bluuebunny/crossref_metadata_2025_split huggingface dataset:
Utilizing a nanobody recruitment approach for assessing serine palmitoyltransferase activity in ER sub-compartments of yeast
Authors: Esch Bianca M. , Walter Stefan , Schmidt Oliver , Fröhlich Florian 
Published: March 2023

AbstractSphingolipids (SP) are one of the three major lipid classes in eukaryotic cells and serve as structural components of the plasma membrane. The rate-limiting step in SP biosynthesis is catalyzed by serine palmitoyltransferase (SPT). In yeast, SPT consists of two catalytic subunits (Lcb1 and Lcb2), a regulatory subunit (Tsc3), negative regulators (Orm1 and Orm2), and the phosphatidylinositol-4-phosphate (PI4P) phosphatase Sac1, collectively known as the SPOTS complex. Regulating SPT activity enables cells to adapt SP metabolism to changing environmental conditions. Therefore, the Orm proteins are phosphorylated by two signaling pathways originating from either the plasma membrane localized target of rapamycin (TOR) complex 2 or the lysosomal/vacuolar TOR complex 1. Moreover, uptake of exogenous serine is necessary for the regulation of SP biosynthesis, which suggests the existence of differentially regulated SPT pools based on their intracellular localization. However, tools for measuring lipid metabolic enzyme activity in different cellular compartments are currently not available. We have developed a nanobody recruitment system that enables the re-localization of the SPOTS complex to the nuclear or peripheral ER. By combining this system with sphingolipid flux analysis, we have identified two distinct active SPT pools in cells. Our method thus serves as a new and versatile tool to measure lipid metabolism with sub-cellular resolution.
Versatile Product Detection via Coupled Assays for Ultra-high-throughput Screening of Carbohydrate-Active-Enzymes in Microfluidic Droplets
Authors: Ladeveze Simon , Zurek Paul J. , Kaminski Tomasz S. , Emond Stephane , Hollfelder Florian 
Published: March 2023

ABSTRACTEnzyme discovery and directed evolution are the two major contemporary approaches for the improvement of industrial processes by biocatalysis in various fields. Customization of catalysts for improvement of single enzyme reactions or de novo reaction development is often complex and tedious. The success of screening campaigns relies on the fraction of sequence space that can be sampled, whether for evolving a particular enzyme or screening metagenomes. Ultrahigh-throughput screening (uHTS) based on in-vitro compartmentalization in water-in-oil emulsion of picolitre droplets generated in microfluidic systems allows screening rates >1 kHz (or >107 per day). Screening for Carbohydrate Active Enzymes (CAZymes) catalysing biotechnologically valuable reactions in this format presents an additional challenge, because the released carbohydrates are difficult to monitor in high throughput. Activated substrates with large optically active hydrophobic leaving groups provide a generic optical readout, but the molecular recognition properties of sugars will be altered by incorporation of such fluoro- or chromophores and their typically higher reactivity, as leaving groups with lowered pKa values compared to native substrates make observation of promiscuous reactions more likely. To overcome these issues, we designed microdroplet assays in which optically inactive carbohydrate products are made visible by specific cascades: the primary reaction of an unlabelled substrate leads to an optical signal downstream. Successfully implementing such assays at the picoliter droplet scale allowed us to detect glucose, xylose, glucuronic acid and arabinose as final products of complex oligosaccharide degradation by glycoside hydrolases by absorbance measurements. Enabling the use of uHTS for screening CAZyme reactions that have been thus far elusive will chart a route towards faster and easier development of specific and efficient biocatalysts for biovalorisation, directing enzyme discovery towards catalysts for their natural rather than model substrates.Graphical abstract / TOC
Altered neurological and neurobehavioral phenotypes in a mouse model of the recurrentKCNB1-p.R306C voltage-sensor variant
Authors: Kang Seok Kyu , Hawkins Nicole A. , Echevarria-Cooper Dennis M. , Baker Erin M. , Dixon Conor J. , Speakes Nathan , Kearney Jennifer A. 
Published: March 2023

