bluuebunny/crossref_metadata_2025 huggingface dataset:
Minimizing Reference Bias with an Impute-First Approach
Authors: Vaddadi Kavya , Mun Taher , Langmead Ben
Published: December 2023

Pangenome indexes reduce reference bias in sequencing data analysis. However, bias can be reduced further by using a personalized reference, e.g. a diploid human reference constructed to match a donor individual’s alleles. We present a novel impute-first alignment framework that combines elements of genotype imputation and pangenome alignment. It begins by genotyping the individual using only a subsample of the input reads. It next uses a reference panel and efficient imputation algorithm to impute a personalized diploid reference. Finally, it indexes the personalized reference and applies a read aligner, which could be a linear or graph aligner, to align the full read set to the personalized reference. This framework achieves higher variant-calling recall (99.54% vs. 99.37%), precision (99.36% vs. 99.18%), and F1 (99.45% vs. 99.28%) compared to a graph pangenome aligner. The personalized reference is also smaller and faster to query compared to a pangenome index, making it an overall advantageous choice for whole-genome DNA sequencing experiments.
Human naïve stem cell models reveal the role of FGF in hypoblast specification in the human embryo
Authors: Dattani Anish , Corujo-Simon Elena , Radley Arthur , Heydari Tiam , Taheriabkenar Yasaman , Carlisle Francesca , Lin Simeng , Mill Jonathan , Zandstra Peter , Nichols Jennifer , Guo Ge
Published: December 2023

The hypoblast is an essential extra-embryonic tissue set aside within the inner cell mass early in mammalian embryo development, in the blastocyst. Research with human embryos is challenging. Thus, stem cell models that reproduce hypoblast differentiation provide valuable alternatives. We show here that human naïve PSC to hypoblast differentiation proceeds via reversion to a transitional ICM-like state, from which hypoblast emerges in concordance with the trajectory in human blastocysts. We identified a window when fibroblast growth factor (FGF) signalling is critical for hypoblast specification. Revisiting FGF signalling in human embryos revealed that inhibition in the early blastocyst suppresses hypoblast formation.In vitro, the induction of hypoblast is synergistically enhanced by limiting trophectoderm and epiblast fates. This finding revises previous reports and establishes a conservation in lineage specification between mouse and human. Overall, this study demonstrates the utility of human naïve PSC-based models in elucidating mechanistic features of early human embryogenesis.
Plasmacytoid dendritic cells (pDCs) display surface but not membrane-bound IgE across a broad range of total serum IgE levels
Authors: Chen Jonathan H. , MacGlashan Donald W. , McCormack Amanda , Schroeder John T. , Tversky Jody R.
Published: December 2023

Introduction

Plasmacytoid dendritic cells (pDCs) are unique antigen presenting cells that may be implicated in allergic disease because they bind IgE on their surface and modulate important Th1/Th2 cytokine responses. While conducting in vitro experiments using excess omalizumab to capture IgE, we discovered that this immunoglobulin is not readily removed from pDC to the same extent observed for basophils suggesting that a portion of the IgE on pDC is membrane bound.

Methods

Basophils and pDC were prepared from leukopacks using established protocols. In order to isolate PBMCs, blood was also drawn from consenting donors with a wide range in total serum IgE levels. B cells and pDCs were identified by flow cytometry by gating on CD19/CD27 and BDCA2/CD123, respectively. Quilizumab, a mouse anti-human monoclonal IgG1 antibody with specificity only for membrane-bound IgE, was used to detect this molecule in both cell types.

Results

When used in vitro at 1.5 mg/ml, omalizumab removed ∼80-90% of the IgE expressed by basophils. In contrast, IgE expression decreased only 30-40% on pDC treated likewise. Upon analyzing pDC for membrane-bound IgE, there was no significant difference in number of target-bound cells and mean fluorescence between the mouse isotype control and quilizumab among pDCs (P= 0.125, 0.165). However, the ratio of the proportion of target-bound CD19+27+B cells compared to other cells was 32:1 for isotype (P< 0.001) and 54:1 for quilizumab (P= 0.015).

