bluuebunny/crossref_metadata_2025 huggingface dataset:
Severe and common mental disorders and risk of hospital admissions for Ambulatory Care Sensitive Conditions (ACSCs): prospective cohort study using UK Biobank
Authors: Niedzwiedz Claire L. , Aragón María José , Breedvelt Josefien J. F. , Smith Daniel J , Prady Stephanie L. , Jacobs Rowena
Published: January 2022

Background

People with mental disorders have an excess chronic disease burden. One mechanism to potentially reduce the public health and economic costs of mental disorders is to reduce preventable hospital admissions. Ambulatory care sensitive conditions (ACSCs) are a defined set of chronic and acute illnesses not considered to require hospital treatment if patients receive adequate primary healthcare. We examined the relationship between both severe and common mental disorders and risk of emergency hospital admissions for ACSCs and factors associated with increased risk.

Methods

Baseline data from England (N=445,814) were taken from UK Biobank, which recruited participants aged 37-73 years during 2006 to 2010, and were linked to hospital admission records up to 31st December 2019. Participants were grouped into those who had a history of either schizophrenia, bipolar disorder, depression or anxiety, or no record of mental disorder. Cox proportional hazard models (for the first admission) and Prentice, Williams and Peterson Total Time models (PWP-TT, which account for all admissions) were used to assess the risk (using hazard ratios (HR)) of hospitalisation for ACSCs among those with mental disorders compared to those without, adjusting for factors in different domains, including sociodemographic (e.g. age, sex, ethnicity), socioeconomic (e.g. deprivation, education level), health and biomarkers (e.g. multimorbidity, inflammatory markers), health-related behaviours (e.g. smoking, alcohol consumption), social isolation (e.g. social participation, social contact) and psychological (e.g. depressive symptoms, loneliness).

Results

People with schizophrenia had the highest risk of hospital admission for ACSCs compared to those with no mental disorder (HR=4.40, 95% CI: 4.04 - 4.80). People with bipolar disorder (HR=2.48, 95% CI: 2.28 – 2.69) and depression or anxiety (HR=1.76, 95% CI: 1.73 – 1.80) also had higher risk. Associations were more conservative when accounting for all admissions. Although adjusting for a range of factors attenuated the observed associations, they still persisted, with socioeconomic and health-related variables contributing most.

Conclusions

People with severe mental disorders had highest risk of preventable hospital admissions, with the risk also elevated amongst those with depression and anxiety. Ensuring people with mental disorders receive adequate ambulatory care is essential to reduce the large health inequalities experienced by these groups.
Vaccinating children against COVID-19: commentary and mathematical modelling
Authors: Hawkes Michael T. , Good Michael F.
Published: January 2022

With the recent licensure of mRNA vaccines against COVID-19 in the 5-11 year old age group, the public health impact of a childhood immunization campaign is of interest. Using a mathematical epidemiological model, we project that childhood vaccination carries minimal risk and yields modest public health benefits. These include large relative reductions in child morbidity and mortality, although the absolute reduction is small because these events are rare. Furthermore, the model predicts “altruistic” absolute reductions in adult cases, hospitalizations, and mortality. However, vaccinating children to benefit adults should be considered from an ethical as well as a public health perspective. From a global health perspective, an additional ethical consideration is the justice of giving priority to children in high-income settings at low risk of severe disease while vaccines have not been made available to vulnerable adults in low-income settings.
Genomic insights into the phylogeny and biomass-degrading enzymes of rumen ciliates
Authors: Li Zongjun , Wang Xiangnan , Zhang Yu , Yu Zhongtang , Zhang Tingting , Dai Xuelei , Pan Xiangyu , Jing Ruoxi , Yan Yueyang , Liu Yangfan , Gao Shan , Li Fei , Huang Youqin , Tian Jian , Yao Junhu , Xing XvPeng , Shi Tao , Ning Jifeng , Yao Bin , Huang Huoqing , Jiang Yu
Published: January 2022

Understanding the biodiversity and genetics of the gut microbiome has important implications for host physiology. One underexplored and elusive group is ciliated protozoa, which play crucial roles in regulating gut microbial interactions. Integrating single-cell sequencing and an assembly-and-identification pipeline, we acquired 52 high-quality ciliate genomes of 22 rumen morphospecies for all major abundant clades. With these genomes, we firstly resolved the taxonomic and phylogenetic framework that reclassified them into 19 species spanning 13 genera and reassigned the genus Dasytricha from Isotrichidae to a new family Dasytrichidae. Via extensive horizontal gene transfer and gene family expansion, rumen ciliates possess a broad array of enzymes to synergistically degrade plant and microbial carbohydrates. In particular, ∼80% of the degrading enzymes in Diplodiniinae and Ophryoscolecinae act on plant cell wall, and the high activities of their cellulase, xylanase and lysozyme reflect the potential of ciliate enzymes for biomass-conversion. Additionally, the new ciliate dataset greatly facilitated the rumen metagenomic analyses by allowing ∼12% of reads to be classified.
Limited consequences for loss of RNA-directed DNA methylation in Setaria viridis domains rearranged methyltransferase (DRM) mutants
Authors: Read Andrew , Weiss Trevor , Crisp Peter A , Liang Zhikai , Noshay Jaclyn , Menard Claire C , Wang Chunfang , Song Meredith , Hirsch Candice N , Springer Nathan M , Zhang Feng
Published: January 2022

