ATF4 links ER stress with reticulophagy in glioblastoma cells

Selective degradation of the endoplasmic reticulum (ER; reticulophagy) is a type of autophagy involved in the removal of ER fragments. So far, amino acid starvation as well as ER stress have been described as inducers of reticulophagy, which in turn restores cellular energy levels and ER homeostasis. Here, we explored the autophagy-inducing mechanisms that underlie the autophagic cell death (ACD)-triggering compound loperamide (LOP) in glioblastoma cells. Interestingly, LOP triggers upregulation of the transcription factor ATF4, which is accompanied by the induction of additional ER stress markers. Notably, knockout of ATF4 significantly attenuated LOP-induced autophagy and ACD. Functionally, LOP also specifically induces the engulfment of large ER fragments within autophagosomes and lysosomes as determined by electron and fluorescence microscopy. LOP-induced reticulophagy and cell death are predominantly mediated through the reticulophagy receptor RETREG1/FAM134B and, to a lesser extent, TEX264, confirming that reticulophagy receptors can promote ACD. Strikingly, apart from triggering LOP-induced autophagy and ACD, ATF4 is also required for LOP-induced reticulophagy. These observations highlight a key role for ATF4, RETREG1 and TEX264 in response to LOP-induced ER stress, reticulophagy and ACD, and establish a novel mechanistic link between ER stress and reticulophagy, with possible implications for additional models of drug-induced ER stress

Improving MCS Enumeration via Caching

Enumeration of minimal correction sets (MCSes) of conjunctive normal form formulas is a central and highly intractable problem in infeasibility analysis of constraint systems. Often complete enumeration of MCSes is impossible due to both high computational cost and worst-case exponential number of MCSes. In such cases partial enumeration is sought for, finding applications in various domains, including axiom pinpointing in description logics among others. In this work we propose caching as a means of further improving the practical efficiency of current MCS enumeration approaches, and show the potential of caching via an empirical evaluation.Peer reviewe

Timed inhibition of CDC7 increases CRISPR-Cas9 mediated templated repair.

Repair of double strand DNA breaks (DSBs) can result in gene disruption or gene modification via homology directed repair (HDR) from donor DNA. Altering cellular responses to DSBs may rebalance editing outcomes towards HDR and away from other repair outcomes. Here, we utilize a pooled CRISPR screen to define host cell involvement in HDR between a Cas9 DSB and a plasmid double stranded donor DNA (dsDonor). We find that the Fanconi Anemia (FA) pathway is required for dsDonor HDR and that other genes act to repress HDR. Small molecule inhibition of one of these repressors, CDC7, by XL413 and other inhibitors increases the efficiency of HDR by up to 3.5 fold in many contexts, including primary T cells. XL413 stimulates HDR during a reversible slowing of S-phase that is unexplored for Cas9-induced HDR. We anticipate that XL413 and other such rationally developed inhibitors will be useful tools for gene modification

Advances in MASELTOV – Serious Games in a Mobile Ecology of Services for Social Inclusion and Empowerment of Recent Immigrants

Immigration imposes a range of challenges with the risk of social exclusion from the information society (Halfman 1998), such as, getting into communication with the local society and understanding the culture of their host nation. Failure to address these challenges can lead to difficulties in the frame of integrating into the society of the host country, leading to fragmented communities and a range of social issues. As part of a comprehensive suite of services for immigrants, the European project seeks to provide both practical tools and learning services via mobile devices, providing a readily usable resource for immigrants. We introduce recent results, such as the game-based learning aspect of the MASELTOV project is introduced, with the rationale behind its design presented. In doing so, the benefits and implications of mobile platforms and emergent data capture techniques for game-based learning are discussed, as are methods for putting engaging gameplay at the forefront of the experience whilst relying on rich data capture and analysis to provide an effective learning solution

High glucose inhibits human epidermal keratinocyte proliferation for cellular studies on diabetes mellitus

In order to more clarify the delayed wound healing in diabetes mellitus, we cultured the human epidermal keratinocytes in both 6 mM (control group) and 12 mM glucose (high-glucose group) of ‘complete’ MCDB 153 medium. Hyperglycaemia slowed the rate of their proliferation and inhibited their DNA synthesis and the production of total proteins. By 1 month after primary seeding in high-glucose group, the cells ceased their proliferation, whereas the cells in control group grew for more than 40 days. Mean population doublings in high-glucose group was 5·27 (vs. 7·25 in control, P = 0·001), and mean population doubling time during 1 month in high glucose group was 5·43 days (vs. 3·65 days in control, P = 0·02). They indicate that prolonged exposure to high glucose decreases the replicative life span of human epidermal keratinocytes in vitro. Furthermore, analysis of fatty acid contents in membrane phospholipids with thin-layer and gas chromatography showed no difference between the cultured keratinocytes in both conditions. Immunocytochemical staining of glucose transporter 1 shows that 28·1% of cells in high-glucose group were almost twice positive of those in control group (13·2%, P = 0·008). The mechanism of the ill effects of high glucose on epidermal keratinocytes is not so far clear, but it indicates the possibility of any direct effect of hyperglycaemia on glucose metabolism without changing lipid metabolism on cell membrane. The high-glucose group presented in this report can be available as an in vitro valuable study model of skin epidermal condition on diabetes mellitus.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72307/1/j.1742-4801.2005.00148.x.pd

Implementation and evaluation of the Level Set method: towards efficient and accurate simulation of wet etching for microengineering applications