Conclusion

Overall, this study demonstrates that while pDCs express significant levels of FcεRI that binds IgE to the surface, there is no appreciable amount of membrane-bound IgE noted. The reason for omalizumab’s poor ability to remove IgE from pDCs remains unknown.
PROTEOME-SCALE RECOMBINANT STANDARDS AND A ROBUST HIGH-SPEED SEARCH ENGINE TO ADVANCE CROSS-LINKING MS-BASED INTERACTOMICS
Authors: Clasen Milan Avila , Ruwolt Max , Kurt Louise U. , Gozzo Fabio C , Wang Shuai , Chen Tao , Carvalho Paulo C , Lima Diogo Borges , Liu Fan
Published: December 2023

Abstract

Advancing data analysis tools for proteome-wide cross-linking mass spectrometry (XL-MS) requires ground-truth standards that mimic biological complexity. Here, we develop wellcontrolled XL-MS standards comprising hundreds of recombinant proteins that are systematically mixed for cross-linking. We use one standard dataset to guide the development of Scout, a search engine for XL-MS with MS-cleavable cross-linkers. Using other, independent datasets from our standards as well as published datasets, we benchmark the performance of Scout and existing XL-MS software. This demonstrates that Scout offers the best combination of speed, sensitivity, and false-discovery rate control. These results illustrate how our large recombinant standards can support the development of XL-MS analysis tools and evaluation of XL-MS results.
Unraveling Unbreakable Hairpins: Characterizing RNA secondary structures that are persistent after dinucleotide shuffling
Authors: Pratt Alyssa A , Hendrix David A
Published: December 2023

The sequence of nucleotides that make up an RNA determines its structure, which determines its function. The RNA hairpin, also known as a stem-loop, is a ubiquitous and fundamental feature of RNA secondary structure. A common method of randomizing an RNA sequence is dinucleotide shuffling with the Altschul-Erickson algorithm, which preserves the dinucleotide content of the sequence. This algorithm generates randomized sequences by sampling Eulerian paths through the de Bruijn graph representation of the original sequence. We identified a subset of RNA hairpins in the bpRNA-1m meta-database that always form hairpins after repeated application of dinucleotide shuffling. We investigated these “unbreakable hairpins” and found several common properties. First, we found that unbreakable hairpins had on average similar folding energies compared to other hairpins of similar lengths, although they frequently contained ultra-stable hairpin loops. We found that they tend to be split by purines and pyrimidines on opposite strands of the stem. Furthermore, we found that this specific sequence feature restricts the number of distinct Eulerian paths through their de Bruijn graph representation, resulting in a small number of distinguishable dinucleotide-shuffled sequences. Beyond this algorithmic means of identification, these distinct sequences may have biological significance because we found that a significant percentage occur in a specific location of 16S ribosomal RNAs. Finally, we present a formula to calculate the number of possible unique dinucleotide shuffled sequences for an input RNA sequence, which has utility for the general application of the Altschul-Erickson algorithm.
Complex ecosystems lose stability when resource consumption is out of niche
Authors: Liu Yizhou , Hu Jiliang , Lee Hyunseok , Gore Jeff
Published: December 2023

Natural communities exhibit diverse dynamics, encompassing global stability, multi-stability, periodic oscillations, and chaotic fluctuations in species abundances. Resource-consumer interactions provide a lens for mechanistic understanding of community behaviors, yet only a globally stable equilibrium can exist in the original MacArthur resource-consumer model. Here we find that diverse dynamics emerges when species consume resources that contribute little to their own growth. Key to understanding these results is comparison of niche range (difference between growth-promoting resources of similar species) and consumption range (difference between growth-promoting resources and the resources that are actually consumed). If the consumption range is small, we observe global stability as in the MacArthur model. But when consumption range increases to about the niche range, stability is lost, giving rise to emergent alternative stable states, globally stable states but with species extinction, and slightly later, persistent fluctuations. Given its importance in dictating stability, we define the ratio between consumption and niche ranges as encroachment, and find that it predicts key community properties like diversity and attractor basins even after the instability transition. In particular, after the loss of stability (encroachment greater than one), species extinction increases approximately linearly with encroachment. Since encroachment relies only on intrinsic species properties, community stability is resilient to environmental changes such as resource supply and mortality rate. Encroachment provides a framework for capturing resource competition due to growth-consumption inconsistency in complex communities as well as a robust quantitative characterization of the resulting emergent dynamics.
CTCF-mediated 3D chromatin predetermines the gene expression program in the male germline
Authors: Kitamura Yuka , Takahashi Kazuki , Maezawa So , Munakata Yasuhisa , Sakashita Akihiko , Kaplan Noam , Namekawa Satoshi H.
Published: December 2023

