bluuebunny/crossref_metadata_2025 huggingface dataset:
Sequenced-based GWAS for linear classification traits in Belgian Blue beef cattle reveals new coding variants in genes regulating body size in mammals
Authors: Gualdron Duarte JL , Yuan C , Gori AS , Moreira GCM , Takeda H , Coppieters W , Charlier C , Georges M , Druet T
Published: June 2023

Cohorts of individuals that have been genotyped and phenotyped for genomic selection programs offer the opportunity to better understand genetic variation associated with complex traits. Here, we perform an association study for traits related to body size and muscular development in intensively selected beef cattle. We leveraged multiple trait information to refine and interpret the significant associations. After a multiple-step genotype imputation to the sequence-level for 14,762 Belgian Blue beef (BBB) cattle cows, we performed a GWAS for 11 traits related to muscular development and body size. The 37 identified genome-wide significant QTL could be condensed in 11 unique QTL regions based on their position. There was evidence for pleiotropic effects in most of these regions (e.g., correlated association signals, overlap between credible sets of candidate variants – CSCV). We consequently applied a multiple-trait approach to combine information from different traits to refine the CSCV. In several QTL regions, we identified strong candidate genes known to be related to growth and height in other species such asLCORL-NCAPGorCCND2. For some of these genes, relevant candidate variants were identified in the CSCV, including three new missense variants inEZH2,PAPPA2andADAM12, possibly two additional coding variants inLCORL, and candidate regulatory variants linked toCCND2andARMC12. Strikingly, four other QTL regions were related to five (recessive) deleterious coding variants previously identified. Heterozygotes for several of these mutations have favorable effects for muscular development traits. Our study further supports that a set of common genes controls body size across mammalian species. In particular, we added new genes to the list of those associated with height in both human and cattle. We also identified new strong candidate causing variants in some of those genes, strengthening the evidence of the causality of these genes. Several breed-specific recessive deleterious variants were identified in our QTL regions, probably as a result of the extreme selection for muscular development in BBB cattle.
GIFT – an R package to access the Global Inventory of Floras and Traits
Authors: Denelle Pierre , Weigelt Patrick , Kreft Holger
Published: June 2023

Advancing knowledge of biodiversity requires open-access global databases and workflows. This appears particularly crucial for plants, as ongoing changes pose major threats to this central group of organisms. Having large-scale information on plant distributions, functional traits and evolutionary history will enable the scientific community to improve its understanding of the patterns and drivers of plant diversity on a global scale.

The Global Inventory of Floras and Traits (GIFT) is a global database of regional plant checklists that has proven successful in documenting biogeographical and geographical patterns of plants. Since the release of the first version of GIFT, the database kept on expanding. We introduce GIFT version 3.0, which contains 5,169 checklists referring to 3,400 regions. These checklists include a total of 371,148 land plant species, mostly vascular plants, of which 354,848 are accepted species names, and 109 functional traits. This new version uses new resources for taxonomic name standardization, is matched to a new plant phylogeny, comes with a new trait aggregation workflow, and includes additional environmental variables.

We also present theGIFT R-package, which contains all necessary functions to retrieve distributional, functional, phylogenetic, and environmental data from the GIFT database. The package comes with a dedicated website,https://biogeomacro.github.io/GIFT/, which includes three rich vignettes to guide users in retrieving data from GIFT.

The recent development of GIFT and its associatedR-package provide ecologists with access to one of the largest plant databases. This will foster research into regional to global patterns of plant diversity and their underlying mechanisms. Proper versioning of the database and the ability to retrieve and cite data from any previous and current instance of the GIFT database will ensure the reproducibility of studies that utilize it.
Mixed data analysis detected Endocrine Fibroblast Growth Factors (FGF19, FGF21, and FGF23) as Prognostic and Diagnostic Markers of Colorectal Neoplasia and Carcinoma
Authors: Rejali Leili , Piroozkhah Moein , Jahanbin Mana , Jalali Pooya , Khanabadi Binazir , Abkenar Elahe Daskar , Tavallaei Mehdi , Niasar Mahsa Saeedi , Hashemi Mehrdad , Sadeghi Amir , Salehi Zahra , Nazemalhosseini-Mojarad Ehsan
Published: June 2023

Background

Endocrine fibroblast growth factors (eFGFs) play important roles in various cellular signaling processes such as development and differentiation. These genes were also found to be significantly related to several cancer. However, little is known about the role of eFGFsin colon neoplasia and colon adenocarcinoma (COAD).

Methods

We performed systematically and comprehensively investigated the gene expression, DNA methylation, prognostic significance, genetic alteration, co-expressed genes, protein-protein interaction, small molecules pathway, and drug interactions ofeFGFsbased on the TIMER2.0, GEPIA2, UALCAN, OncoDB, cBioPortal, LinkedOmics, STRING, SMPDB, htfTarget, mirTarBase, circBank and DGIdb databases. Ultimately, the correlations ofeFGFsexpressions between polyp and COAD tissues compared to normal mucosa were validated using qRT-PCR as a cross-sectional part of our study.