AbstractPathogenic variants inKCNB1are associated with a neurodevelopmental disorder spectrum that includes global developmental delays, cognitive impairment, abnormal electroencephalogram (EEG) patterns, and epilepsy with variable age of onset and severity. Additionally, there are prominent behavioral disturbances, including hyperactivity, aggression, and features of autism spectrum disorder. The most frequently identified recurrent variant isKCNB1-p.R306C, a missense variant located within the S4 voltage-sensing transmembrane domain. Individuals with the R306C variant exhibit mild to severe developmental delays, behavioral disorders, and a diverse spectrum of seizures. Previousin vitrocharacterization of R306C described loss of voltage sensitivity and cooperativity of the sensor and inhibition of repetitive firing. ExistingKcnb1mouse models include dominant negative missense variants, as well as knockout and frameshifts alleles. While all models recapitulate key features ofKCNB1encephalopathy, mice with dominant negative alleles were more severely affected. In contrast to existing loss-of-function and dominant-negative variants,KCNB1-p.R306C does not affect channel expression, but rather affects voltage-sensing. Thus, modeling R306C in mice provides a novel opportunity to explore impacts of a voltage-sensing mutation inKcnb1. Using CRISPR/Cas9 genome editing, we generated theKcnb1R306Cmouse model and characterized the molecular and phenotypic effects. Heterozygous and homozygous R306C mice exhibited pronounced hyperactivity, altered susceptibility to flurothyl and kainic acid induced-seizures, and frequent, long runs of spike wave discharges on EEG. This novel model of channel dysfunction inKcnb1provides an additional, valuable tool to studyKCNB1encephalopathies. Furthermore, this allelic series ofKcnb1mouse models will provide a unique platform to evaluate targeted therapies.
CsrA Shows Selective Regulation of sRNA-mRNA Networks
Authors: Rojano-Nisimura Alejandra Matsuri , Simmons Trevor R. , Leistra Abigail N. , Mihailovic Mia K. , Buchser Ryan , Ekdahl Alyssa M. , Joseph Isabella , Curtis Nicholas C. , Contreras Lydia M. 
Published: March 2023

AbstractPost-transcriptional regulation, by small RNAs (sRNAs) as well as the global Carbon Storage Regulator A (CsrA) protein, play critical roles in bacterial metabolic control and stress responses. The CsrA protein affects selective sRNA-mRNA networks, in addition to regulating transcription factors and sigma factors, providing additional avenues of cross talk between other stress-response regulators. Here, we expand the known set of sRNA-CsrA interactions and study their regulatory effects.In vitrobinding assays confirm novel CsrA interactions with ten sRNAs, many of which are previously recognized as key regulatory nodes. Of those 10 sRNA, we identify that McaS, FnrS, SgrS, MicL, and Spot42 interact with CsrAin vivo. We find that the presence of CsrA impacts the downstream regulation of mRNA targets of the respective sRNA.In vivoevidence supports enhanced CsrA McaS-csgDmRNA repression and showcase CsrA-dependent repression of thefucPmRNA via the Spot42 sRNA. We additionally identify SgrS and FnrS as potential new sRNA sponges of CsrA. Overall, our results further support the expanding impact of the Csr system on cellular physiology via CsrA impact on the regulatory roles of these sRNAs.
Anti-CRISPR protein mediated degradation of Cas9 in human cells
Authors: de Tacca Luisa Arake , Bondy-Denomy Joseph , Rabuka David , Schelle Michael 
Published: March 2023

AbstractBacteriophages encode anti-CRISPR (Acr) proteins that inactivate CRISPR-Cas bacterial immune systems, allowing successful invasion, replication, and prophage integration. Acr proteins inhibit CRISPR-Cas systems using a wide variety of mechanisms. AcrIIA1 is encoded by numerous phages and plasmids, binds specifically to the Cas9 HNH domain, and was the first Acr discovered to inhibit SpyCas9. Here we report the observation of AcrIIA1-induced degradation of SpyCas9 and SauCas9 in human cell culture, the first example of Acr-induced degradation of CRISPR-Cas nucleases in human cells. Optimized expression of AcrIIA1 in human cells provided robust inhibition of SpyCas9 editing but had no impact on Type V CRIPSR-Cas12a, consistent with its binding site on the HNH domain. Targeted Cas9 protein degradation by AcrIIA1 could modulate Cas9 nuclease activity in human therapies. The small size and specificity of AcrIIA1 could be used in a CRISPR-Cas proteolysis-targeting chimera (PROTAC), providing a tool for developing safe and precise gene editing applications.
Cell wall proteomics in liveMycobacterium tuberculosisuncovers exposure of ESX substrates to the periplasm
Authors: Jaisinghani Neetika , Previti Mary L. , Andrade Joshua , Askenazi Manor , Ueberheide Beatrix , Seeliger Jessica C. 
Published: March 2023