The Domains Rearranged Methyltransferases (DRMs) are crucial for RNA-directed DNA methylation (RdDM) in plant species. Setaria viridis is a model monocot species with a relatively compact genome that has limited transposable element content. CRISPR-based genome editing approaches were used to create loss-of-function alleles for the two putative functional DRM genes in S. viridis to probe the role of RdDM. The analysis of drm1ab double mutant plants revealed limited morphological consequences for the loss of RdDM. Whole-genome methylation profiling provided evidence for wide-spread loss of methylation in CHH sequence contexts, particularly in regions with high CHH methylation in wild-type plants. Evidence was also found for locus-specific loss of CG and CHG methylation, even in some regions that lack CHH methylation. Transcriptome profiling identified a limited number of genes with altered expression in the drm1ab mutants. The majority of genes with elevated CHH methylation directly surrounding the transcription start site or in nearby promoter regions do not have altered expression in the drm1ab mutant even when this methylation is lost, suggesting limited regulation of gene expression by RdDM. Detailed analysis of the expression of transposable elements identified several transposons that are transcriptionally activated in drm1ab mutants. These transposons likely require active RdDM for maintenance of transcriptional repression.

Significance statement

Methylation profiling of Setaria viridis plants that lack functional Domains Rearranged Methyltransferase genes reveal widespread loss of DNA methylation in the CHH sequence context. Transcriptome analysis reveals a small set of genes and transposons that are silenced by RNA-directed DNA methylation.
Insights into I-motif stabilization by high resolution primer extension assays: Its strengths and limitations
Authors: Jamroskovic Jan , Deiana Marco , Sabouri Nasim
Published: January 2022

Cytosine-rich DNA can fold into four-stranded intercalated structures, i-motif (iM), in acidic pH and require hemi-protonated C:C+ base pairs to form. However, its formation and stability rely on many other factors that are not yet fully understood. In here, we combined biochemical and biophysical approaches to determine the factors influencing iM stability in a wide range of experimental conditions. By using high resolution primer extension assays, circular dichroism and absorption spectroscopies, we demonstrate that the stability of three different biologically relevant iMs are not dependent on molecular crowding agents. Instead, some of the crowding agents affected overall DNA synthesis. We also tested a range of small molecules to determine their effect on iM stabilization at physiological temperature, and demonstrated that the G-quadruplex-specific molecule, CX-5461, is also a promising candidate for selective iM stabilization. This work provides important insights into the requirements needed for different assays to accurately study iM stabilization, which will serve as important tools for understanding iMs’ biological roles and their potential as therapeutic targets.
Intracellular copper storage and delivery in a bacterium
Authors: Lee Jaeick , Dennison Christopher
Published: January 2022

A family of cytosolic copper (Cu) storage proteins (the Csps) are widespread in bacteria. The Csps can bind large quantities of Cu(I) via their Cys-lined four-helix bundles, and the majority are cytosolic (Csp3s). This is inconsistent with the current dogma that bacteria, unlike eukaryotes, have evolved not to maintain intracellular pools of Cu due to its potential toxicity. Sporulation in Bacillus subtilis has been used to investigate if a Csp3 can store Cu(I) in the cytosol for a target enzyme. The activity of the Cu-requiring endospore multi-Cu oxidase BsCotA (a laccase) increases under Cu-replete conditions in wild type B. subtilis, but not in the strain lacking BsCsp3. Cuprous ions readily transfer from BsCsp3, but not from the cytosolic copper metallochaperone BsCopZ, to BsCotA in vitro producing active enzyme. Both BsCsp3 and BsCotA are upregulated during late sporulation. The hypothesis we propose is that BsCsp3 acquires and stores Cu(I) in the cytosol for BsCotA.
Humanization of wildlife gut microbiota in urban environments
Authors: Dillard Brian A. , Chung Albert K. , Gunderson Alex R. , Campbell-Staton Shane C. , Moeller Andrew H.
Published: January 2022

Urbanization is rapidly altering Earth’s environments, demanding investigations of the impacts on resident wildlife. Here, we show that urban populations of coyotes (Canis latrans) and crested anole lizards (Anolis cristatellus) acquire gut microbiota constituents found in humans, including the gut bacterial lineages most significantly associated with urbanization in humans (e.g., Bacteroides). Comparisons of urban and rural wildlife and human populations revealed significant convergence of the gut microbiota among urban host populations. Remarkably, all microbial lineages found in humans that were overrepresented in urban wildlife relative to rural wildlife were also overrepresented in urban humans relative to rural humans. These results indicate parallel effects of urbanization on human and wildlife gut microbiota and suggest spillover of bacteria from humans into wildlife in cities.
Analytical time-dependent distributions for gene expression models with complex promoter switching mechanisms
Authors: Jia Chen , Li Youming
Published: January 2022

