bluuebunny/crossref_metadata_2025_split huggingface dataset:
Short distance non-autonomy and intercellular transfer of chitin synthase in Drosophila
Authors: Adler Paul N. 
Published: May 2020

AbstractThe complex structure of insect exoskeleton has inspired material scientists and engineers. Chitin is a major component of the cuticle and it is synthesized by the enzyme chitin synthase. There is a single chitin synthase gene (kkv) in Drosophila facilitating research on the function of chitin. Previous editing ofkkvlead to the recovery of a viable hypomorphic allele. Experiments described in this paper argue that a reduction in chitin synthase leads to the shafts of sensory bristles becoming fragile and frequently breaking off as the animals age. This is likely due to reduced chitin levels and further suggests that chitin plays a role in resilience of insect cuticle. The different layers in cuticle are continuous across the many epidermal cells that secrete the cuticle that covers the body. Little is known about the mechanisms responsible for this continuity. Using genetic mosaics and scanning electron microscopy this paper establishes thatkkvshows short range cell non-autonomy. It also provides evidence for 2 possible mechanisms. One is the intercellular transfer of Kkv protein from one cell to its neighbors and the second is the deposition of cuticular material across the boundaries of neighboring cells.
Tolerance induction in memory CD4 T cells is partial and reversible
Authors: Gray Joshua I , Al-Khabouri Shaima , Morton Fraser , Clambey Eric T , Gapin Laurent , Matsuda Jennifer L , Kappler John W , Marrack Philippa , Garside Paul , Otto Thomas D , MacLeod Megan KL 
Published: May 2020

AbstractMemory T cells respond rapidly in part because they are less reliant on heightened levels of costimulatory molecules. This presents challenges to silencing memory T cells in tolerance strategies for autoimmunity or allergy. We find that memory CD4 T cells generated by infection or immunisation survive secondary activation with antigen delivered without adjuvant, regardless of their location in secondary lymphoid organs or peripheral tissues. These cells were, however, functionally altered following a tertiary immunisation with antigen and adjuvant, proliferating poorly but maintaining their ability to produce inflammatory cytokines. Transcriptional and cell cycle analysis of these memory CD4 T cells suggest they are unable to commit fully to cell division potentially because of low expression of DNA repair enzymes. In contrast, these memory CD4 T cells could proliferate following tertiary reactivation by viral re-infection. These data suggest that tolerance induction in memory CD4 T cells is partial and can be reversed.
A genetically encoded red fluorescence dopamine biosensor enables dual imaging of dopamine and norepinephrine
Authors: Nakamoto Chihiro , Goto Yuhei , Tomizawa Yoko , Fukata Yuko , Fukata Masaki , Harpsøe Kasper , Gloriam David E. , Aoki Kazuhiro , Takeuchi Tomonori 
Published: May 2020