Results

The results indicated thateFGFsare highly expressed in COAD, and abnormal gene expressions may be related to promoter methylation. In this matter, methylation analysis revealed promotor hypermethylation ofFGF19andFGF21. Conversely,FGF23was shown to have a tendency for promotor hypomethylation. Moreover, hypermethylation ofFGF21andFGF23and downregulation ofFGF23were found to be detrimental to the survival of COAD patients. KEGG pathway analyses indicated that the co-expressed genes ofeFGFfamily members were mainly related to the regulation of the actin cytoskeleton and, more notably, in Ras signaling, PI3k-Akt signaling, Rap1 signaling, and cancer pathways. Based on qRT-PCR results,FGF21was significantly overexpressed in the colon polyps compared to normal mucosa. Additionally, RNA expression ofFGF21andFGF23was markedly elevated in adenomatous polyps as opposed to hyperplastic polyps.

Conclusion

Collectively, these findings reveal the critical roles ofeFGFsin COAD tumorigenesis and suggesteFGFfamily members as promising prognostic and diagnostic markers for CRC as well as discriminating markers for high-risk from low-risk polyps.
Identification of a recombinant human astrovirus type 5 strain from an acute gastroenteritis outbreak in Beijing, China
Authors: Mao Tongyao , Zhang Peng , Li Lili , Yang Mengyao , Wang Yalu , Ma Yalin , Li Dandi , Li Jinsong , Duan Zhaojun
Published: June 2023

Two samples positive for the same strain (2103CP) of human astrovirus (HAstV) were identified during an acute gastroenteritis outbreak in Beijing, China. A full genomic analysis showed that for both samples, open reading frame 1a (ORF1a) clustered with HAstV-1, whereas ORF1b and ORF2 clustered with HAstV-5. The recombination site was detected upstream of the region where ORF1a and ORF1b overlapped. Recombinant HAstV-5 strains were reported previously in China in 2013 and 2021. Our results indicate the need for continuous surveillance of the prevalence and evolution of this recombinant HAstV-5 in China.
A role for BCL6 in maintaining CX3CR1+CD4+T cells during helminth infection
Authors: Loredan Denis G. , Devlin Joseph C. , Khanna Kamal M. , Loke P’ng
Published: June 2023

Distinct subsets of T lymphocytes express CX3CR1 under inflammatory conditions, but little is known about CX3CR1+CD4+T cells during Type 2 inflammation in helminth infections. Here, we used a fate-mapping mouse model to characterize CX3CR1+CD4+T cells during both acuteNippostrongylus brasiliensisand chronicSchistosoma mansonihelminth infections, revealing CX3CR1+CD4+T cells to be an activated tissue homing subset with varying capacity for cytokine production. Tracking these cells over time revealed that maintenance of CX3CR1 itself along with a TH2 phenotype conferred a survival advantage in the inflamed tissue. Single-cell RNA-sequencing analysis of fate-mapped CX3CR1+CD4+T cells from both the peripheral tissue and the spleen revealed a considerable level of diversity and identified a distinct population of BCL6+TCF-1+PD1+CD4+T cells in the spleen during helminth infections. Conditional deletion of BCL6 in CX3CR1+cells result in fewer CX3CR1+CD4+during infection, indicating a role in sustaining CD4+T cell responses to helminth infections. Overall, our studies revealed the behavior and heterogeneity of CX3CR1+CD4+T cells during Type 2 inflammation in helminth infections and identified BCL6 to be important in their maintenance.
Direct and indirect fitness effects of plant metabolites and genetic constraints limit evolution of allelopathy in an invading plant
Authors: Honor Richard , Marcellus Mia , Colautti Robert I.
Published: June 2023

Invading species encounter novel communities of consumers, pathogens, and competitors. Both phenotypic plasticity and rapid evolution can facilitate invasion across these heterogenous communities. However, the rate and extent of adaptive evolution on contemporary timescales can be constrained by adaptive phenotypic plasticity and the genetic architecture of traits under selection.

We measured phenotypic plasticity and quantified genetic variation for growth, leaf chlorophylla(Chla) and glucosinolates, and lifetime fitness among 23 naturally inbred seed families ofAlliaria petiolata(garlic mustard) collected across its invasive range in eastern North America. After growing a self-pollinated generation in a uniform common garden to reduce maternal effects, we reared second-generation plants in a two-year greenhouse and field experiment with naïve soil from an uninvaded habitat. We estimated selection gradients and causal factors affecting lifetime fitness when reared alone, with an intraspecific competitor, and under interspecific competition with naïveAcer saccharum(sugar maple) saplings.

We defined Total Metabolite Production (TMP) as the first principal component of Chlaand glucosinolate concentrations, accounting for 84% of variation in these two traits. TMP was significantly plastic across growing environments (p< 0.001) with limited broad-sense heritability (H2= 2.91;p= 0.08). Path analysis revealed that plastic phenotypes with higher TMP had an indirect positive effect onA. petiolatafitness via a direct, negative effect on performance ofA. saccharumcompetitors. In contrast, the second principal component defined Relative Glucosinolate Investment (RGI), which was significantly heritable (H2= 16.91, p < 0.001) with no detectable plasticity across treatments. Variation in RGI amongA. petiolatagenotypes had a direct, positive effect onA. saccharumperformance and an indirect negative effect onA. petiolatafitness.

