bluuebunny/crossref_metadata_2025_split huggingface dataset:
Targeted mutagenesis of the Arabidopsis GROWTH-REGULATING FACTOR (GRF) gene family suggests competition of multiplexed sgRNAs for Cas9 apoprotein
Authors: Angulo Juan , Astin Christopher P , Bauer Olivia , Blash Kelan J , Bowen Natalee M , Chukwudinma Nneoma J , Dinofrio Austin S , Faletti Donald O , Ghulam Alexa M , Gusinde-Duffy Cloe M , Horace Kamaria J , Ingram Andrew M , Isaack Kylie E , Jeong Geon , Kiser Randolph J , Kobylanski Jason S , Long Madeline R , Manning Grace A , Morales Julie M , Nguyen Kevin H , Pham Robin T , Phillips Monthip H , Reel Tanner W , Seo Jenny E , Vo Hiep D , Wukuson Alexander M , Yeary Kathryn A , Zheng Grace Y , Lukowitz Wolfgang 
Published: August 2020

AbstractGenome editing in plants typically relies on T-DNA plasmids that are mobilized by Agrobacterium-mediated transformation to deliver the CRISPR/Cas9 machinery. Here, we introduce a series of CRISPR/Cas9 T-DNA vectors for minimal lab settings, such as in the classroom or citizen science projects. Spacer sequences targeting genes of interest can be inserted as annealed short oligonucleotides in a single straightforward cloning step. Fluorescent markers expressed in mature seeds enable reliable selection of transgenic as well as transgene-free individuals using a combination of inexpensive LED lamps and colored-glass alternative filters. Testing these tools on the Arabidopsis GROWTH-REGULATING FACTOR (GRF) gene family, we found that Cas9 expression from an EGG CELL1 (EC1) promoter resulted in tenfold lower mutation rates than expression from a UBIQUITIN10 (UBQ10) promoter. A collection of bona fide null mutations in all nine GRF genes could be established with little effort. Finally, we explored the effects of simultaneously targeting two, four and eight GRF genes on the rate of induced mutations at each target locus. Multiplexing caused strong interference effects: while mutation rates at some loci remained consistently high, mutation rates at other loci dropped dramatically with increasing number of single guide RNA species. Our results suggest potential detrimental genetic interaction between induced mutations as well as competition of CRISPR RNAs for a limiting amount of Cas9 apoprotein.
Sleep-dependent offline performance gain in visual perceptual learning is consistent with a learning-dependent model
Authors: Tamaki Masako , Sasaki Yuka 
Published: August 2020

SummaryAre the sleep-dependent offline performance gains of visual perceptual learning (VPL) consistent with a use-dependent or learning-dependent model? Here, we found that a use-dependent model is inconsistent with the offline performance gains in VPL. In two training conditions with matched visual usages, one generated VPL (learning condition), while the other did not (interference condition). The use-dependent model predicts that slow-wave activity (SWA) during posttraining NREM sleep in the trained region increases in both conditions, in correlation with offline performance gains. However, compared with those in the interference condition, sigma activity, not SWA, during NREM sleep and theta activity during REM sleep, source-localized to the trained early visual areas, increased in the learning condition. Sigma activity correlated with offline performance gain. These significant differences in spontaneous activity between the conditions suggest that there is a learning-dependent process during posttraining sleep for the offline performance gains in VPL.
Alternative catalytic mechanisms driven by structural plasticity is an emerging theme in HAS-GTPases, Era and FeoB
Authors: Batra Sahil , Kumar Ashok , Prakash Balaji 
Published: August 2020