SUMMARYThe cell wall of mycobacteria plays a key role in interactions with the environment and its ability to act as a selective filter is crucial to bacterial survival. Proteins in the cell wall enable this function by mediating the import and export of diverse metabolites from ions to lipids to proteins. Accurately identifying cell wall proteins is an important step in assigning function, especially as many mycobacterial proteins lack functionally characterized homologues. Current methods for protein localization have inherent limitations that reduce accuracy. Here we showed that protein tagging by the engineered peroxidase APEX2 within liveMycobacterium tuberculosisenabled the accurate identification of the cytosolic and cell wall proteomes. Our data indicate that substrates of the virulence-associated Type VII ESX secretion system are exposed to theMtbperiplasm, providing insight into the currently unknown mechanism by which these proteins cross the mycobacterial cell envelope.
Hydrogel Capsule-Based Digital Quantitative Polymerase Chain Reaction
Authors: Tan Zheng Lin , Yasuura Masato , Horiguchi Yukichi , Ashiba Hiroki , Fukuda Takashi 
Published: March 2023

AbstractDroplet digital PCR (ddPCR) is accurate in nucleic acid quantification owing to its linearity and high sensitivity. Amplification of nucleic acid in droplets, however, is limited by the stability of droplets against thermal cycling. While the use of fluorinated oil or supplementation of surfactant could improve the stability of droplets, this process has also greatly increased the cost of ddPCR and limited post-PCR analysis. Here, we report a novel method known as gel capsule-based digital PCR (gc-dPCR) which includes method to prepare hydrogel capsules encapsulating PCR reaction mix, conducting PCR reaction, and readout by either qPCR system or fluorescence microplate reader. We have compared our method to existing methods, i.e., quantitative PCR or vortex ddPCR. Our approach results in higher fluorescence intensity compared to ddPCR suggesting higher sensitivity of the system. As hydrogel capsules are more stable than droplets in fluorinated oil throughout thermal cycling, all partitions can be quantified, thus preventing loss of information from low-concentration samples. Our approach should extend to all droplet-based PCR methods. Generally, our approach has greatly improved ddPCR by increasing droplets stability and sensitivity, and reducing the cost of ddPCR, which help to remove the barrier of ddPCR in settings with limited resources.Abstract Figure
Improving mRNA vaccine safety and efficiency with cationized lipid nanoparticle formula
Authors: Peng Xu , Liao Guangneng , Ren Dongsheng , Zhou Yucheng , Wu Xiujin , Lei Yingxue , Zhang Yan , Chen Liang , He Chen , Zhang Yaoyi , Yin Hailin , Yang Guang , Xu Kai 
Published: March 2023

AbstractThe widespread use of Covid-19 mRNA vaccines has highlighted the need to address rare but concerning side effects. Systemic off-target gene expression has been identified as a primary cause of acute adverse reactions and side effects associated with nucleoside-modified mRNA vaccines. In this study, we incorporated the permanent cationic lipid Dotap component into the mRNA-LNP formula associated with the FDA-approved mRNA vaccine Comirnaty to create a novel positively charged LNP carrier for mRNA vaccine delivery. Using the optimized LNP formula to prepare SARS-Cov-2 Spike mRNA vaccines for immunogenicity testing, Balb/c mice exhibited improved immunogenicity kinetics with initial antibody titers being lower but showing a continuous upward trend, ultimately reaching levels comparable to those of control mRNA vaccines 8 weeks after boost immunization. The mRNA vaccines encapsulated in the modified LNPs have demonstrated a superior safety profile in respect to systemic delivery of LNP constituents, off-target gene expression, and the systemic pro-inflammatory stimulation. Consequently, it may represent a safer alternative of conventional mRNA-LNP vaccines.
ChatGPT in Healthcare: A Taxonomy and Systematic Review
Authors: Li Jianning , Dada Amin , Kleesiek Jens , Egger Jan 
Published: March 2023