Synthesis.Adaptive evolution of allelopathy during invasion has been constrained by (i) a lack of heritable genetic variation for allelopathy, (ii) high plasticity for TMP across competition treatments, and (iii) selection for lower RGI under interspecific and intraspecific competition. As an alternative to eco-evolutionary feedbacks, plasticity in TMP may be an overlooked explanation for variable performance ofA. petiolataacross its introduced range.
Two birds with one stone: human SIRPα nanobodies for functional modulation and in vivo imaging of myeloid cells
Authors: Wagner Teresa R. , Blaess Simone , Leske Inga B. , Frecot Desiree I. , Gramlich Marius , Traenkle Bjoern , Kaiser Philipp D. , Seyfried Dominik , Maier Sandra , Rezza Amélie , Sônego Fabiane , Thiam Kader , Pezzana Stefania , Zeck Anne , Gouttefangeas Cécile , Scholz Armin M. , Nueske Stefan , Maurer Andreas , Kneilling Manfred , Pichler Bernd J. , Sonanini Dominik , Rothbauer Ulrich
Published: June 2023

Signal-regulatory protein α (SIRPα) expressed by myeloid cells is of particular interest for therapeutic strategies targeting the interaction between SIRPα and the "don’t eat me" ligand CD47 and as a marker to monitor macrophage infiltration into tumor lesions. To address both approaches, we developed a set of novel human SIRPα (hSIRPα)-specific nanobodies (Nbs). We identified three high-affinity Nbs targeting the hSIRPα/hCD47 interface, thereby enhancing antibody-dependent cellular phagocytosis (ADCP). For non-invasivein vivoimaging, we chose S36 Nb as a non-modulating binder. By quantitative positron emission tomography (PET) in novel hSIRPα/hCD47 knock-in (KI) mice, we demonstrated the applicability of64Cu-hSIRPα-S36 Nb to visualize tumor infiltration of myeloid cells. We envision that the hSIRPα-Nbs presented in this study have potential as versatile probes, including novel myeloid-specific checkpoint inhibitors for combinatorial treatment approaches and forin vivostratification and monitoring of individual responses during cancer immunotherapies.
Data-driven identification of potentially successful intervention implementations: a proof of concept using five years of opioid prescribing data from over 7000 practices in England
Authors: Hopcroft Lisa E. M. , Curtis Helen J. , Croker Richard , Pretis Felix , Inglesby Peter , Evans Dave , Bacon Seb , Goldacre Ben , Walker Alex J. , MacKenna Brian
Published: June 2023

Background

We have previously demonstrated that opioid prescribing increased by 127% between 1998 and 2016. New policies aimed at tackling this increasing trend have been recommended by public health bodies and there is some evidence that progress is being made. We sought to extend our previous work and develop an unbiased, data-driven approach to identify general practices and clinical commissioning groups (CCGs) whose prescribing data suggest that interventions to reduce the prescribing of opioids may have been successfully implemented.

Methods

We analysed five years of prescribing data for three opioid prescribing measures: one capturing total opioid prescribing and two capturing regular prescribing of high dose opioids. Using a data-driven approach, we applied a modified version of our change detection Python library to identify changes in these measures over time, consistent with the successful implementation of an intervention. This analysis was carried out for general practices and CCGs, and organisations were ranked according to the change in prescribing rate.

Results

We present data for the three CCGs and practices demonstrating the biggest reduction in opioid prescribing across the three opioid prescribing measures. We observed a 40% drop in the regular prescribing of high dose opioids (measured as a percentage of regular opioids) in the highest ranked CCG (North Tyneside); a 99% drop in this same measure was found in several practices. Decile plots demonstrate that CCGs exhibiting large reductions in opioid prescribing do so via slow and gradual reductions over a long period of time (typically over two years); in contrast, practices exhibiting large reductions do so rapidly over a much shorter period of time.

Conclusions

By applying one of our existing analysis tools to a national dataset, we were able to rank NHS organisations by reduction in opioid prescribing rates. Highly ranked organisations are candidates for further qualitative research into intervention design and implementation.

Contributions to the literature

Demonstrating that a data-driven approach can identify and quantify changes in important clinical measures in publicly available NHS data

Identifying changes in this way allows the unbiased identification of candidates for further qualitative research into intervention design and implementation

Large reductions observed at the CCG level (which are more robust to local circumstances) demonstrate that it is possible to reduce opioid prescribing and that continued and wider success in reducing opioid prescribing is dependent, at least in part, to closing an implementation gap
Investigating of the causes of seizures in children admitted to Kerman hospitals in 2017
Authors: Ghaedamini Hossein , Nejad Biglari Habibe , Saghafi Zahra , Hosseininasab Ali , Amirbeigi Ali , Ahmadabadi Farzad , Ghaedamini Amirmahdi
Published: June 2023

Introduction

Seizure in children with the prevalence of 4 to 6 cases in 1000 is the most common neurological disorder in pediatrics. This study aimed to determine the causes of seizures in children admitted to Kerman hospitals in 2017

Materials and methods

In this retrospective study, 250 hospitalized children suffering from seizure were selected through census method. Data collection tool was a checklist. The obtained data was analyzed employing SPSS24software, using chi-square, and correlation coefficient statistical tests at the significant level P < 0.05.