Classical gene expression models assume exponential switching time distributions between the active and inactive promoter states. However, recent experiments have shown that many genes in mammalian cells may produce non-exponential switching time distributions, implying the existence of multiple promoter states and molecular memory in the promoter switching dynamics. Here we analytically solve a gene expression model with random bursting and complex promoter switching, and derive the time-dependent distributions of the mRNA and protein copy numbers, generalizing the steady-state solution obtained in [SIAM J. Appl. Math. 72, 789-818 (2012)] and [SIAM J. Appl. Math. 79, 1007-1029 (2019)]. Using multiscale simplification techniques, we find that molecular memory has no influence on the time-dependent distribution when promoter switching is very fast or very slow, while it significantly affects the distribution when promoter switching is neither too fast nor too slow. By analyzing the dynamical phase diagram of the system, we also find that molecular memory in the inactive gene state weakens transient and stationary bimodality of the copy number distribution, while molecular memory in the active gene state enhances such bimodality.
Climatic niche conservatism in a clade of disease vectors (Diptera: Phlebotominae)
Authors: D’Agostino Emmanuel R.R. , Vivero Rafael , Romero Luis , Bejarano Eduar , Hurlbert Allen H. , Comeault Aaron A. , Matute Daniel R.
Published: January 2022

Sandflies of the family Psychodidae show notable diversity in both disease vector status and climatic niche. Some species (in the subfamily Phlebotominae) transmitLeishmaniaparasites, responsible for the disease leishmaniasis. Other Psychodidae species do not. Psychodid species’ ranges can be solely tropical, confined to the temperate zones, or span both. Studying the relationship between the evolution of disease vector status and that of climatic niche affords an understanding not only of the climate conditions associated with the presence and species richness ofLeishmaniavectors, but also allows the study of the extent to which psychodid flies’ climatic niches are conserved, in a context with implications for global human health. We obtained observation site data, and associated climate data, for 223 psychodid species to understand which aspects of climate most closely predict distribution. Temperature and seasonality are strong determinants of species occurrence within the clade. We built a mitochondrial DNA phylogeny of Psychodidae, and found a positive relationship between pairwise genetic distance and climate niche differentiation, which indicates strong niche conservatism. This result is also supported by strong phylogenetic signals of metrics of climate differentiation. Finally, we used ancestral trait reconstruction to infer the tropicality (i.e., proportion of latitudinal range in the tropics minus the proportion of the latitudinal range in temperate areas) of ancestral species, and counted transitions to and from tropicality states, finding that tropical and temperate species respectively produced almost entirely tropical and temperate descendant species, a result consistent for vector and non-vector species. Taken together, these results imply that while vectors ofLeishmaniacan survive in a variety of climates, their climate niches are strongly predicted by phylogeny.
Mutant KRAS-associated proteome is mainly controlled by exogenous factors
Authors: Carvalho Patrícia Dias , Martins Flávia , Carvalho Joana , Oliveira Maria José , Velho Sérgia
Published: January 2022

KRAS signaling has been extensively studied, yet the clarification between KRAS-autonomous and non-autonomous mechanisms are still less explored. Understanding how KRAS signaling and effects are affected by exogenous stimuli can provide valuable insights not only to understand resistance mechanisms that justify pathway inhibition failure, but also to uncover novel therapeutic targets for mutant KRAS patients. Hence, aiming at understanding KRAS-autonomous versus non autonomous mechanisms, we studied the response of two mutant KRAS colorectal cancer cell lines (HCT116 and LS174T) - control and KRAS silenced- to TGFβ1-activated fibroblasts secretome. By performing a total proteome analysis, we observed that TGFβ1-activated fibroblast-secreted factors triggered cell line-specific proteome alterations and that mutant KRAS governs approximately 1/3 of those alterations. Moreover, the analysis of the impact of exogenous factors on the modulation of KRAS proteome revealed that, in both cell lines, more than 2/3 of the KRAS-associated proteome is controlled in a KRAS-non-autonomous manner and dependent on the exogenous factors. This work highlights the context-dependency of KRAS-associated signaling and reinforces the importance of establishing more integrative models resembling the complexity of the tumor microenvironment to study KRAS-associated signals.
A CNN model with feature integration for MI EEG subject classification in BMI
Authors: Roy Arunabha M.
Published: January 2022

Objective

Electroencephalogram (EEG) based motor imagery (MI) classification is an important aspect in brain-machine interfaces (BMIs) which bridges between neural system and computer devices decoding brain signals into recognizable machine commands. However, the MI classification task is challenging due to inherent complex properties, inter-subject variability, and low signal-to-noise ratio (SNR) of EEG signals. To overcome the above-mentioned issues, the current work proposes an efficient multi-scale convolutional neural network (MS-CNN).

Approach

In the framework, discriminant user-specific features have been extracted and integrated to improve the accuracy and performance of the CNN classifier. Additionally, different data augmentation methods have been implemented to further improve the accuracy and robustness of the model.

Main results

The model achieves an average classification accuracy of 93.74% and Cohen’s kappa-coefficient of 0.92 on the BCI competition IV2b dataset outperforming several baseline and current state-of-the-art EEG-based MI classification models.

Significance

The proposed algorithm effectively addresses the shortcoming of existing CNN-based EEG-MI classification models and significantly improves the classification accuracy.
Neomorphic DNA-binding enables tumor-specific therapeutic gene expression in fusion-addicted childhood sarcoma
Authors: Hölting Tilman L. B. , Cidre-Aranaz Florencia , Matzek Dana , Popper Bastian , Jacobi Severin J. , Li Jing , Piseddu Ignazio , Cadilha Bruno L. , Ledderose Stephan , Zwilling Jennifer , Ohmura Shunya , Anz David , Künkele Annette , Klauschen Frederick , Grünewald Thomas G. P. , Knott Maximilian M. L.
Published: January 2022

Chimeric fusion transcription factors are oncogenic hallmarks of several devastating cancer types including pediatric sarcomas, such as Ewing sarcoma (EwS) and alveolar rhabdomyosarcoma (ARMS). Despite their exquisite specificity, these driver oncogenes have been considered largely undruggable due to their lack of enzymatic activity.