AbstractDopamine (DA) and norepinephrine (NE) are pivotal neuromodulators that regulate a broad range of brain functions, often in concert. Despite their physiological importance, untangling the relationship between DA and NE in finely controlling output functions is currently challenging, primarily due to a lack of techniques to visualize spatiotemporal dynamics with sufficiently high selectivity. Although genetically encoded fluorescent biosensors have been developed to detect DA, their poor selectivity prevents distinguishing DA from NE. Here, we report the development of a red fluorescent genetically encoded GPCR (G protein-coupled receptor)-activation reporter for DA termed ‘R-GenGAR-DA’. More specifically, a circular permutated red fluorescent protein (cpmApple) was inserted into the third intracellular loop of human DA receptor D1 (DRD1) followed by the screening of mutants within the linkers between DRD1 and cpmApple. We developed two variants: R-GenGAR-DA1.1, which brightened following DA stimulation, and R-GenGAR-DA1.2, which dimmed. R-GenGAR-DA1.2 demonstrated reasonable dynamic range (ΔF/F0= –50%) and DA affinity (EC50= 0.7 µM) as well as the highest selectivity for DA over NE (143-fold) amongst available DA biosensors. Due to its high selectivity, R-GenGAR-DA1.2 allowed dual-color fluorescence live imaging for monitoring DA and NE, combined with the existing green-NE biosensor GRABNE1m, which has high selectivity for NE over DA (>350-fold) in HeLa cells and hippocampal neurons grown from primary culture. By enabling precise measurement of DA, as well as simultaneous visualization of DA and NE, the red-DA biosensor R-GenGAR-DA1.2 is promising in advancing our understanding of the interplay between DA and NE in organizing key brain functions.Significance StatementThe neuromodulators dopamine and norepinephrine modulate a broad range of brain functions, often in concert. One current challenge is to measure dopamine and norepinephrine dynamics simultaneously with high spatial and temporal resolution. We therefore developed a red-dopamine biosensor that has 143-fold higher selectivity for dopamine over norepinephrine. Taking advantage of its high selectivity for dopamine over norepinephrine, this red-dopamine biosensor allowed dual-color fluorescence live imaging for monitoring dopamine and norepinephrine in both HeLa cells and hippocampal neuronsin vitrocombined with the existing green-norepinephrine biosensor that has 350-fold selectivity for norepinephrine over dopamine. Thus, this approach can provide new opportunities to advance our understanding of high spatial and temporal dynamics of dopamine and norepinephrine in normal and abnormal brain functions.
Human H-ferritin presenting RBM of spike glycoprotein as potential vaccine of SARS-CoV-2
Authors: Yao Dehui , Lao Fang , Zhang Zeyi , Liu Yan , Cheng Jianwei , Ding Fengjiao , Wang Xiaofei , Xi Lun , Wang Chuang , Yan Xichong , Zhang Rongkun , Ouyang Fangxing , Ding Hui , Ke Tianyi 
Published: May 2020

AbstractThe outbreak of COVID-19 has so far inflicted millions of people all around the world and will have a long lasting effect on every aspect of everyone’s life. Yet there is no effective approved treatment for the disease. In an effort of utilizing human ferritin as nanoplatform for drug delivery, we engineered a fusion protein by presenting receptor-binding motif (RBM) of SARS-CoV-2 virus spike glycoprotein on the N-terminus of ferritin subunits. The designed fusion protein with a cage-like structure, similar to that of corona virus, is a potential anti-SARS-CoV-2 vaccine. We hereby show the construction, preparation, and characterization of the fusion protein RBM-HFtn. Our initial affinity study confirmed its biological activity towards ACE2 receptor which suggests its mode of action against SARS-CoV-2 could be either through vaccine therapy or blocking the cellular entry of virus as antagonist of ACE2 receptor.
Cell states beyond transcriptomics: integrating structural organization and gene expression in hiPSC-derived cardiomyocytes
Authors: Gerbin Kaytlyn A. , Grancharova Tanya , Donovan-Maiye Rory , Hendershott Melissa C. , Brown Jackson , Dinh Stephanie Q. , Gehring Jamie L. , Hirano Matthew , Johnson Gregory R. , Nath Aditya , Nelson Angelique , Roco Charles M. , Rosenberg Alexander B. , Filip Sluzewski M. , Viana Matheus P. , Yan Calysta , Zaunbrecher Rebecca J. , Cordes Metzler Kimberly R. , Menon Vilas , Palecek Sean P. , Seelig Georg , Gaudreault Nathalie , Knijnenburg Theo , Rafelski Susanne M. , Theriot Julie A. , Gunawardane Ruwanthi N. 
Published: May 2020

SummaryWe present a quantitative co-analysis of RNA abundance and sarcomere organization in single cells and an integrated framework to predict subcellular organization states from gene expression. We used human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes expressing mEGFP-tagged alpha-actinin-2 to develop quantitative image analysis tools for systematic and automated classification of subcellular organization. This captured a wide range of sarcomeric organization states within cell populations that were previously difficult to quantify. We performed RNA FISH targeting genes identified by single cell RNA sequencing to simultaneously assess the relationship between transcript abundance and structural states in single cells. Co-analysis of gene expression and sarcomeric patterns in the same cells revealed biologically meaningful correlations that could be used to predict organizational states. This study establishes a framework for multi-dimensional analysis of single cells to study the relationships between gene expression and subcellular organization and to develop a more nuanced description of cell states.Graphical AbstractTranscriptional profiling and structural classification was performed on human induced pluripotent stem cell-derived cardiomyocytes to characterize the relationship between transcript abundance and subcellular organization.
Sirtuin-1 Sensitive Lysine-136 Acetylation Drives Phase Separation and Pathological Aggregation of TDP-43
Authors: Morato Jorge Garcia , Hans Friederike , von Zweydorf Felix , Feederle Regina , Elsässer Simon J. , Skodras Angelos A. , Gloeckner Christian Johannes , Buratti Emanuele , Neumann Manuela , Kahle Philipp J. 
Published: May 2020