Results

Out of 250 children studied, 55.6%were male and 44.4%were female. The mean age of the children was 10.54 ± 3.7 years. 80.4%were born with vaginal delivery and 19.6%were born with cesarean section. 44%of the patients had febrile seizure, 23.2%epilepsy, 12.4%infection, 10%gastroenteritis, 5.2%static encephalopathy, 2.8%meningitis, 1.6%hypocalcaemia, 0.4 %metabolic disorders and 0.4 %tumor. That underlying factors of febrile seizure were upper respiratory infection (43.6%), gastroenteritis (15.7%), shigellosis (10.5%), urinary infection (9.9%), pneumonia (8.1%), unidentified fever (7.8%), otitis (3%) and dental abscess (1.4%) respectively.

Conclusion

According to this study, the most common cause of the seizure is a febrile seizure, which confirms the results of other studies. Common seizure require diagnosis, planning, and special treatments that can be obtained by taking a complete history, accurate examination, and effective Para clinical evaluations.
Crystal structures reveal nucleotide-induced conformational changes in G motifs and distal regions in guanylate-binding protein 2
Authors: Roy Sayantan , Wang Bing , Tian Yuan , Yin Qian
Published: June 2023

Guanylate-binding proteins (GBPs) are interferon-inducible GTPases that confer protective immunity against a variety of intracellular pathogens including bacteria, viruses, and protozoan parasites. GBP2 is one of the two highly inducible GBPs, yet the precise mechanisms underlying the activation and regulation of GBP2, in particular the nucleotide-induced conformational changes in GBP2, remain poorly understood. In this study, we elucidate the structural dynamics of GBP2 upon nucleotide binding through crystallographic analysis. GBP2 dimerizes upon GTP hydrolysis and returns to monomer state once GTP is hydrolyzed to GDP. By determining the crystal structures of GBP2 G domain (GBP2GD) in complex with GDP and nucleotide-free full-length GBP2, we unveil distinct conformational states adopted by the nucleotide-binding pocket and distal regions of the protein. Our findings demonstrate that the binding of GDP induces a distinct closed conformation both in the G motifs and the distal regions in the G domain. The conformational changes in the G domain are further transmitted to the C-terminal helical domain, leading to large-scale conformational rearrangements. Through comparative analysis, we identify subtle but critical differences in the nucleotide-bound states of GBP2, providing insights into the molecular basis of its dimer-monomer transition and enzymatic activity. Overall, our study expands the understanding of the nucleotide-induced conformational changes in GBP2, shedding light on the structural dynamics governing its functional versatility. These findings pave the way for future investigations aimed at elucidating the precise molecular mechanisms underlying GBP2’s role in the immune response and may facilitate the development of targeted therapeutic strategies against intracellular pathogens.
Lipid droplets as substrates for protein phase separation
Authors: Kamatar Advika , Bravo Jack P.K. , Yuan Feng , Wang Liping , Lafer Eileen M. , Taylor David W. , Stachowiak Jeanne C. , Parekh Sapun H.
Published: June 2023

Membrane-associated protein phase separation plays critical roles in cell biology, driving essential cellular phenomena from immune signaling to membrane traffic. Importantly, by restricting diffusion to a two-dimensional surface, lipid bilayers can nucleate phase separation at far lower concentrations compared to those required for phase separation in solution. How might other intracellular lipid substrates, such as lipid droplets, contribute to nucleation of phase separation? Distinct from bilayer membranes, lipid droplets consist of a phospholipid monolayer surrounding a core of neutral lipids, and they are energy storage organelles that protect cells from lipotoxicity and oxidative stress. Here, we show that intrinsically disordered proteins can undergo phase separation on the surface of synthetic and cell-derived lipid droplets. Specifically, we find that model disordered domains, FUS LC and LAF1-RGG, separate into protein-rich and protein-depleted phases on the surfaces of lipid droplets. Owing to the hydrophobic nature of interactions between FUS LC proteins, increasing ionic strength drives an increase in its phase separation on droplet surfaces. The opposite is true for LAF1-RGG, owing to the electrostatic nature of its interprotein interactions. In both cases, protein-rich phases on the surfaces of synthetic and cell-derived lipid droplets demonstrate molecular mobility indicative of a liquid-like state. Our results show that lipid droplets can nucleate protein condensates, suggesting that protein phase separation could be key in organizing biological processes involving lipid droplets.
Short 2′-O-methyl/LNA oligomers as highly-selective inhibitors of miRNA productionin vitroandin vivo
Authors: Corradi Eloina , Koralewska Natalia , Milewski Marek C. , Masante Linda , Kierzek Ryszard , Figlerowicz Marek , Baudet Marie-Laure , Kurzynska-Kokorniak Anna
Published: June 2023

MicroRNAs (miRNAs) that share identical or near-identical sequences constitute miRNA families and are predicted to act redundantly. Yet recent evidence suggests that members of the same miRNA family with high sequence similarity might have different roles and that this functional divergence might be rooted in their precursors’ sequence. Current knock-down strategies such as antisense oligonucleotides (ASOs) or miRNA sponges cannot distinguish between identical or near identical miRNAs originating from different precursors to allow exploring unique functions of these miRNAs. We now develop a method based on short 2′-OMe/LNA-modified oligonucleotides to selectively target specific precursor molecules and ablate the production of individual members of miRNA familiesin vitroandin vivo. Using the highly conservedXenopusmiR-181a family as a proof-of-concept, we demonstrate that 2′-OMe/LNA-ASOs targeting pre-miRNA apical region elicit a precursor-selective inhibition of mature miRNA-5p production. The levels of miRNAs released from the 3′-arm of these precursors are not reduced, suggesting that our approach is also arm-selective. Overall, we show that this strategy can be successfully appliedin vivoto achieve high target selectivity to study identical or highly similar miRNAs stemming from different precursors.
Coordination of transporter, cargo, and membrane properties during non-vesicular lipid transport
Authors: Koukalova Alena , Eisenreichova Andrea , Różycki Bartosz , Boura Evzen , Humpolickova Jana
Published: June 2023