AbstractGTP hydrolysis is the underlying basis for functioning of ‘biological switches’ or GTPases. Extensively studied GTPases, Ras and EF-Tu, use a conserved Gln/His that facilitates the activation of attacking water for nucleophilic attack. However, this is insufficient to explain catalysis inHydrophobicAmino acidSubstituted (HAS)-GTPases that naturally possess a hydrophobic residue in lieu of Gln/His. We had previously reported a bridging water-chain mediated catalytic mechanism for HAS-GTPase FeoB; which utilizes two distantly-located but conserved glutamates. Curiously, mutating these does not abolish GTP hydrolysis. Similarly, in this study we report our observations on another HAS-GTPase Era, wherein the mutants of catalytically important residues continue to hydrolyze GTP. We attempt to rationalize these inquisitive observations on GTP hydrolysis by FeoB and Era mutants. We propose a general theory that appears common to at least three classes of GTPases, where ‘alternative mechanisms’ emerge when the primary mechanism is disrupted. Based on the analysis of crystal structures of FeoB and Era mutants, bound to the transition state analogue GDP.AlFx, this work suggests that in the absence of catalytically important residues, the active site waters in both FeoB and Era undergo re-arrangements, which in turn helps in sustaining GTP hydrolysis. Similar employment of alternative mechanisms was also suggested for the catalytic mutants of hGBP1. Importantly, such alternatives underscore the robustness of GTP hydrolysis mechanisms in these systems, and raise important questions regarding the need for persistent GTP hydrolysis and the physiological relevance of structural plasticity seen here.
Flagella Ca2+elevations regulate pausing of retrograde intraflagellar transport trains in adherentChlamydomonasflagella
Authors: Fort Cecile , Collingridge Peter , Brownlee Colin , Wheeler Glen 
Published: August 2020

AbstractThe movement of ciliary membrane proteins is directed by transient interactions with intraflagellar transport (IFT) trains. The green algaChlamydomonashas adapted this process for gliding motility, using IFT to move adhesive glycoproteins (FMG-1B) in the flagella membrane. Although Ca2+signalling contributes directly to the gliding process, uncertainty remains over the mechanisms through which Ca2+acts to influence the movement of IFT trains. Here we show that flagella Ca2+elevations regulate IFT primarily by initiating the movement of paused retrograde IFT trains. Flagella Ca2+elevations exhibit complex spatial and temporal properties, including high frequency repetitive Ca2+elevations that prevent the accumulation of paused retrograde IFT trains. We show that flagella Ca2+elevations disrupt the IFT-dependent movement of microspheres along the flagella membrane. The results suggest that flagella Ca2+elevations directly disrupt the interaction between retrograde IFT particles and flagella membrane glycoproteins to modulate gliding motility and the adhesion of the flagellum to a surface.
Binocular disparity-based learning does not generalize to the other visual hemifield
Authors: Klinzing Jens G. , Nienborg Hendrikje , Rauss Karsten 
Published: August 2020

AbstractVisual perceptual learning refers to long-lasting performance improvements on a visual skill, an ability presumably supported by plastic changes in early visual brain areas. Visual perceptual learning has been shown to be induced by training and to benefit from consolidation during sleep, presumably via the reactivation of learning-associated neuronal firing patterns. However, previous studies have almost exclusively relied on a single paradigm, the texture discrimination task, on which performance improvements may not rely on strictly perceptual skills. Here, we tested whether sleep has beneficial effects on a visual disparity discrimination task. We confirm previous findings in showing that the ability to discriminate different disparities is unaffected by sleep during a 12-hour retention period after training. Importantly, we extend these results by providing evidence against an effect of sleep on the generalization of improved disparity discrimination across the vertical meridian. We further rule out carry-over effects as a possible confound in our earlier within-subject study by relying on a between-subject design. The combined data from both studies argue against sleep as an important factor in the consolidation of a strictly perceptual skill. This sets important constraints on models of the role of sleep and sleep-associated neural reactivation in the consolidation of non-declarative memories.
Complexome profile ofToxoplasma gondiimitochondria identifies a divergent cytochromebc1complex
Authors: Maclean Andrew E. , Bridges Hannah R. , Silva Mariana F. , Ding Shujing , Hirst Judy , Sheiner Lilach 
Published: August 2020