Here, we show in the EwS model that – capitalizing on neomorphic DNA-binding preferences – the addiction to the respective fusion transcription factor EWSR1-FLI1 can be leveraged to express therapeutic genes.

We genetically engineered a de novo enhancer-based, synthetic and highly potent expression cassette that can elicit EWSR1-FLI1-dependent expression of a therapeutic payload as evidenced by episomal and CRISPR-edited genomic reporter assays. Combining in silico screens and immunohistochemistry, we identified GPR64 as a highly specific cell surface antigen for targeted transduction strategies in EwS. Functional experiments demonstrated that anti-GPR64-pseudotyped lentivirus harboring our expression cassette can specifically transduce EwS cells to promote the expression of viral thymidine kinase sensitizing EwS for treatment to the otherwise relatively non-toxic (Val)ganciclovir and leading to strong anti-tumorigenic, but no adverse effects in vivo. Further, we prove that similar vector designs can be applied in PAX3-FOXO1-driven ARMS, and to express immunomodulatory cytokines, such as IL-15 and XCL1, in tumor types typically considered to be immunologically ‘cold’.

Collectively, these results generated in pediatric sarcomas indicate that exploiting, rather than suppressing, the neomorphic functions of chimeric transcription factors may open inroads to innovative and personalized therapies, and that our highly versatile approach may be translatable to other cancers addicted to oncogenic transcription factors with unique DNA-binding properties.
A Novel Virus Discovered in the Yeast Pichia membranifaciens
Authors: Lee Mark D. , Creagh Jack W. , Fredericks Lance R. , Crabtree Angela M. , Suresh Patel Jagdish , Rowley Paul A.
Published: January 2022

Mycoviruses are widely distributed across fungi, including yeasts of the Saccharomycotina subphylum. It was recently discovered that the yeast species Pichia membranifaciens contained double stranded RNAs (dsRNAs) that were predicted to be of viral origin. The fully sequenced dsRNA is 4,578 bp in length, with RNA secondary structures similar to the packaging, replication, and frameshift signals of totiviruses of the family Totiviridae. This novel virus has been named Pichia membranifaciens virus L-A (PmV-L-A) and is related to other totiviruses previously described within the Saccharomycotina yeasts. PmV-L-A is part of a monophyletic subgroup within the I-A totiviruses, implying a common ancestry between mycoviruses isolated from the Pichiaceae and Saccharomycetaceae yeasts. Energy minimized AlphaFold2 molecular models of the PmV-L-A Gag protein revealed structural conservation with the previously solved structure of the Saccharomyces cerevisiae virus L-A (ScV-L-A) Gag protein. The predicted tertiary structure of the PmV-L-A Pol and its homologs provide details of the potential mechanism of totivirus RNA-dependent RNA polymerases (RdRps) because of structural similarities to the RdRps of mammalian dsRNA viruses. Insights into the structure, function, and evolution of totiviruses gained from yeasts is important because of their parallels with mammalian viruses and the emerging role of totiviruses in animal disease.
Alternative splicing analysis benchmark with DICAST
Authors: Fenn Amit , Tsoy Olga , Faro Tim , Rössler Fanny , Dietrich Alexander , Kersting Johannes , Louadi Zakaria , Lio Chit Tong , Völker Uwe , Baumbach Jan , Kacprowski Tim , List Markus
Published: January 2022

Alternative splicing is a major contributor to transcriptome and proteome diversity in health and disease. A plethora of tools have been developed for studying alternative splicing in RNA-seq data. Previous benchmarks focused on isoform quantification and mapping. They neglected event detection tools, which arguably provide the most detailed insights into the alternative splicing process. DICAST offers a modular and extensible framework for the analysis of alternative splicing integrating 11 splice-aware mapping and eight event detection tools. We benchmark all tools extensively on simulated as well as whole blood RNA-seq data. STAR and HISAT2 demonstrated the best balance between performance and run time. The performance of event detection tools varies widely with no tool outperforming all others. DICAST allows researchers to employ a consensus approach to consider the most successful tools jointly for robust event detection. Furthermore, we propose the first reporting standard to unify existing formats and to guide future tool development.
Conserved meiotic mechanisms in the cnidarian Clytia hemisphaerica revealed by Spo11 knockout
Authors: Munro Catriona , Cadis Hugo , Pagnotta Sophie , Houliston Evelyn , Huynh Jean-René
Published: January 2022

During meiosis, DNA recombination allows the shuffling of genetic information between the maternal and paternal chromosomes. Recombination is initiated by double strand breaks (DSBs) catalyzed by the conserved enzyme Spo11. How this crucial event is connected to other meiotic processes is surprisingly variable depending on the species. Here, we knocked down Spo11 by CRISPR in the jellyfish Clytia hemisphaerica, belonging to Cnidaria, the sister group to Bilateria (where classical animal models are found). Spo11 mutants fail to assemble synaptonemal complexes and chiasmata, and in consequence homologous chromosome pairs disperse during oocyte growth, creating aneuploid but fertilizable eggs that develop into viable larvae. Clytia thus shares an ancient eukaryotic dependence of synapsis and chromosome segregation on Spo11-generated DSBs, and provides new evolutionary perspectives on meiosis regulation.
Discovery of small molecule antagonists of human Retinoblastoma Binding Protein 4 (RBBP4)
Authors: Perveen Sumera , Zepeda-Velázquez Carlos A. , McLeod David , Marcellus Richard , Mohammed Mohammed , Abbey Megha , Ensan Deeba , Panagopoulos Dimitrios , Trush Viacheslav , Gibson Elisa , Dong Aiping , Halabelian Levon , Poda Gennady , Owen Julie , Aman Ahmed , Kiyota Taira , Mamai Ahmed , Arrowsmith Cheryl H. , Brown Peter J. , Schapira Matthieu , Al-awar Rima , Vedadi Masoud
Published: January 2022