Homeostasis of cellular membranes is maintained by fine-tuning their lipid composition. Yeast lipid transporter Osh6, belonging to the oxysterol-binding protein-related proteins family, was found to participate in the transport of phosphatidylserine (PS). PS synthesized in the endoplasmic reticulum is delivered to the plasma membrane, where it is exchanged for phosphatidylinositol-4 phosphate (PI4P). PI4P provides the driving force for the directed PS transport against its concentration gradient. In this study, we employed an in vitro approach to reconstitute the transport process into the minimalistic system of large unilamellar vesicles to reveal its fundamental biophysical determinants. Our study draws a comprehensive portrait of the interplay between the structure and dynamics of Osh6, the carried cargo lipid, and the physical properties of the involved membranes, with particular attention to their charge and fluidity. Specifically, we address the role of the cargo lipid, which, by occupying the transporter, imposes changes in its dynamics and, consequently, predisposes the cargo to disembark in the correct target membrane.
Therapy-associated remodeling of pancreatic cancer revealed by single-cell spatial transcriptomics and optimal transport analysis
Authors: Shiau Carina , Cao Jingyi , Gregory Mark T. , Gong Dennis , Yin Xunqin , Cho Jae-Won , Wang Peter L. , Su Jennifer , Wang Steven , Reeves Jason W. , Kim Tae Kyung , Kim Youngmi , Guo Jimmy A. , Lester Nicole A. , Schurman Nathan , Barth Jamie L. , Weissleder Ralph , Jacks Tyler , Qadan Motaz , Hong Theodore S. , Wo Jennifer Y. , Roberts Hannah , Beechem Joseph M. , Fernandez-del Castillo Carlos , Mino-Kenudson Mari , Ting David T. , Hemberg Martin , Hwang William L.
Published: June 2023

In combination with cell intrinsic properties, interactions in the tumor microenvironment modulate therapeutic response. We leveraged high-plex single-cell spatial transcriptomics to dissect the remodeling of multicellular neighborhoods and cell–cell interactions in human pancreatic cancer associated with specific malignant subtypes and neoadjuvant chemotherapy/radiotherapy. We developed Spatially Constrained Optimal Transport Interaction Analysis (SCOTIA), an optimal transport model with a cost function that includes both spatial distance and ligand–receptor gene expression. Our results uncovered a marked change in ligand–receptor interactions between cancer-associated fibroblasts and malignant cells in response to treatment, which was supported by orthogonal datasets, including anex vivotumoroid co-culture system. Overall, this study demonstrates that characterization of the tumor microenvironment using high-plex single-cell spatial transcriptomics allows for identification of molecular interactions that may play a role in the emergence of chemoresistance and establishes a translational spatial biology paradigm that can be broadly applied to other malignancies, diseases, and treatments.
Effects of destruxin A on the trehalose content, trehalase activity and their gene expression ofBemisia tabaci
Authors: Zhang Can , Guo Jianling , Ali Shaukat , Qiu Bao-Li
Published: June 2023

Bemisia tabaci(Gennadius) (Hemiptera: Aleyrodidae) is a serious pest of agricultural crops in different regions of the world. Destruxin A (DA) is one of 39 derivatives of destruxins, which are secondary metabolites produced by entomopathogenic fungi. Due to the high insecticidal activity, DA have great application potential in biological control ofB. tabaci. In order to understand the impact of destruxin A on the energy substances of the global pestB. tabaci, we measured the changes in energy substance content of theB. tabaciadults treated with DA at LC10and LC50concentrations after 4, 8, and 12 hours. The results revealed that DA caused varying degrees of decline in trehalose, soluble sugars, and glycogen content, indicating its ability to influence the energy substance content of whiteflies. After DA treatment, the content of trehalose inB. tabacidecreased significantly, while the activity of trehalase, a specific hydrolase for trehalose, increased significantly. Further study of the expression levels of two genes (BtTre-1andBtTre-2) encoding trehalases inB. tabaciafter DA treatment showed that the soluble trehalase gene,BtTre-1, was induced in both low and high concentrations of DA, while the membrane-bound trehalase gene,BtTre-2, did not show significant changes in expression level.
Total cell N-glycosylation is altered during neuronal differentiation of iPSC to NSC and is disturbed by trisomy 21
Authors: Cindrić Ana , Vučković Frano , Murray Aoife , Klarić Thomas , Alić Ivan , Petrović Dražen Juraj , Krištić Jasminka , Nižetić Dean , Lauc Gordan
Published: June 2023