AbstractThe mitochondrial electron transport chain (mETC) and F1Fo-ATP synthase are of central importance for energy and metabolism in eukaryotic cells. The Apicomplexa, important pathogens of humans causing diseases such as toxoplasmosis and malaria, depend on their mETC in every known stage of their complicated life cycles. Here, using a complexome profiling proteomic approach, we have characterised theToxoplasmamETC complexes and F1Fo-ATP synthase. We identified and assigned 60 proteins to complexes II, IV and F1Fo-ATP synthase ofToxoplasma, of which 16 have not been identified previously. Notably, our complexome profile elucidates the composition of theToxoplasmacomplex III, the target of clinically used drugs such as atovaquone. We identified two new homologous subunits and two new parasite-specific subunits, one of which is broadly conserved in myzozoans. We demonstrate all four proteins are essential for complex III stability and parasite growth, and show their depletion leads to decreased mitochondrial potential, supporting their role as complex III subunits. Our study highlights the divergent subunit composition of the apicomplexan mETC complexes and sets the stage for future structural and drug discovery studies.Author SummaryApicomplexan parasites, such asToxoplasmaandPlasmodium, cause diseases of global importance, such as toxoplasmosis and malaria. The mitochondrial electron transport chain (mETC) and F1Fo-ATP synthase, which provide the parasite with energy and important metabolites, are essential for parasite function. Here, using a proteomic technique called complexome profiling, we report the composition of theToxoplasmamETC and F1Fo-ATP synthase. In particular, we reveal the compositions of complexes II and III for the first time. Complex III is an important drug target, yet its full protein composition was unknown. We identify new parasite-specific complex III subunits and demonstrate that they are essential for parasite survival and for proper functioning of the mETC. Our study highlights the divergent nature of the apicomplexan mETC and F1Fo-ATP synthase.
The conserved ER-transmembrane protein TMEM39 coordinates with COPII to promote collagen secretion and prevent ER stress
Authors: Zhang Zhe , Luo Shuo , Barbosa Guilherme Oliveira , Bai Meirong , Kornberg Thomas B. , Ma Dengke K. 
Published: August 2020

AbstractDysregulation of collagen production and secretion contributes to aging and tissue fibrosis of major organs. How premature collagen proteins in the endoplasmic reticulum (ER) route as specialized cargos for secretion remains to be fully elucidated. Here, we report that TMEM39, an ER-localized transmembrane protein, regulates production and secretory cargo trafficking of procollagen. We identify the C. elegans ortholog TMEM-39 from an unbiased RNAi screen and show that deficiency of tmem-39 leads to striking defects in cuticle collagen production and constitutively high ER stress response. RNAi knockdown of the tmem-39 ortholog in Drosophila causes similar defects in collagen secretion from fat body cells. The cytosolic domain of human TMEM39A binds to Sec23A, a vesicle coat protein that drives collagen secretion and vesicular trafficking. TMEM-39 regulation of collagen secretion is independent of ER stress response and autophagy. We propose that roles of TMEM-39 in collagen secretion and preventing ER stress are likely evolutionarily conserved.
Human ES and iPS Cells Display Less Drug Resistance Than Differentiated Cells, and Naïve-State Induction Further Decreases Drug Resistance
Authors: Panina Yulia , Yamane Junko , Kobayashi Kenta , Sone Hideko , Fujibuchi Wataru 
Published: August 2020