Spermatogenesis is a unidirectional differentiation process that generates haploid sperm, but how the gene expression program that directs this process is established is largely unknown. Here we determine the high-resolution 3D chromatin architecture of male germ cells during spermatogenesis and show that CTCF-mediated 3D chromatin predetermines the gene expression program required for spermatogenesis. In undifferentiated spermatogonia, CTCF-mediated chromatin contacts on autosomes pre-establish meiosis-specific super-enhancers (SE). These meiotic SE recruit the master transcription factor A-MYB in meiotic spermatocytes, which strengthens their 3D contacts and instructs a burst of meiotic gene expression. We also find that at the mitosis-to-meiosis transition, the germline-specific Polycomb protein SCML2 resolves chromatin loops that are specific to mitotic spermatogonia. Moreover, SCML2 and A-MYB establish the unique 3D chromatin organization of sex chromosomes during meiotic sex chromosome inactivation. We propose that CTCF-mediated 3D chromatin organization enforces epigenetic priming that directs unidirectional differentiation, thereby determining the cellular identity of the male germline.
Preference and Perception of Indian Caregivers Towards Formal Long–Term Supportive Services – A Mixed–Methods Study
Authors: Ghosh Subharati , Mathur Arvind
Published: December 2023

Purpose of the Study

In India, the rate of population aging, the greater burden of chronic disease-related care, and smaller families question the sustainability of traditional family based, home long term support and services (LTSS). However, little is still known about Indians’ perception of formal LTSS.

Design and Methods

A mixed-methods design of family caregivers for older adults from Jodhpur, Rajasthan (n=30 in-depth interviews; n=100 quantitative survey). Inductive qualitative data analysis identified emergent themes about perceptions of either informal or formal LTSS. Caregivers self-reported which common LTSS they needed in a quantitative survey. Multivariable Poisson and logistic regression models were used to estimate the average total number of LTSS and probabilities of self reporting wanting individual LTSS, respectively.

Results

The central theme was a negative perception of formal LTSS, especially the idea of paid helpers. A second theme served as the rationale for the first theme: caregivers reported a “duty” to provide care to one’s family that “others” and those “doing it for money” could not meet. Caregivers reported on average 2.8 LTSS needs of 10 options. Formal LTSS, like home-health care assistance with instrumental activities of daily living, were least frequently reported; caregiver education and self-care activities were the most reported.

Implications

Despite providing intensive amounts of informal LTSS and care for their family members, Indian caregivers consistently reported disinterest in using formal LTSS alternatives in qualitative and quantitative data. Caregivers reported a stronger desire for services that support their ability to carry out their caregiving roles.
Leveraging DNA Methylation to Predict Treatment Response in Major Depressive Disorder: A Critical Review
Authors: Dahrendorff Jan , Currier Glenn , Uddin Monica
Published: December 2023

Background

Major depressive disorder (MDD) is a debilitating and prevalent mental disorder with a high disease burden. Despite a wide array of different treatment options, many patients do not respond to initial treatment attempts. Selection of the most appropriate treatment remains a significant clinical challenge in psychiatry, highlighting the need for the development of biomarkers with predictive utility. Recently, the epigenetic modification DNA methylation (DNAm) has emerged to be of great interest as a potential predictor of MDD treatment outcomes. Here we review efforts to date that seek to identify DNAm signatures associated with treatment response in individuals with MDD.

Methods

Searches were conducted in the databases PubMed, Scopus, and Web of Science with the concepts and keywords major depressive disorder, DNA methylation, antidepressants, psychotherapy, cognitive behavior therapy, electroconvulsive therapy, transcranial magnetic stimulation, and brain stimulation therapies.

Results

We identified 31 studies implicating DNAm patterns associated with MDD treatment outcomes. The majority of studies (N=25) are focused on selected target genes exploring treatment outcomes in pharmacological treatments (N=22) with a few studies assessing treatment response to electroconvulsive therapy (N=3). Additionally, there are few genome-scale efforts (N=6) to characterize DNAm patterns associated with treatment outcomes.

Limitations

There is a relative dearth of studies investigating DNAm patterns in relation to psychotherapy, electroconvulsive therapy, or transcranial magnetic stimulation; importantly, most existing studies have limited sample sizes.

