. Eb1 and XMAP215/Msps promote each other’s localisation at polymerising microtubule plus-ends. Tau outcompetes Eb1-binding along microtubule lattices, thus preventing depletion of Eb1 tip pools. The three factors genetically interact and show shared mutant phenotypes: reductions in axon growth, comet size, comet number and comet velocity, as well as prominent deterioration of parallel microtubule bundles into disorganised curled conformations. This microtubule curling is caused by Eb1 plus-end depletion which impairs spectraplakin-mediated guidance of extending microtubules into parallel bundles. Our demonstration that Eb1, XMAP215/Msps and Tau co-operate during the regulation of microtubule polymerisation and bundle organisation, offers new conceptual explanations for developmental and degenerative axon pathologies and how to treat them.Summary statementEb1, XMAP215 and tau co-operate interdependently in axons to promote the polymerisation of microtubules and their organisation into the parallel bundles required for axonal transport.</jats:sec

Tau, XMAP215/Msps and Eb1 co-operate interdependently to regulate microtubule polymerisation and bundle formation in axons

From PLOS via Jisc Publications RouterHistory: received 2021-04-23, accepted 2021-06-07, collection 2021-07, epub 2021-07-06Publication status: PublishedFunder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/I002448/1Funder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/P020151/1Funder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/L000717/1Funder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/M007553/1Funder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/M007456/1Funder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268; Grant(s): BB/R018960/1Funder: Leverhulme Trust; funder-id: http://dx.doi.org/10.13039/501100000275; Grant(s): ECF-2017-247Funder: Deutsche Forschungsgemeinschaft; funder-id: http://dx.doi.org/10.13039/501100001659; Grant(s): VO 2071/1-1Funder: National Institutes of Health; funder-id: http://dx.doi.org/10.13039/100000002; Grant(s): R01 MH109651Funder: Consejo Nacional de Innovación, Ciencia y Tecnología; funder-id: http://dx.doi.org/10.13039/501100009068Funder: Biotechnology and Biological Sciences Research Council; funder-id: http://dx.doi.org/10.13039/501100000268Funder: Wellcome Trust; funder-id: http://dx.doi.org/10.13039/100010269Funder: University of Manchester Strategic FundFunder: Wellcome Trust; funder-id: http://dx.doi.org/10.13039/100004440; Grant(s): 087742/Z/08/ZFunder: National Institutes of Health; funder-id: http://dx.doi.org/10.13039/100000002; Grant(s): P40OD018537The formation and maintenance of microtubules requires their polymerisation, but little is known about how this polymerisation is regulated in cells. Focussing on the essential microtubule bundles in axons of Drosophila and Xenopus neurons, we show that the plus-end scaffold Eb1, the polymerase XMAP215/Msps and the lattice-binder Tau co-operate interdependently to promote microtubule polymerisation and bundle organisation during axon development and maintenance. Eb1 and XMAP215/Msps promote each other’s localisation at polymerising microtubule plus-ends. Tau outcompetes Eb1-binding along microtubule lattices, thus preventing depletion of Eb1 tip pools. The three factors genetically interact and show shared mutant phenotypes: reductions in axon growth, comet sizes, comet numbers and comet velocities, as well as prominent deterioration of parallel microtubule bundles into disorganised curled conformations. This microtubule curling is caused by Eb1 plus-end depletion which impairs spectraplakin-mediated guidance of extending microtubules into parallel bundles. Our demonstration that Eb1, XMAP215/Msps and Tau co-operate during the regulation of microtubule polymerisation and bundle organisation, offers new conceptual explanations for developmental and degenerative axon pathologies

Tau, XMAP215/Msps and Eb1 co-operate interdependently to regulate microtubule polymerisation and bundle formation in axons

The formation and maintenance of microtubules requires their polymerisation, but little is known about how this polymerisation is regulated in cells. Focussing on the essential microtubule bundles in axons of Drosophila and Xenopus neurons, we show that the plus-end scaffold Eb1, the polymerase XMAP215/Msps and the lattice-binder Tau co-operate interdependently to promote microtubule polymerisation and bundle organisation during axon development and maintenance. Eb1 and XMAP215/Msps promote each other’s localisation at polymerising microtubule plus-ends. Tau outcompetes Eb1-binding along microtubule lattices, thus preventing depletion of Eb1 tip pools. The three factors genetically interact and show shared mutant phenotypes: reductions in axon growth, comet sizes, comet numbers and comet velocities, as well as prominent deterioration of parallel microtubule bundles into disorganised curled conformations. This microtubule curling is caused by Eb1 plus-end depletion which impairs spectraplakin-mediated guidance of extending microtubules into parallel bundles. Our demonstration that Eb1, XMAP215/Msps and Tau co-operate during the regulation of microtubule polymerisation and bundle organisation, offers new conceptual explanations for developmental and degenerative axon pathologies