AbstractThe recent release of ChatGPT, a chat bot research project/product of natural language processing (NLP) by OpenAI, stirs up a sensation among both the general public and medical professionals, amassing a phenomenally large user base in a short time. This is a typical example of the ‘productization’ of cutting-edge technologies, which allows the general public without a technical background to gain firsthand experience in artificial intelligence (AI), similar to the AI hype created by AlphaGo (DeepMind Technologies, UK) and self-driving cars (Google, Tesla, etc.). However, it is crucial, especially for healthcare researchers, to remain prudent amidst the hype. This work provides a systematic review of existing publications on the use of ChatGPT in healthcare, elucidating the ‘status quo’ of ChatGPT in medical applications, for general readers, healthcare professionals as well as NLP scientists. The large biomedical literature databasePubMedis used to retrieve published works on this topic using the keyword ‘ChatGPT’. An inclusion criterion and a taxonomy are further proposed to filter the search results and categorize the selected publications, respectively. It is found through the review that the current release of ChatGPT has achieved only moderate or ‘passing’ performance in a variety of tests, and is unreliable for actual clinical deployment, since it is not intended for clinical applications by design. We conclude that specialized NLP models trained on (bio)medical datasets still represent the right direction to pursue for critical clinical applications.
WITHDRAWN: Assessing the Causal Effects of Environmental Tobacco Smoke Exposure: A meta-analytic Mendelian randomisation study
Authors: Woolf Benjamin , Rajasundaram Skanda , Gill Dipender , Sallis Hannah M. , Munafò Marcus R. 
Published: March 2023

Withdrawal StatementThe authors have withdrawn their manuscript owing to an issue, which they were recently alerted to, with GWAS of paternal smoking used here. They hope to update the manuscript with an alternative GWAS as soon as possible. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
Validation of the Strengths and Difficulties Questionnaire (SDQ) emotional subscale in assessing depression and anxiety across development
Authors: Armitage Jessica M , Tseliou Foteini , Riglin Lucy , Dennison Charlotte , Eyre Olga , Bevan Jones Rhys , Rice Frances , Thapar Ajay K , Thapar Anita , Collishaw Stephan 
Published: March 2023

AbstractEmotional disorders are common in childhood, and their prevalence sharply increases during adolescence. The Strengths and Difficulties Questionnaire (SDQ) is widely used for screening emotional and behavioural difficulties in children and young people, but little is known about the accuracy of the emotional subscale (SDQ-E) in detecting emotional disorders, and whether this changes over development. Such knowledge is important in determining whether symptom changes across age are due to developmental or measurement differences. This study assessed the validity of the SDQ-E and two individual items (low mood and general worry) in differentiating between cases and non-cases of Major Depressive Disorder (MDD), Generalised Anxiety Disorder (GAD), and other anxiety disorders across ages 7, 10, 13, 15, and 25 years in a UK population cohort. Analyses showed moderate accuracy of the subscale in discriminating cases of MDD (AUC=0.67-0.85), and high accuracy for discriminating cases of GAD (AUC=0.80-0.93) and any anxiety disorder (AUC=0.74-0.83) compared to non-cases. The SDQ-E performed well across ages and sex, and generally performed better than the two individual items. Together our findings validate the SDQ-E as a screen for emotional disorders during childhood, adolescence, and early adulthood, and as a tool for longitudinal research on depression and anxiety disorders.
Evaluation of factors determining the development and progress of library and information science in different Pakistani universities
Authors: Shah Suhaib Hussain , Pei Lei , Chen Tianyu 
Published: March 2023

Purpose
The field of library and information sciences (LIS) is crucial to our educational system. Across the globe, the LIS systems operate at varying levels and rates of efficiency. One of the developing nations is Pakistan, which has LIS systems in all of its colleges, universities and schools. This multimethod study aims to identify and quantify elements that are detrimental to LIS progress as well as evaluate the faculty and infrastructure profiles of universities that offer LIS undergraduate and graduate level programmes.

Design/methodology/approach
Data was collected from the study's participants, who were mainly LIS professionals and faculty at 17 different universities, using survey questionnaires and in-person interviews. This study used a descriptive survey methodology, gathering information through a Google Survey and filling it out with a premade survey proforma. The survey responses were examined using content analysis. The development of LIS instructional and scholarly output is influenced by a variety of factors, which were investigated using a generalized linear model (GLM). To determine whether there was a statistically significant difference in opinion between faculty members and working professionals, as well as between men and women, the outcomes of an independent sample t-test were examined.