Conclusions

Given the heterogeneity in both methods and results of studies to date, there is a need for additional studies before existing findings can inform clinical decisions.
Donor-Specific Antibody Testing is an Effective Surveillance Strategy for High-Risk Antibody Mediated Rejection in Heart Transplant Patients in the Contemporary Era
Authors: Cusi Vincenzo , Cardenas Ashley , Tada Yuko , Vaida Florin , Wettersten Nicholas , Chak Jennifer , Pretorius Victor , Urey Marcus Anthony , Morris Gerald P. , Lin Grace , Kim Paul J.
Published: December 2023

Background:

Pathologic antibody mediated rejection (pAMR) evaluation and donor specific antibody (DSA) testing are recommended in the first year after heart transplantation (HTx) in adult patients. Whether DSA testing adds prognostic information to contemporary pAMR surveillance has not been fully studied.

Methods:

This was a single center study of consecutive endomyocardial biopsies (EMB) performed between November 2010 and February 2023 in adult HTx patients. The primary objective was to evaluate whether DSA testing contributes additional information to pAMR surveillance to better predict overall survival. Secondary endpoints included cardiac allograft dysfunction and loss.

Results:

A total of 6,033 EMBs from 544 HTx patients were reviewed for the study. The pAMR+/DSA+ group had significantly lower overall survival versus the pAMR-/DSA-group (hazard ratio [HR] = 2.63; 95% confidence interval [CI], 1.35-5.11; pc= 0.013). In the pAMR+/DSA+ group, patients with cardiac allograft dysfunction, compared to those without allograft dysfunction, had significantly lower overall and cardiac survival (pc< 0.001 for both). In contrast, pAMR+/DSA+ and pAMR-/DSA-patients without cardiac allograft dysfunction showed no difference in overall and cardiac survival. Primary graft dysfunction (PGD) was a novel risk factor for development ofde novoDSAs (dnDSA) three weeks post-HTx (p = 0.007).

Conclusions:

DSA testing as the primary surveillance method can identify high-risk pAMR+/DSA+ patients. Surveillance pAMR testing in the contemporary era may need to be reevaluated. Earlier DSA testing at 10-14 days post-HTx should be considered in PGD patients.
Multiscale Bayesian Simulations Reveal Functional Chromatin Condensation of Gene Loci
Authors: Brandani Giovanni B , Gu Chenyang , Gopi Soundhararajan , Takada Shoji
Published: December 2023

Chromatin, the complex assembly of DNA and associated proteins, plays a pivotal role in orchestrating various genomic functions. To aid our understanding of the principles underlying chromatin organization, we introduce Hi-C metainference, a Bayesian approach that integrates Hi-C contact frequencies into multiscale prior models of chromatin. This approach combines both bottom-up (the physics-based prior) and top-down (the data-driven posterior) strategies to characterize the 3D organization of a target genomic locus. We first demonstrate the capability of this method to accurately reconstruct the structural ensemble and the dynamics of a system from contact information. We then apply the approach to investigate the Sox2, Pou5f1, and Nanog loci of mouse embryonic stem cells using a bottom-up chromatin model at 1kb resolution. We observe that the studied loci are conformationally heterogeneous and organized as crumpled globules, favoring contacts between distant enhancers and promoters. Using nucleosome-resolution simulations, we then reveal how the Nanog gene is functionally organized across the multiple scales of chromatin. At the local level, diverse chromatin folding motifs correlate with epigenetics, with open chromatin predominantly observed at cis-regulatory elements and compact tetranucleosomes in between. At the larger scale, we find that enhancer-promoter contacts are driven by the transient condensation of chromatin into compact domains stabilized by extensive inter-nucleosome interactions. Overall, this work highlights the condensed, but dynamic nature of chromatinin vivo, contributing to a deeper understanding of gene structure-function relationships.
Immediate impact of the 2021 harmful algal bloom in southeast Hokkaido on the rocky intertidal community and its spatial variation
Authors: Yao Yuan , Noda Takashi
Published: December 2023