The use of atomistic methods, such as the Continuous Cellular Automaton (CCA), is currently regarded as a computationally efficient and experimentally accurate approach for the simulation of anisotropic etching of various substrates in the manufacture of Micro-electro-mechanical Systems (MEMS). However, when the features of the chemical process are modified, a time-consuming calibration process needs to be used to transform the new macroscopic etch rates into a corresponding set of atomistic rates. Furthermore, changing the substrate requires a labor-intensive effort to reclassify most atomistic neighborhoods. In this context, the Level Set (LS) method provides an alternative approach where the macroscopic forces affecting the front evolution are directly applied at the discrete level, thus avoiding the need for reclassification and/or calibration. Correspondingly, we present a fully-operational Sparse Field Method (SFM) implementation of the LS approach, discussing in detail the algorithm and providing a thorough characterization of the computational cost and simulation accuracy, including a comparison to the performance by the most recent CCA model. We conclude that the SFM implementation achieves similar accuracy as the CCA method with less fluctuations in the etch front and requiring roughly 4 times less memory. Although SFM can be up to 2 times slower than CCA for the simulation of anisotropic etchants, it can also be up to 10 times faster than CCA for isotropic etchants. In addition, we present a parallel, GPU-based implementation (gSFM) and compare it to an optimized, multicore CPU version (cSFM), demonstrating that the SFM algorithm can be successfully parallelized and the simulation times consequently reduced, while keeping the accuracy of the simulations. Although modern multicore CPUs provide an acceptable option, the massively parallel architecture of modern GPUs is more suitable, as reflected by computational times for gSFM up to 7.4 times faster than for cSFM. (c) 2013 Elsevier B.V. All rights reserved.We thank the anonymous reviewers for their valuable comments and suggestions. This work has been supported by the Spanish FPI-MICINN BES-2011-045940 grant and the Ramon y Cajal Fellowship Program by the Spanish Ministry of Science and Innovation. Also, we acknowledge support by the JAE-Doc grant from the Junta para la Ampliacion de Estudios program co-funded by FSE and the Professor Partnership Program by NVIDIA Corporation.Montoliu Álvaro, C.; Ferrando Jódar, N.; Gosalvez, MÁ.; Cerdá Boluda, J.; Colom Palero, RJ. (2013). Implementation and evaluation of the Level Set method: towards efficient and accurate simulation of wet etching for microengineering applications. Computer Physics Communications. 184(10):2299-2309. https://doi.org/10.1016/j.cpc.2013.05.016S229923091841

Citizen science versus professional data collection: Comparison of approaches to mosquito monitoring in Germany

Due to the recent emergence of invasive mosquito species and the outbreaks of mosquito-borne diseases in Europe, research on the ecology and diversity of the mosquito fauna has returned to scientific agendas. Through a nationwide surveillance programme in Germany, mosquitoes have been monitored actively by systematically operated traps since 2011, and passively by the 'Mückenatlas' (mosquito atlas) citizen science project launched in 2012. To assess the performance of both monitoring methods we compared the two respective datasets with regard to habitat coverage, species composition and the ability to detect invasive mosquitoes. The datasets include observations from the beginning of the project until the end of 2017. We found significant differences in species composition caused by land use types and the participants' recording activity. Active monitoring performed better in mapping mosquito diversity, whereas passive monitoring better detected invasive species, thereby using data from private premises scientists usually cannot access. Synthesis and applications. Active and passive monitoring is complementary. Combining them allows for the determination of mosquito diversity, efficient detection of emerging invasive species and the initiation of rapid-response actions against such invaders. The 'Mückenatlas' sets an example for the usefulness of citizen science when included in a national monitoring programme, an approach that may be worth copying for tackling the global spread of arthropod vectors of disease agents

Developmental changes in human dopamine neurotransmission: cortical receptors and terminators

<p>Abstract</p> <p>Background</p> <p>Dopamine is integral to cognition, learning and memory, and dysfunctions of the frontal cortical dopamine system have been implicated in several developmental neuropsychiatric disorders. The dorsolateral prefrontal cortex (DLPFC) is critical for working memory which does not fully mature until the third decade of life. Few studies have reported on the normal development of the dopamine system in human DLPFC during postnatal life. We assessed pre- and postsynaptic components of the dopamine system including tyrosine hydroxylase, the dopamine receptors (D1, D2 short and D2 long isoforms, D4, D5), catechol-<it>O</it>-methyltransferase, and monoamine oxidase (A and B) in the developing human DLPFC (6 weeks -50 years).</p> <p>Results</p> <p>Gene expression was first analysed by microarray and then by quantitative real-time PCR. Protein expression was analysed by western blot. Protein levels for tyrosine hydroxylase peaked during the first year of life (p < 0.001) then gradually declined to adulthood. Similarly, mRNA levels of dopamine receptors D2S (p < 0.001) and D2L (p = 0.003) isoforms, monoamine oxidase A (p < 0.001) and catechol-<it>O</it>-methyltransferase (p = 0.024) were significantly higher in neonates and infants as was catechol-<it>O</it>-methyltransferase protein (32 kDa, p = 0.027). In contrast, dopamine D1 receptor mRNA correlated positively with age (p = 0.002) and dopamine D1 receptor protein expression increased throughout development (p < 0.001) with adults having the highest D1 protein levels (p ≤ 0.01). Monoamine oxidase B mRNA and protein (p < 0.001) levels also increased significantly throughout development. Interestingly, dopamine D5 receptor mRNA levels negatively correlated with age (r = -0.31, p = 0.018) in an expression profile opposite to that of the dopamine D1 receptor.</p> <p>Conclusions</p> <p>We find distinct developmental changes in key components of the dopamine system in DLPFC over postnatal life. Those genes that are highly expressed during the first year of postnatal life may influence and orchestrate the early development of cortical neural circuitry while genes portraying a pattern of increasing expression with age may indicate a role in DLPFC maturation and attainment of adult levels of cognitive function.</p