AbstractPluripotent stem cells (PSCs) possess unique characteristics that distinguish them from other cell types. Human embryonic stem (ES) cells are recently gaining attention as a powerful tool for human toxicity assessment without the use of experimental animals, and an embryonic stem cell test (EST) was introduced for this purpose. However, human PSCs have not been thoroughly investigated in terms of drug resistance or compared with other cell types or cell states, such as naïve state, to date. Aiming to close this gap in research knowledge, we assessed and compared several human PSC lines for their resistance to drug exposure. Firstly, we report that RIKEN-2A human induced pluripotent stem (iPS) cells possessed approximately the same sensitivity to selected drugs as KhES-3 human ES cells. Secondly, both ES and iPS cells were several times less resistant to drug exposure than other non-pluripotent cell types. Finally, we showed that iPS cells subjected to naïve-state induction procedures exhibited a sharp increase in drug sensitivity. Upon passage of these naïve-like cells in non-naïve PSC culture medium, their sensitivity to drug exposure decreased. We thus revealed differences in sensitivity to drug exposure among different types or states of PSCs and, importantly, indicated that naïve-state induction could increase this sensitivity.
Inhibition of mTOR decreases insoluble protein burden by reducing translation inC. elegans
Authors: Yee Zhuangli , Lim Shaun Hsien Yang , Ng Li Fang , Gruber Jan 
Published: August 2020

AbstractAging animals accumulate insoluble protein as a consequence of a decline of proteostatic maintenance with age. InCaenorhabditis elegans, for instance, levels of detergent-insoluble protein increase with age. In longer-lived strains ofC. elegans, this accumulation occurs more slowly, implying a link to lifespan determination. We further explored this link, and found that detergent-insoluble protein accumulates more rapidly at higher temperatures, a condition where lifespan is short. We employed aC. elegansstrain carrying a GFP transcriptional reporter under the control of a heat shock (hsp-16.2) promoter to investigate the dynamics of proteostatic failure in individual nematodes. We found that early, sporadic activation ofhsp-16.2was predictive of shorter remaining lifespan in individual nematodes. Exposure to rapamycin, resulting in reduced mTOR signaling, delayed spurious expression, extended lifespan, and delayed accumulation of insoluble protein, suggesting that targets downstream of the mTOR pathway regulate the accumulation of insoluble protein. We specifically explored ribosomal S6 kinase (rsks-1) as one such candidate and found that RNAi againstrsks-1also resulted in less age-dependent accumulation of insoluble protein and extended lifespan. Our results demonstrate that inhibition of protein translationviareduced mTOR signaling resulted in slower accumulation of insoluble protein, delayed proteostatic crisis and extended lifespan inC. elegans.
Estimating plant abundance in irregular polygons: plot-based sampling with a conditional spatial configuration
Authors: Damgaard Christian 
Published: August 2020

AbstractWhen estimating plant species abundance using plot-based methods in irregular polygons, there are some constraints to the spatial configuration of the sampling plots: i) the sample plots must be positioned within the polygon, ii) the sample plots may not overlap, iii) due to the spatial variation in abundance often found in natural plant communities, the spatial coverage must be relatively large in order to decrease the sampling variance, iv) due to a possible mosaic structure of the vegetation, systematic designs should be avoided due to the possibility of biased estimates. Here, we suggest a protocol using simulations of the possible random spatial configurations to select a conditional spatial configuration that meets these constraints.
Adaptive value of circadian rhythms in High Arctic Svalbard ptarmigan
Authors: Appenroth Daniel , Wagner Gabriela C. , Hazlerigg David G. , West Alexander C. 
Published: August 2020

SUMMARYThe arctic archipelago of Svalbard (74 to 81° North) experiences extended periods of uninterrupted daylight in summer and uninterrupted darkness in winter. Species native to Svalbard display no daily rhythms in behaviour or physiology during these seasons, leading to the view that circadian rhythms may be redundant in arctic environments [1, 2]. Nevertheless, seasonal changes in the physiology and behaviour of arctic species rely on photoperiodic synchronisation to the solar year. Since this phenomenon is generally circadian-based in temperate species, we investigated if this might be a preserved aspect of arctic temporal organisation.Here, we demonstrate the involvement of the circadian clock in the seasonal photoperiodic response of the Svalbard ptarmigan (Lagopus muta hyperborea), the world’s northernmost resident bird species. First, we show the persistence of rhythmic clock gene expression under constant conditions within the mediobasal hypothalamus and pars tuberalis, the key tissues in the seasonal neuroendocrine cascade. We then employ a “sliding skeleton photoperiod” protocol, revealing that the driving force behind seasonal biology of the Svalbard ptarmigan is rhythmic sensitivity to light, a feature that depends on a functioning circadian rhythm. Our results suggest that the unusual selective pressure of the Arctic relaxes the adaptive value of the circadian clock for organisation of daily activity patterns, whilst preserving its importance for seasonal synchronisation. Thus, our data simultaneously reconnects circadian rhythms to life in the Arctic and establishes a universal principle of evolutionary value for circadian rhythms in seasonal biology.
Dysfunctional perceptual antecedent can justify the social orienting deficit in autism spectrum disorder: an eye-tracking study
Authors: Carrozza Cristina , Fabio Rosa Angela 
Published: August 2020