There has been a limited number of studies on the effects of harmful algal blooms (HABs) on natural rocky intertidal ecosystems. From mid-September to early November 2021, an unprecedented HAB caused byKarenia selliformishit the Pacific coast of southeast Hokkaido, Japan, for the first time, causing massive mortalities among marine organisms. To clarify the immediate impacts of the HAB on abundance of 10 rocky intertidal species in four functional groups (macroalgae, sessile invertebrates, molluscan grazers, and molluscan carnivores), we focused on two questions. 1) How did the HAB affect the abundance of each species differently at the regional scale? 2) How did the impacts of the HAB on the abundance of each functional groups vary spatially, and was the spatial variation of the HAB impacts related to the spatial distribution of cell density of HAB species? To study these issues, we compared census data for 17 years before the HAB and within one month after it for five shores on the southeast coast of Hokkaido. Results showed that two macroalgae species and all three molluscan grazer species declined significantly after the HAB. Moreover, the decrease of molluscan grazers was significantly correlated with cell density. These results suggest that the impacts of the HAB in southeast Hokkaido on abundance of rocky intertidal organisms are highly variable depending on species and locality, presumably because of differences in species-specific tolerances to HAB toxins and spatial variation in the density of the HAB organisms.
MetageNN: a memory-efficient neural network taxonomic classifier robust to sequencing errors and missing genomes
Authors: da Silva Rafael Peres , Suphavilai Chayaporn , Nagarajan Niranjan
Published: December 2023

Background

With the rapid increase in throughput of long-read sequencing technologies, recent studies have explored their potential for taxonomic classification by using alignment-based approaches to reduce the impact of higher sequencing error rates. While alignment-based methods are generally slower, k-mer-based taxonomic classifiers can overcome this limitation, potentially at the expense of lower sensitivity for strains and species that are not in the database.

Results

We present MetageNN, a memory-efficient long-read taxonomic classifier that is robust to sequencing errors and missing genomes. MetageNN is a neural network model that uses short k-mer profiles of sequences to reduce the impact of distribution shifts on error-prone long reads. Benchmarking MetageNN against other machine learning approaches for taxonomic classification (GeNet) showed substantial improvements with long-read data (20% improvement in F1 score). By utilizing nanopore sequencing data, MetageNN exhibits improved sensitivity in situations where the reference database is incomplete. It surpasses the alignment-based MetaMaps and MEGAN-LR, as well as the k-mer-based Kraken2 tools, with improvements of 100%, 36%, and 23% respectively at the read-level analysis. Notably, at the community level, MetageNN consistently demonstrated higher sensitivities than the previously mentioned tools. Furthermore, MetageNN requires < 1/4thof the database storage used by Kraken2, MEGAN-LR and MMseqs2 and is >7x faster than MetaMaps and GeNet and >2x faster than MEGAN-LR and MMseqs2.

Conclusion

This proof of concept work demonstrates the utility of machine-learning-based methods for taxonomic classification using long reads. MetageNN can be used on sequences not classified by conventional methods and offers an alternative approach for memory-efficient classifiers that can be optimized further.
TRIB3 silencing promotes the downregulation of Akt pathway and PAX3-FOXO1 in high-risk rhabdomyosarcoma
Authors: Gallo-Oller Gabriel , Pons Guillem , Sansa-Girona Julia , Navarro Natalia , Zarzosa Patricia , García-Gilabert Lia , Cabré Fernandez Paula , Guillén Burrieza Gabriela , Valero-Arrese Lorena , Segura Miguel F. , Lizcano José M. , Sánchez de Toledo José , Moreno Lucas , Gallego Soledad , Roma Josep
Published: December 2023

Background

Rhabdomyosarcoma (RMS), such as other childhood tumors, has witnessed treatment advancements in recent years. However, high-risk patients continue to face poor survival rates, often attributed to the presence of the PAX3/7-FOXO1 fusion proteins, which has been associated with metastasis and treatment resistance. Despite efforts to directly target these chimeric proteins, clinical success remains elusive. In this study, the main aim was to address this challenge by investigating regulators of FOXO1. Specifically, we focused on TRIB3, a potential regulator of the fusion protein in RMS.

Methods

TRIB3 expression was examined through the analysis of patient datasets, including gene expression profiling and gene set enrichment analyses. In cell lines, the DepMap dataset for RMS was utilized alongside Western blot analysis to assess TRIB3 expression. The functional significance of TRIB3 in RMS was assessed through constitutive and inducible shRNA-mediated knockdowns. Subsequentin vitroandin vivoanalyses, including orthotopic tumor models in immune-compromised mice, were conducted to delineate the role and underlying molecular mechanisms exerted by TRIB3 in RMS

Results

Our findings revealed a prominent TRIB3 expression in RMS tumors, highlighting its correlation with several clinical features. By conducting TRIB3 genetic inhibition experiments, we observed an impairment on cell proliferation. Notably, the knockdown of TRIB3 led to a decrease in PAX3-FOXO1 and its target genes at protein level, accompanied by a reduction in the activity of the Akt signaling pathway. Furthermore, TRIB3 influenced posttranslational modifications, such as phosphorylation together with proteasomal degradation of PAX3-FOXO1 protein. Additionally, inducible silencing of TRIB3 significantly delayed tumor growth and improved overall survivalin vivo.