AbstractThe trans-activation response DNA-binding protein TDP-43 regulates RNA processing and forms neuropathological aggregates in patients with amyotrophic lateral sclerosis and frontotemporal lobar degeneration. Investigating TDP-43 post-translational modifications, we discovered that K84 acetylation reduced nuclear import whereas K136 acetylation impaired RNA binding and splicing capabilities of TDP-43. Such failure of RNA interaction triggered TDP-43 phase separation mediated by the C-terminal low complexity domain, leading to the formation of insoluble aggregates with pathologically phosphorylated and ubiquitinated TDP-43. Confirming the results from site-directed mutagenesis, we succeeded to introduce authentic acetyl-lysine at the identified sites via amber suppression. [AcK84]TDP-43 showed cytoplasmic mislocalization and the aggregation propensity of [acK136]TDP-43 was confirmed. With newly developed antibodies, we found that the nuclear sirtuin SIRT1 can potently deacetylate [acK136]TDP-43. Moreover, SIRT1 reduced the aggregation propensity of [acK136]TDP-43. Thus, distinct lysine acetylations modulate nuclear import, RNA binding and phase separation of TDP-43, suggesting novel regulatory mechanisms for TDP-43 pathogenesis.
A model of the PI cycle reveals the regulating roles of lipid-binding proteins and pitfalls of using mosaic biological data
Authors: Mazet Francoise , Tindall Marcus J. , Gibbins Jonathan M. , Fry Michael J. 
Published: May 2020

AbstractThe phosphatidylinositol (PI) cycle is central to eukaryotic cell signaling. Its complexity, due to the number of reactions and lipid and inositol phosphate intermediates involved makes it difficult to analyze experimentally. Computational modelling approaches are seen as a way forward to elucidate complex biological regulatory mechanisms when this cannot be achieved solely through experimental approaches. Whilst mathematical modelling is well established in informing biological systems, many models are often informed by data sourced from different cell types (mosaic data), to inform model parameters. For instance, kinetic rate constants are often determined from purified enzyme datain vitroor use experimental concentrations obtained from multiple unrelated cell types. Thus they do not represent any specific cell type nor fully capture cell specific behaviours. In this work, we develop a model of the PI cycle informed byin-vivoomics data taken from a single cell type, namely platelets. Our model recapitulates the known experimental dynamics before and after stimulation with different agonists and demonstrates the importance of lipid- and protein-binding proteins in regulating second messenger outputs. Furthermore, we were able to make a number of predictions regarding the regulation of PI cycle enzymes and the importance of the number of receptors required for successful GPCR signaling. We then consider how pathway behavior differs, when fully informed by data for HeLa cells and show that model predictions remain relatively consistent. However, when informed by mosaic experimental data model predictions greatly vary. Our work illustrates the risks of using mosaic datasets from unrelated cell types which leads to over 75% of outputs not fitting with expected behaviors.Authors summaryComputational models of cellular complexity offer much in terms of understanding cell behaviors and in informing experimental design, but their usefulness is limited in them being built with mosaic data not representing specific cell types and tested against limited experimental outputs. In this work we demonstrate an approach using quantitative proteomic datasets and temporal experimental data from a single cell type (platelets) to inform kinetic rate constants and protein concentrations for a mathematical model of a key signaling pathway - the phosphatidylinositol (PI) cycle; known for its central role in numerous cell functions and diseases. After using our model to make novel predictions regarding how aspects of the pathway are regulated, we demonstrate its versatile nature by utilising proteomic data from other cell types to generate similar predictions for those cells while highlighting the pitfalls of using mosaic data when constructing computational models.
Structure, function and variants analysis of the androgen-regulatedTMPRSS2, a drug target candidate for COVID-19 infection
Authors: David Alessia , Khanna Tarun , Beykou Melina , Hanna Gordon , Sternberg Michael J E 
Published: May 2020