Down syndrome (DS) is a genetic condition caused by trisomy 21 (T21) and characterized by a range of neurological symptoms including intellectual disability, early onset of neurodegeneration and dementia, some of which can be attributed to accelerated aging. N-glycosylation is a protein modification that plays a critical role in numerous biological processes and its dysregulation is associated with a wide range of diseases, in some even before the onset of symptoms. N-glycosylation of total plasma proteins, as well as specific plasma proteins, such as immunoglobulin G, has been shown to change in DS, displaying an accelerated aging phenotype consistent with the various symptoms of premature aging that occur in DS. However, little is known about how T21 affects the N-glycosylation of other cellular proteins. To better understand how T21 affects N-glycosylation during neural differentiation, we characterized and compared the total released N-glycans of induced pluripotent stem cells (iPSCs) and their neural stem cell (NSC) derivatives. We analyzed six different isogenic clones all derived from a single individual with mosaic DS and thus all sharing the same genetic background; however, three had a normal disomic karyotype (D21), while the other three had an additional copy of chromosome 21 (T21). We characterized the total cell N-glycosylation profiles using ultra high performance liquid chromatography (UHPLC) and subsequent tandem mass spectrometry analysis to determine proposed glycan structures. Our results revealed both qualitative and quantitative differences in the composition of N-glycomes between iPSCs and NSCs, with NSCs showing a higher amount of complex N-glycans and a lower amount of mannosidic N-glycans when compared to iPSCs. Moreover, we found differences in N-glycosylation patterns between D21 and T21 cells. Notably, T21 cells exhibited a significant increase in the amount of pseudohybrid N-glycans. Trisomy 21 also caused a significant decrease in the abundance of a hybrid monoantennary fucosylated glycan (H6N3F1). Our findings define the released N-glycan profile of total cells for both D21 and T21 iPSCs and NSCs and suggest that the presence of a third copy of chromosome 21 impacts N-glycosylation patterns already in the stem cell state.
The interplay of DNA repair context with target sequence predictably biases Cas9-generated mutations
Authors: Pallaseni Ananth , Peets Elin Madli , Girling Gareth , Crepaldi Luca , Kuzmin Ivan , Moor Marilin , Muñoz-Subirana Núria , Schimmel Joost , Serçin Özdemirhan , Mardin Balca R. , Tijsterman Marcel , Peterson Hedi , Kosicki Michael , Parts Leopold
Published: June 2023

Mutagenic outcomes of CRISPR/Cas9-generated double-stranded breaks depend on both the sequence flanking the cut and cellular DNA damage repair. The interaction of these features has been largely unexplored, limiting our ability to understand and manipulate the outcomes. Here, we measured how the absence of 18 repair genes changed frequencies of 83,680 unique mutational outcomes generated by Cas9 double-stranded breaks at 2,838 synthetic target sequences in mouse embryonic stem cells. This large scale survey allowed us to classify the outcomes in an unbiased way, generating hypotheses about new modes of double-stranded break repair. Our data indicate a specialised role forPrkdc(DNA-PKcs protein) andPolm(Polμ) in creating 1bp insertions that match the nucleotide on the proximal side of the Cas9 cut with respect to the protospacer-adjacent motif (PAM), a variable involvement ofNbn(NBN) andPolq(Polθ) in the creation of different deletion outcomes, and a unique class of uni-directional deletion outcomes that are dependent on both end-protection geneXrcc5(Ku80) and the resection geneNbn(NBN). We used the knowledge of the reproducible variation across repair milieus to build predictive models of the mutagenic outcomes of Cas9 scission that outperform the current standards. This work improves our understanding of DNA repair gene function, and provides avenues for more precise modulation of CRISPR/Cas9-generated mutations.
Is a Picture Worth 1,000 SNPs? Effects of User-Submitted Photographs on Ancestry Estimates from Direct-to-Consumer Canine Genetic Tests
Authors: Rando Halie M. , Graim Kiley , Hampikian Greg , Greene Casey S.
Published: June 2023

Objective

To evaluate whether the breed ancestry predictions of direct-to-consumer (DTC) genetic tests for dogs are influenced by the user-provided photograph.

Animals

Twelve pet dogs considered purebred (i.e., registered with a breed organization) representing twelve different breeds.

Methods

Six buccal swabs per dog were collected by the owners and submitted to six DTC genetic testing companies. The experimenters registered each sample with the company. For half of the dogs, the registration included a photograph of the DNA donor. For the other half of the dogs, photographs were swapped between dogs. Analysis of the DNA and breed ancestry prediction was conducted by each company. Each company’s breed predictions were evaluated to assess whether the condition (i.e., matching versus shuffled photograph) affected the odds of identifying the DNA donor’s registered breed. A convolutional neural network was also used to predict breed based solely on the photograph as a positive control.

Results

Five of the six tests always produced results that included the registered breed. One test and the convolutional neural network were unlikely to identify the registered breed and frequently returned results that included the breed in the photograph. This result suggests that one test on the market is relying on the photograph more than the DNA sample. Additionally, differences in the predictions made across all tests underscore the challenge of identifying breed ancestry, even in purebred dogs.