Conclusions

Based on our comprehensive analysis, we propose that TRIB3 holds therapeutic potential for treating the most aggressive subtype of RMS. The findings herein reported contribute to our understanding of the underlying molecular mechanisms driving RMS progression and provide novel insights into the potential use of TRIB3 as a therapeutic intervention for high-risk RMS patients.
Comparative analysis of methodologies for detecting extrachromosomal circular DNA
Authors: Gao Xuyuan , Liu Ke , Luo Songwen , Tang Meifang , Liu Nianping , Jiang Chen , Fang Jingwen , Li Shouzhen , Hou Yanbing , Guo Chuang , Qu Kun
Published: December 2023

Extrachromosomal circular DNA (eccDNA) is crucial in oncogene amplification, gene transcription regulation, and intratumor heterogeneity. While various analysis pipelines and experimental methods have been developed for eccDNA identification, their detection efficiencies have not been systematically assessed. To address this, we evaluated the performance of 7 analysis pipelines using three simulated datasets, in terms of accuracy, similarity, duplication rate, and computational resource consumption. We also compared the eccDNA detection efficiency of 7 experimental methods through twenty-one real sequencing datasets. Our results identified Circle-Map and CReSIL as the most effective pipelines for eccDNA detection through short-read and long-read sequencing data, respectively. Moreover, third-generation sequencing-based Circle-Seq showed superior efficiency in detecting copy number-amplified eccDNA over 10 kb in length. These results offer valuable insights for researchers in choosing the most suitable methods for eccDNA research.
Early Pastoralism in Central European Forests: Insights from Ancient Environmental Genomics
Authors: Zampirolo Giulia , Holman Luke E. , Sawafuji Rikai , Ptáková Michaela , Kovačiková Lenka , Šída Petr , Pokorný Petr , Pedersen Mikkel Winther , Walls Matthew
Published: December 2023

Central European forests have been shaped by complex human interactions throughout the Holocene, with significant changes following the introduction of domesticated animals in the Neolithic (∼7.5 – 6.0 kyr BP). However, understanding early pastoral practices and their impact on forests is limited by methods for detecting animal movement across past landscapes. Here we examine ancient sedimentary DNA (sedaDNA) preserved at the Velký Mamuťák rock shelter, in northern Bohemia (Czech Republic), which has been a forested enclave since the early Holocene. We find that domesticated animals, their associated microbiomes, and plants potentially gathered for fodder, have clear representation by the Late Neolithic, around 6.0 kyr BP, and persist throughout the Bronze Age into recent times. We identify a change in dominant grazing species from sheep to pigs in the Bronze Age (∼4.1 – 3.0 kyr BP) and interpret the impact this had in the mid-Holocene retrogressions that still define the structure of Central European forests today. This study highlights the ability of ancient metagenomics to bridge archaeological and paleoecological methods and provide an enhanced perspective on the roots of the Anthropocene.
Mitochondrial Apolipoprotein MIC26 is a metabolic rheostat regulating central cellular fuel pathways
Authors: Lubeck Melissa , Naha Ritam , Schaumkessel Yulia , Westhoff Philipp , Stefanski Anja , Petzsch Patrick , Stühler Kai , Köhrer Karl , Weber Andreas P. M. , Anand Ruchika , Reichert Andreas S. , Kondadi Arun Kumar
Published: December 2023