RBBP4 is a nuclear WD40 motif-containing protein widely implicated in various cancers and a putative drug target. It interacts with multiple proteins within diverse complexes such as nucleosome remodeling and deacetylase (NuRD) complex and polycomb repressive complex 2 (PRC2), as well as histone H3 and H4 through two distinct binding sites. B-cell lymphoma/leukemia 11A (BCL11A), friend of GATA-1 (FOG-1), plant homeodomain finger protein 6 (PHF6) and histone H3 bind to the top of the donut-shaped seven-bladed β-propeller fold of RBBP4, while suppressor of zeste 12 (SUZ12), metastasis associated protein 1 (MTA1) and histone H4 bind to a pocket on the side of the WD40 repeats of this protein. Here, we report the discovery of the first small molecule antagonists of the RBBP4 top pocket, competing with interacting peptides from proteins such as BCL11A and histone H3. We also determined the first crystal structure of RBBP4 in complex with a small molecule (OICR17251), paving the path for structure-guided design and optimization towards more potent antagonists.
Three million images and morphological profiles of cells treated with matched chemical and genetic perturbations
Authors: Chandrasekaran Srinivas Niranj , Cimini Beth A. , Goodale Amy , Miller Lisa , Kost-Alimova Maria , Jamali Nasim , Doench John G. , Fritchman Briana , Skepner Adam , Melanson Michelle , Kalinin Alexandr A. , Arevalo John , Haghighi Marzieh , Caicedo Juan , Kuhn Daniel , Hernandez Desiree , Berstler Jim , Shafqat-Abbasi Hamdah , Root David , Swalley Susanne E. , Garg Sakshi , Singh Shantanu , Carpenter Anne E.
Published: January 2022

Identifying genetic and chemical perturbations with similar impacts on cell morphology can reveal compounds’ mechanisms of action or novel regulators of genetic pathways. Research on methods for identifying such similarities has lagged due to a lack of carefully designed and well-annotated image sets of cells treated with chemical and genetic perturbations. Here, we create such a Resource dataset, CPJUMP1, where each perturbed gene is a known target of at least two chemical compounds in the dataset. We systematically explore the directionality of correlations among perturbations that target the same gene, and we find that identifying matches between chemical perturbations and genetic perturbations is a challenging task. Our dataset and baseline analyses provide a benchmark for evaluating methods that measure perturbation similarities and impact, and more generally, learn effective representations of cellular state from microscopy images. Such advancements would accelerate the applications of image-based profiling, such as functional genomics and drug discovery.
Structure-based identification of naphthoquinones and derivatives as novel inhibitors of main protease Mpro and papain-like protease PLpro of SARS-CoV-2
Authors: Santos Lucianna H. , Kronenberger Thales , Almeida Renata G. , Silva Elany B. , Rocha Rafael E. O. , Oliveira Joyce C. , Barreto Luiza V. , Skinner Danielle , Fajtová Pavla , Giardini Miriam A. , Woodworth Brendon , Bardine Conner , Lourenço André Luiz , Craik Charles S. , Poso Antti , Podust Larissa M. , McKerrow James H. , Siqueira-Neto Jair L. , O’Donoghue Anthony J. , da Silva Júnior Eufrânio N. , Ferreira Rafaela S.
Published: January 2022

The worldwide COVID-19 pandemic caused by the coronavirus SARS-CoV-2 urgently demands novel direct antiviral treatments. The main protease (Mpro) and papain-like protease (PLpro) are attractive drug targets among coronaviruses due to their essential role in processing the polyproteins translated from the viral RNA. In the present work, we virtually screened 688 naphthoquinoidal compounds and derivatives against Mpro of SARS-CoV-2. Twenty-four derivatives were selected and evaluated in biochemical assays against Mpro using a novel fluorogenic substrate. In parallel, these compounds were also assayed with SARS-CoV-2 PLpro. Four compounds inhibited Mpro with half-maximal inhibitory concentration (IC50) values between 0.41 µM and 66 µM. In addition, eight compounds inhibited PLpro with IC50 ranging from 1.7 µM to 46 µM. Molecular dynamics simulations suggest stable binding modes for Mpro inhibitors with frequent interactions with residues in the S1 and S2 pockets of the active site. For two PLpro inhibitors, interactions occur in the S3 and S4 pockets. In summary, our structure-based computational and biochemical approach identified novel naphthoquinonal scaffolds that can be further explored as SARS-CoV-2 antivirals.
Biochemical characterization of actin assembly mechanisms with ALS-associated profilin variants
Authors: Liu Xinbei , Pimm Morgan L. , Haarer Brian , Brawner Andrew T. , Henty-Ridilla Jessica L.
Published: January 2022