AbstractSARS-CoV-2 is a novel virus causing mainly respiratory, but also gastrointestinal symptoms. Elucidating the molecular processes underlying SARS-CoV-2 infection, and how the genetic background of an individual is responsible for the variability in clinical presentation and severity of COVID-19 is essential in understanding this disease.Cell infection by the SARS-CoV-2 virus requires binding of its Spike (S) protein to the ACE2 cell surface protein and priming of the S by the serine protease TMPRSS2. One may expect that genetic variants leading to a defective TMPRSS2 protein can affect SARS-CoV-2 ability to infect cells. We used a range of bioinformatics methods to estimate the prevalence and pathogenicity of TMPRSS2 genetic variants in the human population, and assess whether TMPRSS2 and ACE2 are co-expressed in the intestine, similarly to what is observed in lungs.We generated a 3D structural model of the TMPRSS2 extracellular domain using the prediction server Phyre and studied 378 naturally-occurring TMPRSS2 variants reported in the GnomAD database. One common variant, p.V160M (rs12329760), is predicted damaging by both SIFT and PolyPhen2 and has a MAF of 0.25. Valine 160 is a highly conserved residue within the SRCS domain. The SRCS is found in proteins involved in host defence, such as CD5 and CD6, but its role in TMPRSS2 remains unknown. 84 rare variants (53 missense and 31 leading to a prematurely truncated protein, cumulative minor allele frequency (MAF) 7.34×10−4) cause structural destabilization and possibly protein misfolding, and are also predicted damaging by SIFT and PolyPhen2 prediction tools. Moreover, we extracted gene expression data from the human protein atlas and showed that both ACE2 and TMPRSS2 are expressed in the small intestine, duodenum and colon, as well as the kidneys and gallbladder.The implications of our study are that: i. TMPRSS2 variants, in particular p.V160M with a MAF of 0.25, should be investigated as a possible marker of disease severity and prognosis in COVID-19 and ii. in vitro validation of the co-expression of TMPRSS2 and ACE2 in gastro-intestinal is warranted.
Pharmacokinetics of anti-tuberculosis drugs in multidrug resistant tuberculosis patients in India
Authors: Hemanth Kumar AK , Natarajan PL , Kannan T , Sridhar R , Kumar S , Vinod Kumar V , Gomathi NS , Bharathiraja T , Sudha V , Balaji S , Rameshkumar S , Nair Dina , Tripathy SP , Geetha R 
Published: May 2020

AbstractProgrammatic Management of multidrug resistant tuberculosis (MDR TB) services were introduced in the Indian TB control programme in 2007. A pharmacokinetic (PK) study of drugs used to treat MDR TB, namely levofloxacin (LFX), ethionamide (ETH), cycloserine (CS), pyrazinamide (PZA), moxifloxacin (MFX) and isoniazid (INH) was undertaken in adult MDR TB patients treated according to the prevailing guidelines in India. Factors influencing drug PK and end-of-intensive phase (IP) status were also determined. We recruited 350 MDR TB patients receiving anti-TB treatment (ATT) in the Indian Government programme in south India. At steady state, serial blood samples were collected, after supervised drug administration. Status at end of IP was noted from the programme records. Of the 303 patients for whom end-of-IP status was known, 214 were culture negative (responders), while 45 patients were either culture positive or required change of regimen or had died before completion of IP (non-responders). The median Cmax (2.0 vs 2.9μg/ml; p = 0.005) and AUC0-12 (12.2 vs 17.0μg/ml.h; p = 0.002) of ETH were significantly lower in non-responders than responders at IP. In multivariate logistic regression analysis, after excluding defaulters and adjusting for confounders, AUC0-12 of ETH significantly influenced end-of-IP status (aOR - 1.065; 95% CI: 1.001 - 1.134; p = 0.047). Drug doses used currently in the programme produced optimal drug concentrations in majority of patients. ETH played a major role in the MDR TB combination regimen and was a key determinant of end-of-IP status.
A pedagogical protocol for iterative robotic fabrication on remote grounds
Authors: Bar-Sinai Karen Lee , Shaked Tom , Sprecher Aaron 
Published: May 2020