Clinical Relevance

Veterinarians are likely to encounter patients who have conducted DTC genetic testing and may find themselves in the position of explaining genetic test results that they did not order. This systematic comparison of tests on the market provides context for interpreting unexpected results from consumer-grade DTC genetic testing kits.
Efficient Visual Perception of Human-Robot Walking Environments using Semi-Supervised Learning
Authors: Kuzmenko Dmytro , Tsepa Oleksii , Kurbis Andrew Garrett , Mihailidis Alex , Laschowski Brokoslaw
Published: June 2023

Convolutional neural networks trained using supervised learning can improve visual perception for human-robot walking. These advances have been possible due to large-scale datasets like ExoNet and StairNet - the largest open-source image datasets of real-world walking environments. However, these datasets require vast amounts of manually annotated data, the development of which is time consuming and labor intensive. Here we present a novel semi-supervised learning system (ExoNet-SSL) that uses over 1.2 million unlabelled images from ExoNet to improve training efficiency. We developed a deep learning model based on mobile vision transformers and trained the model using semi-supervised learning for image classification. Compared to standard supervised learning (98.4%), our ExoNet-SSL system was able to maintain high prediction accuracy (98.8%) when tested on previously unseen environments, while requiring 35% fewer labelled images during training. These results show that semi-supervised learning can improve training efficiency by leveraging large amounts of unlabelled data and minimize the size requirements for manually annotated images. Future research will focus on model deployment for onboard real-time inference and control of human-robot walking.
A drop dispenser for simplifying on-farm detection of foodborne pathogens
Authors: Ranjbaran Mohsen , Kaur Simerdeep , Wang Jiangshan , Raut Bibek , Verma Mohit S.
Published: June 2023

Rapid nucleic-acid biosensors are useful for on-farm detection of foodborne pathogens on fresh produce during pre-season and pre-harvest stages. Such tools aim to be user-friendly so that a producer could operate them in a few simple steps and detect multiple targets. Currently, an easy-to-use device for on-farm applications does not exist commercially. One of the bottlenecks is the delivery of a prescribed amount of sample to the reaction sites of the biosensor using a simple and precise approach. Here, we developed drop dispensers using 3D printing and a hydrophilic surface chemistry treatment to generate precise drops (DNA/bacterial samples) of a few micro-liters (∼20 to ∼33 µL). We tested multiple copies of these devices over time of repeated application to estimate their shelf-lives which is about one month. In addition to drop generation tests, we employed these devices in nucleic-acid testing. The tests used loop-mediated isothermal amplification (LAMP) to detect DNA or whole cells of Shiga-toxin-producingEscherichia coliO157:H7. These tests were performed to simulate the on-farm sample collection (using collection flags that we previously designed) and delivery using the drop dispensers. Our results showed that these devices performed similarly to standard commercial pipettors in LAMP assays, providing a limit of detection of 7.8×106cell/mL for whole-cell detection. This drop dispenser will eventually be part of a user-friendly consumable kit that will enable performing LAMP assays by non-specialist users for a cost of USD 4 per test.
Distinct Conformations of Mirabegron Determined by MicroED
Authors: Lin Jieye , Unge Johan , Gonen Tamir
Published: June 2023

Mirabegron, commonly known as “Myrbetriq”, has been widely prescribed as a medicine for overactive bladder syndrome for over a decade. However, the structure of the drug and what conformational changes it may undergo upon binding its receptor remain unknown. In this study, we employed microcrystal electron diffraction (MicroED) to reveal its elusive three-dimensional (3D) structure. We find that the drug adopts two distinct conformational states (conformers) within the asymmetric unit. Analysis of hydrogen bonding and packing demonstrated that the hydrophilic groups were embedded within the crystal lattice, resulting in a hydrophobic surface and low water solubility. Structural comparison revealed the presence oftrans- andcis-forms in conformers1and2, respectively. Comparison of the structures of Mirabegron alone with that of the drug bound to its receptor,1the beta 3 adrenergic receptor (β3AR) suggests that the drug undergoes major conformational change to fit in the receptor agonist binding site. This research highlights the efficacy of MicroED in determining the unknown and polymorphic structures of active pharmaceutical ingredients (APIs) directly from powders.
Decoding the comprehensive substrate-specificity and evidence of altered site-specific collagen prolyl-3-hydroxylation, lysyl-hydroxylation, and lysyl O-glycosylation in P4ha1 and P4ha2 deleted mutant mice
Authors: Sarohi Vivek , Basak Trayambak
Published: June 2023

Collagens, the most abundant proteins in mammals, play pivotal roles in the maintenance of tissue structure, functions, cell-to-cell communication, cellular migration, behavior, and growth. Collagens are highly complex in structure due to the dynamic post-translational modifications (PTMs) such as hydroxylations (on prolines and lysine residues) and O-glycosylation (on hydroxylysines) enzymatically catalyzed during biosynthesis. The most prevalent modification in fibrillar collagens is prolyl 4-hydroxylation catalyzed by collagen prolyl 4-hydroxylases (C-P4hs). Prolyl 4-hydroxylation on collagens plays a critical role in collagen biosynthesis, thermostability, and cell-collagen interactions. However, the site-specificity of prolyl 4-hydroxylase 1 (P4ha1) and P4ha2 is not comprehensively studied yet. Further, the effect of P4ha1 and P4ha2 on the plethora of other site-specific collagen PTMs is not known to date. In-depth mass-spectrometry data (PXD008802) analysis of mice skin collagen I extracted from wild-type and different deletion mutants of C-P4hs revealed that partial or full deletion of prolyl 4-hydroxylases (P4ha1 and P4ha2) significantly decreases collagen deposition in ECM hinting towards perturbed biosynthesis. A total of421site-specific PTMs on fibrillar collagen chains (Col1a1, Col1a2, and Col3a1) were identified. Further, novel23P4ha1 specific,8P4ha2 specific, and18C-P4hs promiscuous sites on fibrillar collagen chains were identified. Partial deletion of P4ha1 and full deletion of P4ha2 also resulted in altered levels of the site-specific prolyl-3-hydroxylation occupancy in collagen I. Surprisingly, an increased level of site-specific lysyl hydroxylation (Col1a1-K731, Col1a2-K183,315) was documented upon partial deletion of P4ha1 and full deletion of P4ha2. Our findings showcased that the activity of prolyl 4-hydroxylases is not limited to 4-hydroxylation of specific proline sites, but simultaneously can perturb the entire biosynthetic network by modulating prolyl 3-hydroxylation and lysyl hydroxylation occupancy levels in the fibrillar collagen chains in a site-specific manner.
Comprehensive profiling of the human intestinal DNA virome and prediction of disease-associated bacterial hosts in severe Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS)
Authors: Hsieh Shen-Yuan , Savva George M. , Telatin Andrea , Tiwari Sumeet K. , Tariq Mohammad A. , Newberry Fiona , Seton Katharine A. , Booth Catherine , Bansal Amolak S. , Wileman Thomas , Adriaenssens Evelien M. , Carding Simon R.
Published: June 2023