Eight separate mutations in the actin-binding protein profilin-1 have been identified as a rare cause of amyotrophic lateral sclerosis (ALS). Profilin is essential for many neuronal cell processes through its regulation of lipids, nuclear signals, and cytoskeletal dynamics, including actin filament assembly. Direct interactions between profilin and actin monomers inhibit actin filament polymerization. In contrast, profilin can also stimulate polymerization by simultaneously binding actin monomers and proline-rich tracts found in other proteins. Whether the ALS-associated mutations in profilin compromise these actin assembly functions is unclear. We performed a quantitative biochemical comparison of the direct and formin-mediated impact for the eight ALS-associated profilin variants on actin assembly using classic protein-binding and single-filament microscopy assays. We determined that the binding constant of each profilin for actin monomers generally correlates with the actin nucleation strength associated with each ALS-related profilin. In the presence of formin, the A20T, R136W, Q139L, and C71G variants failed to activate the elongation phase of actin assembly. This diverse range of formin-activities is not fully explained through profilin-PLP interactions, as all ALS-associated variants bind a formin-derived PLP peptide with similar affinities. However, chemical denaturation experiments suggest that the folding stability of these profilins impact some of these effects on actin assembly. Thus, changes in profilin protein stability and alterations in actin filament polymerization may both contribute to the profilin-mediated actin disruptions in ALS.
A western representative of an eastern clade: phylogeographic history of the gypsophilous plant Nepeta hispanica
Authors: Ramos-Gutiérrez Ignacio , Moreno-Saiz Juan Carlos , Fernández-Mazuecos Mario
Published: January 2022

The preference of certain plant species for gypsum soils leads to disjunct population structures that are thought to generate island-like dynamics potentially influencing biogeographic patterns at multiple evolutionary scales. Here, we study the evolutionary and biogeographic history of Nepeta hispanica, a western Mediterranean plant associated with gypsum soils and displaying a patchy distribution with populations very distant from each other. Three approaches were used: (a) interspecific phylogenetic analyses based on nuclear DNA sequences of the ITS region to unveil the relationships and times of divergence between N. hispanica and its closest relatives; ((b)phylogeographic analyses using plastid DNA regions trnS-trnG and psbJ-petA to evaluate the degree of genetic isolation between populations of N. hispanica, their relationships and their genetic diversity; and (c) ecological niche modelling to evaluate historical distributional changes. Results reveal that N. hispanica belongs to an eastern Mediterranean and Asian clade diversified in arid environments since the Miocene-Pliocene. This species represents the only extant lineage of this clade that colonized the western Mediterranean, probably through the northern Mediterranean coast (southern Europe). Present Iberian populations display a high plastid genetic diversity and, even if geographically distant from each other, they are highly connected according to the distribution of plastid haplotypes and lineages. This can be explained by a scenario involving a complex history of back-and-forth colonisation events, facilitated by a relative stability of suitable conditions for the species across the Iberian Peninsula throughout the Quaternary.
The role of collegiality in academic review, promotion, and tenure
Authors: Dawson Diane , Morales Esteban , McKiernan Erin C. , Schimanski Lesley A. , Niles Meredith T. , Alperin Juan Pablo
Published: January 2022

Review, promotion, and tenure (RPT) processes at universities typically assess candidates along three dimensions: research, teaching, and service. In recent years, some have argued for the inclusion of a controversial fourth criterion: collegiality. While collegiality plays a role in the morale and effectiveness of academic departments, it is amorphic and difficult to assess, and could be misused to stifle dissent or enforce homogeneity. Despite this, some institutions have opted to include this additional element in their RPT documents and processes, but it is unknown the extent of this practice and how it varies across institution type and disciplinary units. This study is based on two sets of data: survey data collected as part of a project that explored the publishing decisions of faculty and how these related to perceived importance in RPT processes, and 864 RPT documents collected from 129 universities from the United States and Canada. We analysed these RPT documents to determine the degree to which collegiality and related terms are mentioned, if they are defined, and if and how they may be assessed during the RPT process. Results show that when collegiality and related terms appear in these documents they are most often just briefly mentioned. It is less common for collegiality and related terms to be defined or assessed in RPT documents. Although the terms are mentioned across all types of institutions, there is a statistically significant difference in how prevalent they are at each. Collegiality is more commonly mentioned in the documents of doctoral research-focused universities (60%), than of master’s universities and colleges (31%) or baccalaureate colleges (15%). Results from the accompanying survey of faculty also support this finding: individuals from R-Types were more likely to perceive collegiality to be a factor in their RPT processes. We conclude that collegiality likely plays an important role in RPT processes, whether it is explicitly acknowledged in policies and guidelines or not, and point to several strategies in how it might be best incorporated in the assessment of academic careers.
Integrin signaling is critical for myeloid-mediated support of T-cell acute lymphoblastic leukemia
Authors: Lyu Aram , Nam Seo Hee , Humphrey Ryan S. , Durham Tyler A. , Hu Zicheng , Arasappan Dhivya , Horton Terzah M. , Ehrlich Lauren I. R.
Published: January 2022

We previously found that T-cell acute lymphoblastic leukemia (T-ALL) requires support from tumor-associated myeloid cells, which activate IGF1R signaling in the leukemic blasts. However, IGF1 is not sufficient to sustain T-ALL survival in vitro, implicating additional myeloid-mediated signals in T-ALL progression. Here, we find that T-ALL cells require close contact with myeloid cells to survive. Transcriptional profiling and in vitro assays demonstrate that integrin-mediated cell adhesion and activation of the downstream FAK/PYK2 kinases are required for myeloid-mediated support of T-ALL cells and promote IGF1R activation. Consistent with these findings, inhibition of integrins or FAK/PYK2 signaling diminishes leukemia burden in multiple organs and confers a survival advantage in a mouse model of T-ALL. Inhibiting integrin-mediated cell adhesion or FAK/PYK2 also diminishes survival of primary patient T-ALL cells co-cultured with myeloid cells. Furthermore, elevated integrin pathway gene signatures correlate significantly with myeloid enrichment and an inferior prognosis in pediatric T-ALL patients.