Findings
According to our data, the factors that have the biggest impact on the caliber and output of LIS research are “poor writing skills” (3.43), “lack of journal publication fees” (3.51) and “lack of research skills” (3.78). The top GLM model identified poor writing skills, a lack of publication fees and a lack of research expertise as bottleneck characteristics for producing high-quality LIS research. The aforementioned factors were 3.62, 2.41 and 2.19 times more significant than the average factor, respectively, to put it another way.

Originality/value
This study’s analysis revealed that there is no real distinction between the two groups' viewpoints. The results of this study can be applied to problems and challenges associated with LIS education in Pakistani educational institutions.
Explore the relationship between leadership skills, organizational citizenship behavior and organizational culture in Vietnamese public organizations
Authors: Tran Quan Hoang Nguyen 
Published: March 2023

PurposeThe purpose of this study is to look into the relationship between leadership skills, especially technical, human and conceptual skills, and organizational citizenship behavior (OCB), with organizational culture acting as a mediator.Design/methodology/approachFor the purpose of this study, a quantitative strategy was adopted. To collect data, paper/online surveys were administered to 294 individuals in various Vietnamese public organizations. Stata 15.1 was used to assess the reliability, validity and consistency of the measurements. Regression analysis was used to evaluate the proposed hypotheses.FindingsIt was discovered that leadership skills have a significant effect on OCB, which partially supported the hypotheses. However, only conceptual skill was found to have a significant impact on individual OCB (OCBI). Conceptual skill was also found to have a significant impact on supportive culture. In addition, only supportive culture had a significant effect on OCBI. Finally, supportive culture was confirmed to mediate the relationship between conceptual skill and OCBI.Originality/valueThis study adds to the social exchange theory by filling three significant gaps in previous research on leadership skills. First, it investigates the impact of three leadership skills on OCB in public organizations. Second, it examines the role of leadership skills in fostering an organizational culture. Third, the paper concludes by investigating the underlying mechanism through which leadership skills contribute to OCB via the mediation of organizational culture. The implications of these findings for practice are discussed.
Applications of six sigma for service quality enhancement in the UAE: a multiple case study analysis and lessons learned
Authors: Bhat Shreeranga , Antony Jiju , Maalouf Maher , E.V. Gijo , Salah Souraj 
Published: March 2023

Purpose
This paper aims to unearth the essential components of Six Sigma for successful deployment and sustainment of service quality in four different organizations in the United Arab Emirates (UAE). More specifically, the paper is intended to determine the motivation to apply Six Sigma, Voice of Customer, Key Performance Indicators (KPIs), Critical-to-Quality (CTQ), readiness factors, Critical Success Factors, sustainment measures, tools and techniques used, challenges/barriers and performance impact on the company.

Design/methodology/approach
An exploratory research methodology with multiple case study analyses was adopted to determine the answers to the research objectives. Four case studies from different service processes of four companies were analyzed. The case studies were collated from these companies via a case study protocol with pre-defined criteria.

Findings
The analysis shows that service operation improvement projects are primarily dependent on the voice of the internal customer, with return on investment in savings as the KPI of the process. Most organizations prefer cycle time and errors as the CTQs in the Six Sigma projects. Even novice users can effectively apply the Six Sigma methodology with external experts’ assistance, mentoring and interventions. Across the case studies, it is observed that the projects were successfully deployed due to the support of top management leadership, effective communication and cross-functional teams. Employee resistance to change is the common barrier observed during the case study analysis. Eventually, in all the four case studies, Six Sigma is executed with standard tools and techniques within the define, measure, analyze, improve, control (DMAIC) approach.

Research limitations/implications
The present study’s findings cannot be generalized due to the limited number of case study analyses in different ecosystems in the UAE. The authors would like to analyse and report more case studies in service quality improvement through the Six Sigma methodology to comprehend and develop a generic roadmap for the deployment of Six Sigma in the UAE service industry.

Practical implications
The study’s findings provide insights into commonalities and differences between the essential factors of Six Sigma deployment and sustainability in UAE companies.

Originality/value
The study results might help the policymakers and key decision makers in UAE and other countries understand the effectiveness of Six Sigma in service quality improvement with its essential factors for deployment.