Mitochondria play central roles in metabolism and metabolic disorders such as type 2 diabetes. MIC26, a MICOS complex subunit, was linked to diabetes and modulation of lipid metabolism. Yet, the functional role of MIC26 in regulating metabolism under hyperglycemia is not understood. We employed a multi-omics approach combined with functional assays using WT andMIC26KO cells cultured in normoglycemia or hyperglycemia, mimicking altered nutrient availability. We show that MIC26 has an inhibitory role in glycolysis and cholesterol/lipid metabolism under normoglycemic conditions. Under hyperglycemia, this inhibitory role is reversed demonstrating that MIC26 is critical for metabolic adaptations. This is partially mediated by alterations of mitochondrial metabolite transporters. Furthermore,MIC26deletion led to a major metabolic rewiring of glutamine utilization as well as oxidative phosphorylation. We propose that MIC26 acts as a metabolic ‘rheostat’, that modulates mitochondrial metabolite exchange via regulating mitochondrial cristae, allowing cells to cope with nutrient overload.
Antigenic and genetic diversity of H1 and H3 influenza A viruses in swine in Brazil
Authors: Lopes Sara , Anderson Tavis K. , Schaefer Rejane , Tochetto Caroline , Gava Danielle , Cantao Mauricio E. , Ciacci-Zanella Janice R. , Vincent Baker Amy L. , Lewis Nicola S.
Published: December 2023

Background

Influenza A virus (IAV) circulates within human and swine populations, and pigs are considered intermediate hosts for the generation of IAV with pandemic potential. Surveillance and characterization of IAVs circulating in pig populations are crucial to strain match vaccines to control IAV transmission in pigs and quantify pandemic potential to humans.

Methods

Here, we characterized the genetic and antigenic diversity of IAVs circulating in Brazilian swine between 2010-2018. Phylogenetic maximum-likelihood trees were generated for 84 Brazilian hemagglutinin (HA) gene segments. Hemagglutination inhibition (HI) assay data was used with antigenic cartography to quantify the antigenic differences among representative H1 and H3 swine viruses and relative cross-reactivity between these viruses and human seasonal vaccine strains.

Results

We identified two genetic lineages of H1 viruses derived from separate human-to-swine transmission events (H1 1B lineage, clades 1B.2.3 and 1B.2.4), an H3 lineage that has diversified into two genetic clades (H3 1990.5.1 and 1990.5.2), and HA genes associated with the 2009 H1N1 pandemic. There was limited cross-reactivity between circulating swine lineages and significant antigenic variation within lineage.

Conclusions

The antigenic diversity among endemic IAV in swine indicates a need for regional strain-specific vaccination strategies in Brazil. Our data supports the need for systematic genomic surveillance and characterization in Brazil to improve the efficacy of swine vaccines and quantify the pandemic potential of endemic swine influenza A viruses.
Mechanical positive feedback and biochemical negative feedback combine to generate complex contractile oscillations in cytokinesis
Authors: Werner Michael E. , Ray Dylan D. , Breen Coleman , Staddon Michael F. , Jug Florian , Banerjee Shiladitya , Maddox Amy Shaub
Published: December 2023

Contractile force generation by the cortical actomyosin cytoskeleton is essential for a multitude of biological processes. The actomyosin cortex behaves as an active material that drives local and large-scale shape changes via cytoskeletal remodeling in response to biochemical cues and feedback loops. Cytokinesis is the essential cell division event during which a cortical actomyosin ring generates contractile force to change cell shape and separate two daughter cells. Our recent work with active gel theory predicts that actomyosin systems under the control of a biochemical oscillator and experiencing mechanical strain will exhibit complex spatiotemporal behavior, but cytokinetic contractility was thought to be kinetically simple. To test whether active materialsin vivoexhibit spatiotemporally complex kinetics, we used 4-dimensional imaging with unprecedented temporal resolution and discovered sections of the cytokinetic cortex undergo periodic phases of acceleration and deceleration. Quantification of ingression speed oscillations revealed wide ranges of oscillation period and amplitude. In the cytokinetic ring, activity of the master regulator RhoA pulsed with a timescale of approximately 20 seconds, shorter than that reported for any other biological context. Contractility oscillated with 20-second periodicity and with much longer periods. A combination ofin vivoandin silicoapproaches to modify mechanical feedback revealed that the period of contractile oscillation is prolonged as a function of the intensity of mechanical feedback. Effective local ring ingression is characterized by slower speed oscillations, likely due to increased local stresses and therefore mechanical feedback. Fast ingression also occurs where material turnover is high,in vivoandin silico. We propose that downstream of initiation by pulsed RhoA activity, mechanical positive feedback, including but not limited to material advection, extends the timescale of contractility beyond that of biochemical input and therefore makes it robust to fluctuations in activation. Circumferential propagation of contractility likely allows sustained contractility despite cytoskeletal remodeling necessary to recover from compaction. Our work demonstrates that while biochemical feedback loops afford systems responsiveness and robustness, mechanical feedback must also be considered to describe and understand the behaviors of active materialsin vivo.
Artificial intelligence-assisted stimulated Raman molecular cytology enables accurate and rapid intraoperative assessment of the surgical margin in pancreatic ductal adenocarcinoma
Authors: Fang Tinghe , Liu Daoning , Chen Xun , Zhou Keji , Hao Chunyi , Yue Shuhua
Published: December 2023