Statement of significance

Although tumor-associated myeloid cells provide critical support for T-ALL, our understanding of the underlying mechanisms remains limited. This study reveals that integrin-mediated adhesion and signaling are key mechanisms by which myeloid cells promote survival and progression of T-ALL blasts in the leukemic microenvironment.
Molecular Mechanism of Hyperactivation Conferred by a Truncated TRPA1 Disease Mutant Suggests New Gating Insights
Authors: Bali Avnika , Schaefer Samantha P. , Trier Isabelle , Zhang Alice L. , Kabeche Lilian , Paulsen Candice E.
Published: January 2022

The wasabi receptor, TRPA1, is a non-selective homotetrameric cation channel expressed in primary sensory neurons of the pain pathway, where it is activated by diverse chemical irritants. A direct role for TRPA1 in human health has been highlighted by the discovery of genetic variants associated with severe pain disorders. One such TRPA1 mutant was identified in a father-son pair with cramp fasciculation syndrome (CFS) and neuronal hyperexcitability-hypersensitivity symptoms that may be caused by aberrant channel activity, though the mechanism of action for this mutant is unknown. Here, we show the CFS-associated R919* TRPA1 mutant is functionally inactive when expressed alone in heterologous cells, which is not surprising since it lacks the 201 C-terminal amino acids that house critical channel gating machinery including the pore-lining transmembrane helix. Interestingly, the R919* mutant confers enhanced agonist sensitivity when co-expressed with wild type (WT) TRPA1. This channel hyperactivation mechanism is conserved in distant TRPA1 species orthologues and can be recapitulated in the capsaicin receptor, TRPV1. Using a combination of ratiometric calcium imaging, immunostaining, surface biotinylation, pulldown assays, fluorescence size exclusion chromatography, and proximity biotinylation assays, we show that the R919* mutant co-assembles with WT subunits into heteromeric channels. Within these heteromers, we postulate that R919* TRPA1 subunits contribute to hyperactivation by lowering energetic barriers to channel activation contributed by the missing regions. Additionally, we show heteromer activation can originate from the R919* TRPA1 subunits, which suggests an unexpected role for the ankyrin repeat and coiled coil domains in concerted channel gating. Our results demonstrate the R919* TRPA1 mutant confers gain-of-function thereby expanding the physiological impact of nonsense mutations, reveals a novel and genetically tractable mechanism for selective channel sensitization that may be broadly applicable to other receptors, and uncovers new gating insights that may explain the molecular mechanism of temperature sensing by some TRPA1 orthologues.
Machine learning guided batched design of a bacterial Ribosome Binding Site
Authors: Zhang Mengyan , Holowko Maciej Bartosz , Zumpe Huw Hayman , Ong Cheng Soon
Published: January 2022

Optimisation of gene expression levels is an essential part of the organism design process. Fine control of this process can be achieved through engineering transcription and translation control elements, including the ribosome binding site (RBS). Unfortunately, design of specific genetic parts can still be challenging due to lack of reliable design methods. To address this problem, we have created a machine learning guided Design-Build-Test-Learn (DBTL) cycle for the experimental design of bacterial RBSs to show how small genetic parts can be reliably designed using relatively small, high-quality data sets. We used Gaussian Process Regression for the Learn phase of cycle and the Upper Confidence Bound multi-armed bandit algorithm for the Design of genetic variants to be tested in vivo. We have integrated these machine learning algorithms with laboratory automation and high-throughput processes for reliable data generation. Notably, by Testing a total of 450 RBS variants in four DBTL cycles, we experimentally validated RBSs with high translation initiation rates equalling or exceeding our benchmark RBS by up to 34%. Overall, our results show that machine learning is a powerful tool for designing RBSs, and they pave the way towards more complicated genetic devices.
CRISPR-LRS for mapping transgenes in the mouse genome
Authors: Bryant W. Bart , Yang Allison , Griffin Susan , Zhang Wei , Long Xiaochun , Miano Joseph M.
Published: January 2022

Microinjected transgenes, including bacterial artificial chromosomes (BACs), insert randomly in the mouse genome. Traditional methods of mapping a transgene are challenging, thus complicating breeding strategies and the accurate interpretation of phenotypes, particularly when a transgene disrupts critical coding or noncoding sequences. Here, we introduce CRISPR-Cas9 long-read sequencing (CRISPR-LRS) to ascertain transgene integration locus and estimated copy number. This method revealed integration loci for both a BAC and Cre-driver line, and estimated the copy numbers for two other BAC mouse lines. CRISPR-LRS offers an easy approach to establish robust breeding practices and accurate phenotyping of most any transgenic mouse line.
Unravelling microalgal-bacterial interactions in aquatic ecosystems through 16S rRNA gene-based co-occurrence networks
Authors: Pushpakumara B.L.D. Uthpala , Tandon Kshitij , Willis Anusuya , Verbruggen Heroen
Published: January 2022