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a disabling disorder of unknown etiology with severely affected patients being house- and/or bedbound. A historical association with chronic virus infection and subsequent recent reports correlating intestinal microbial dysbiosis with disease pathology prompted us to analyze the intestinal virome in a small cohort of severely-affected ME/CFS patients and same household healthy controls (SHHC). Datasets from whole metagenomic sequencing (WMS) and sequencing of virus-like particles (VLP)-enriched metagenomes from the same fecal sample yielded diverse, high-quality vOTUs with high read coverage and high genome completeness. The core intestinal virome was largely composed of tailed phages in the classCaudoviriceteswith no significant differences in alpha diversity between ME/CFS and SHHC groups. However, the WMS dataset had a higher Shannon measure than the VLP dataset (p< 0.0001), with VLP- and WMS-derived sequences indicating differential abundances within several viral families and different viral compositions in beta diversity. This confirms that combining different isolation methodologies identifies a greater diversity of viruses including extracellular phages and integrated prophages. DNA viromes and bacteriomes from ME/CFS and SHHC groups were comparable with no differences in any alpha or beta diversity measures. One vOTU derived from the VLP-derived dataset was assigned to ssDNA human virus smacovirus 1. Using anin-silicoapproach to predict cohort-based bacterial hosts, we identified members of theAnaerotruncusgenus interacting with unique viruses present in ME/CFS microbiomes; this may contribute to the GI microbial dysbiosis described in ME/CFS patients.
Nanoparticle endocytosis is driven by monocyte phenotype rather than nanoparticle size under high shear flow conditions
Authors: Aldarondo Dasia A. , Huynh Chris , Dickey Leah , Bilynsky Colette , Lee Yerim , Wayne Elizabeth C.
Published: June 2023

Monocytes are members of the mononuclear phagocyte system involved in pathogen clearance and nanoparticle pharmacokinetics. Monocytes play a critical role in the development and progression of cardiovascular disease and, recently, in SARS-CoV-2 pathogenesis. While studies have investigated the effect of nanoparticle modulation on monocyte uptake, their capacity for nanoparticle clearance is poorly studied. In this study, we investigated the impact ofACE2deficiency, frequently observed in individuals with cardiovascular complications, on monocyte nanoparticle endocytosis. Moreover, we investigated nanoparticle uptake as a function of nanoparticle size, physiological shear stress, and monocyte phenotype. Our Design of Experiment (DOE) analysis found that the THP-1ACE2-cells showed a greater preference for 100nm particles under atherosclerotic conditions than THP-1 wild-type cells. Observing how nanoparticles can modulate monocytes in the context of disease can inform precision dosing.
The Antitumor Activities of Anti-CD47 Antibodies Require Fc-FcγR interactions
Authors: Osorio Juan C. , Smith Patrick , Knorr David A. , Ravetch Jeffrey V.
Published: June 2023

While anti-CD47 antibodies hold promise for cancer immunotherapy, early phase clinical trials have shown limited signs of clinical benefit, suggesting that blockade of CD47 alone may not be sufficient for effective tumor control. Here, we investigate the contributions of the Fc domain of anti-CD47 antibodies required for optimal in vivo antitumor activity across multiple species-matched models, providing new insights into the mechanisms underlying the efficacy of this emerging class of therapeutic antibodies. Using a novel mouse model humanized for CD47, SIRPα and FcγRs, we demonstrate that local administration of an Fc-engineered anti-CD47 antibody with enhanced binding to activating FcγRs modulates myeloid and T-cell subsets in the tumor microenvironment, resulting in improved long-term systemic antitumor immunity and minimal on-target off-tumor toxicity. Our results highlight the importance of Fc optimization in the development of effective anti-CD47 therapies and provide a novel approach for enhancing the antitumor activity of this promising immunotherapy.

Highlights

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Engagement of activating FcγRs augments the in vivo antitumor activity of CD47 blocking antibodies

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Humanized mice for CD47, SIRPα and FcγRs allow assessment of hFcγRs contribution to the activity of anti-hCD47 Abs

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Fc-optimized anti-hCD47 ab promotes systemic antitumor immunity with abscopal effect and minimal on-target toxicity

Graphical Abstract