PurposeThe purpose of the paper is to advance remote robotic fabrication through an iterative and pedagogical protocol for shaping architectural grounds. Advancements in autonomous robotic tools enable to reach increasingly larger scales of architectural and landscape construction and operate in remote and inaccessible sites. In parallel, the relation of architecture to its environment is significantly reconsidered, as the building industry's contribution to the environmental stress increases. In response, new practices emerge, addressing the reshaping and modulation of environments using digital tools. The context of extra-terrestrial architecture provides a ground for exploring these issues, as future practice in this domain relies on the use of remote autonomous means for repurposing local matter. As a result, the novelty in robotic construction laboratories is tied to innovation in architectural pedagogy.Design/methodology/approachThis paper puts forth a pedagogical protocol and iterative framework for digital groundscaping using robotic tools. The framework is demonstrated through an intensive workshop led by the authors. To situate the discussion, digital groundscaping is linked to several conditions that characterize practice and relate to pedagogy. These conditions include the experimental dimension of knowledge in digital fabrication, the convergence of knowledge as part of the blur between the fields of architecture and landscape architecture and the bridging of heterogeneous knowledge sets (virtual and physical), which robotic fabrication on natural terrains entails.FindingsThe outcomes of the workshop indicate that iterative processes can assist in applying autonomous design protocols on remote grounds. The protocols were assessed in light of the roles of technological tools, design iterations and material agency in the robotic fabrication.Originality/valueThe paper concludes with observations linking the iterative protocol to new avenues in architectural pedagogy as means of advancing the capacity to digitally design, modulate and transform natural grounds.
Factors leading to design changes in Jordanian construction projects
Authors: Gharaibeh Lina Ghazi , Matarneh Sandra T. , Arafeh Mazen , Sweis Ghaleb 
Published: May 2020

PurposeThe problem of design changes in the construction industry is common worldwide, and the Jordanian market is no exception. The purpose of this paper is to identify the factors causing design changes in construction projects in Jordan in both the public and private sectors. Furthermore, this research will examine the effect of these factors on project's performance during the construction phase.Design/methodology/approachThis research commences by identifying the factors causing design changes in construction projects worldwide through an intensive literature review. The identified factors were then filtered to those applicable to the Jordanian construction market based on the results obtained from a questionnaire survey and real case construction projects. In total, 252 professionals from the Jordanian construction industry and 10 completed and/or ongoing construction projects in different parts of Jordan were compared.FindingsThe results find that the top major factors affecting design changes are owner's requirements; design errors and omissions and value engineering. The research also studies and documents the impacts of design changes on project cost, schedule and quality.Originality/valueThe results obtained from this research will assist the construction professionals representing owners, consultants and contractors in applying control measures to minimize the occurrence of the identified factors causing design changes and to mitigate their sever impacts on projects in terms of cost, schedule and quality.
Distribution channels of vegetable industry in Indonesia: impact on business performance
Authors: Mariyono Joko , Waskito Jaka , Kuntariningsih Apri , Gunistiyo Gunistiyo , Sumarno Sumarno 
Published: May 2020

Purpose
The purpose of this paper is to analyse the distribution channels of vegetable sectors in Indonesia, its economic impact on the performance of vegetable sales and the factors affecting marketing channels selected by producers.

Design/methodology/approach
The study employed qualitative and quantitative methods. A market survey was qualitatively conducted at producer, intermediary, wholesaler, hotel and food processing company as well as retailer levels. Producer survey was quantitatively conducted at the farm level, by interviewing 556 randomly selected farm households. Structural equation modelling was employed to accomplish the objectives of the study.

Findings
Marketing channels for vegetables in Indonesia was complex and relatively long. Farmers decided to select particular channels because of business circumstance and their knowledge. Distance and gentleman’s agreement with traders limited farmers to choose the desirable marketing channel. Marketing channels affect business performance in terms of high sales and profit.

Research limitations/implications
This study only pays attention to the supply side of vegetables. The effect of marketing channels also encumbers the consumers, which are beyond this study. Other studies are expected to highlight the consumer side.

Originality/value
This study focused on smallholder agribusiness players. This study uses two surveys as data sources: market survey and producer survey. The market survey serves as vital information to design producer surveys.