Interactions between microalgae and bacteria can directly influence the global biogeochemical cycles but the majority of such interactions remain unknown. 16S rRNA gene-based co-occurrence networks have potential to help identify microalgal-bacterial interactions. Here, we used data from 10 Earth microbiome projects to identify potential microalgal-bacterial associations in aquatic ecosystems. A high degree of clustering was observed in microalgal-bacterial modules, indicating densely connected neighbourhoods.ProteobacteriaandBacteroidetespredominantly co-occurred with microalgae and represented hubs of most modules. Our results also indicated that species-specificity may be a global characteristic of microalgal associated microbiomes. Several previously known associations were recovered from our network modules, validating that biologically meaningful results can be inferred using this approach. A range of previously unknown associations were recognised such as co-occurrences ofBacillariophytawith unculturedPlanctomycetes OM190andDeltaproteobacteriaorderNB1-j. PlanctomycetesandVerrucomicrobiawere identified as key associates of microalgae due to their frequent co-occurrences with several microalgal taxa. Despite no clear taxonomic pattern, bacterial associates appeared functionally similar across different environments. To summarise, we demonstrated the potential of 16S rRNA gene-based co-occurrence networks as a hypothesis-generating framework to guide more focused research on microalgal-bacterial associations.
Function and constraint in enhancers with multiple evolutionary origins
Authors: Fong Sarah L. , Capra John A.
Published: January 2022

Motivation

Thousands of human gene regulatory enhancers are composed of sequences with multiple evolutionary origins. These evolutionarily “complex” enhancers consist of older “core” sequences and younger “derived” sequences. However, the functional relationship between the sequences of different evolutionary origins within complex enhancers is poorly understood.

Results

We evaluated the function, selective pressures, and sequence variation across core and derived components of human complex enhancers. We find that both components are older than expected from the genomic background, and cores are enriched for derived sequences of similar evolutionary ages. Both components show strong evidence of biochemical activity in massively parallel report assays (MPRAs). However, core and derived sequences have distinct transcription factor (TF) binding preferences that are largely stable across evolutionary origins. Given these signatures of function, both core and derived sequences have substantial evidence of purifying selection. Nonetheless, derived sequences exhibit weaker purifying selection than adjacent cores. Derived sequences also tolerate more common genetic variation and are enriched compared to cores for eQTL associated with gene expression variability in human populations.

Conclusions

Both core and derived sequences have strong evidence of gene regulatory function, but derived sequences have distinct constraint profiles, TF binding preferences, and tolerance to variation compared with cores. We propose that the step-wise integration of younger derived and older core sequences has generated regulatory substrates with robust activity and the potential for functional variation. Our analyses demonstrate that synthesizing study of enhancer evolution and function can aid interpretation of regulatory sequence activity and functional variation across human populations.

Significance Statement

Thousands of human gene regulatory enhancers are mosaics of sequences from multiple evolutionary origins, yet how these different segments contribute to enhancer function is poorly understood. By dissecting their regulatory functions, transcription factor binding, constraint, and human genetic variation, we show that both older “core” and younger “derived” sequences in complex enhancers have strong evidence of gene regulatory function, but derived sequences are more likely to harbor genetic variants that influence function. Together, our results support a model in which the integration of sequences of different origins generates regulatory substrates with robust activity and the potential for functional variation.
An integrated lab-on-a-chip device for RNA extraction, amplification and CRISPR-Cas12a-assisted detection for COVID-19 screening in resource-limited settings
Authors: Ngamsom Bongkot , Iles Alexander , Kamita Moses , Kimani Racheal , Rodriguez-Mateos Pablo , Mungai Mary , Dyer Charlotte E. , Walter Cheryl , Gitaka Jesse , Pamme Nicole
Published: January 2022

In response to the ongoing COVID-19 pandemic and disparities of vaccination coverage in low- and middle-income countries, it is vital to adopt a widespread testing and screening programme, combined with contact tracing, to monitor and effectively control the infection dispersion in areas where medical resources are limited. This work presents a lab-on-a-chip platform, namely “IFAST-CRISPR”, as an affordable, rapid and high-precision molecular diagnostic means for SARS-CoV-2 detection. The herein proposed “sample-to-answer” platform integrates RNA extraction, amplification and CRISPR-Cas-based detection with lateral flow readout in one device. The microscale dimensions of the device containing immiscible liquids, coupled with the use of silica paramagnetic beads and GuHCl, streamline sample preparation, including RNA concentration, extraction and purification, in 15 min with minimal hands-on steps. By combining RT-LAMP with CRISPR-Cas12 assays targeting the nucleoprotein (N) gene, visual identification of ≥ 470 copies mL-1 genomic SARS-CoV-2 samples was achieved in 45 min, with no cross-reactivity towards HCoV-OC43 nor H1N1. On-chip assays showed the ability to isolate and detect SARS-CoV-2 from 1,000 genome copies mL-1 of replication-deficient viral particles in 1 h. This simple, affordable and integrated platform demonstrated a visual, faster, and yet specificity and sensitivity-comparable alternative to the costly gold-standard RT-PCR assay, requiring only a simple heating source. Further investigations on multiplexing and direct interfacing of the accessible Swan-brand cigarette filter for saliva sample collection could provide a complete work flow for COVID-19 diagnostics from saliva samples suitable for low-resource settings.