Purpose
Children with Autism Spectrum Disorder (ASD) show reduced attention to social stimuli. The reasons for these impairments are still being debated by researchers. The aim of this study is to analyse if reduced attention towards social stimuli is determined by initial underlying difficulties in the control of visual attention. Among the variables that could produce these difficulties, the authors considered geometric complexity and typology of geometric figures.

Design/methodology/approach
To test this hypothesis, in this paper, an eye-tracker paradigm was used for assessing visual exploration and recognition memory towards geometric figures (curved vs rectilinear) with two levels of geometric complexity (low and high) in 17 children with ASD matched with 17 children with typical development (TD).

Findings
The results showed that the ASD group seemed indifferent to both the geometric complexity and the typology of figures (curved and rectilinear), whereas the TD group showed higher performances with highly complex and curved geometric figures than with low complex and rectilinear geometric figures.

Research limitations/implications
Because of the chosen research approach, the research results may lack generalizability. Therefore, researchers are encouraged to test the proposed hypotheses further.

Practical implications
This paper includes implications upon the presence of an unspecified visual attention deficit that is present from the early stages of the processing of stimuli.

Social implications
The understanding of this deficit from the early stages of the processing of stimuli can help educators to intervene at an early stage when disturbances in social relationships are starting.

Originality/value
This study contributes to understanding the presence of dysfunctional perceptual antecedents that could determine general difficulties in paying attention to social stimuli in ASD subjects.
Comparative analysis of employment trend in African agriculture relative to other regions: a gender perspective
Authors: Mkpado Mmaduabuchukwu , Mkpado Ndidiamaka Sandra 
Published: August 2020

PurposeLucrative employment in agriculture is fundamental to poverty alleviation in Africa. The paper examined employment along gender, impact of materials and proportion of female employment in African agriculture.Design/methodology/approachTime series econometrics was employed in the framework of production function analysis involving 36 years of data.FindingsResults show that world labour in agriculture decreased from 49.77 to 40.04% but increased from 12.43 to 16.94% in Africa. World female employment in agriculture ranged from 40.56 to 42.81% and from 40.40 to 43.02% in developing economies, but decreased from 40.39 to 36.08% in developed economies. Total agricultural labour in Africa was negatively and significantly related to agricultural gross production index number (APIN).Research limitations/implicationsInteraction of cattle stock and females employed in agriculture was positive and significant at pooled African values. Interaction of irrigation facilities and female labour was positive and significant in West Africa. Interaction of cattle stock and total labour in Southern Africa had negative relationship with APIN. Interaction of total labour and irrigation had negative relationship with APIN in Africa. Insufficient agricultural facilities in terms of cattle stock and irrigation infrastructure for the populace exist. It recommends increased investments to expand irrigated lands and livestock.Practical implicationsAfrican governments need to use good political will to effect the needed transformation in agriculture. It is possible for agriculture to offer lucrative employment to both males and females in less developed world as in developed economies.Originality/valueThe paper noted very limited agricultural facilities in terms of cattle stock and irrigation facilities for the populace engaged in agriculture. It recommends investments to expand irrigated lands and livestock.