Pancreatic ductal adenocarcinoma (PDAC) is a highly mortal cancer with surgical resection the only potentially curative treatment. The accurate intraoperative assessment of PDAC surgical margins is significant in guaranteeing resection adequacy and improving the patient’s survival. However, the commonly used frozen-section pancreatic biopsy is limited by its strict requirement of practitioners and lack of objectivity. Here, we developed the multi-instance cytology with learned Raman embedding (MICLEAR), a label-free cytology based on chemical information reflected by Raman spectra. First, 4085 cells collected from 41 patients were imaged with stimulated Raman scattering (SRS) microscopy. Then, a contrastive learning (CL)-based cell embedding model was obtained to represent each cell with a concise vector that contained its morphological and componential information. Finally, a multi-instance learning (MIL)-based diagnosis model using cell vectors predicted the probability that the margin was positive. MICLEAR reached 80% sensitivity, 94.1% specificity, and 0.86 AUC on the pancreatic neck margin (PNM) samples from 27 patients. It holds promise for rapid and accurate assessment of PDAC surgical margins.
Towards ultra-low-cost smartphone microscopy
Authors: Zhang Haoran , Zhang Weiyi , Zuo Zirui , Yang Jianlong
Published: December 2023

The outbreak of COVID-19 exposed the inadequacy of our technical tools for home health surveillance, and recent studies have shown the potential of smartphones as a universal optical microscopic imaging platform for such applications. However, most of them use laboratory-grade optomechanical components and transmitted illuminations to ensure focus tuning capability and imaging quality, which keeps the cost of the equipment high. Here we propose an ultra-low-cost solution for smartphone microscopy. To realize focus tunability, we designed a seesaw-like structure capable of converting large displacements on one side into small displacements on the other (reduced to ∼9.1%), which leverages the intrinsic flexibility of 3D printing materials. We achieved a focus-tuning accuracy of ∼ 5µm, which is 40 times higher than the machining accuracy of the 3D-printed lens holder itself. For microscopic imaging, we use an off-the-shelf smartphone camera lens as the objective and the built-in flashlight as the illumination. To compensate for the resulting image quality degradation, we developed a learning-based image enhancement method. We use the CycleGAN architecture to establish the mapping from smartphone microscope images to benchtop microscope images without pairing. We verified the imaging performance on different biomedical samples. Except for the smartphone, we kept the full costs of the device under 4 USD. We think these efforts to lower the costs of smartphone microscopes will benefit their applications in various scenarios, such as point-of-care testing, on-site diagnosis, and home health surveillance.

Research highlights

We propose a solution for ultra-low-cost smartphone microscopy. Utilizing the flexibility of 3D-printed material, we can achieve focusing accuracy of ∼ 5µm. Such a low-cost device will benefit point-of-care diagnosis and home health surveillance.
In situlipidomics ofStaphylococcus aureusosteomyelitis using imaging mass spectrometry
Authors: Good Christopher J. , Butrico Casey E. , Colley Madeline E. , Gibson-Corley Katherine N. , Cassat James E. , Spraggins Jeffrey M. , Caprioli Richard M.
Published: December 2023

Osteomyelitis occurs whenStaphylococcus aureusinvades the bone microenvironment, resulting in a bone marrow abscess with a spatially defined architecture of cells and biomolecules. Imaging mass spectrometry and microscopy are invaluable tools that can be employed to interrogate the lipidome ofS. aureus-infected murine femurs to reveal metabolic and signaling consequences of infection. Here, nearly 250 lipids were spatially mapped to healthy and infection-associated morphological features throughout the femur, establishing composition profiles for tissue types. Ether lipids and arachidonoyl lipids were significantly altered between cells and tissue structures in abscesses, suggesting their roles in abscess formation and inflammatory signaling. Sterols, triglycerides, bis(monoacylglycero)phosphates, and gangliosides possessed ring-like distributions throughout the abscess, indicating dysregulated lipid metabolism in a subpopulation of leukocytes that cannot be discerned with traditional microscopy. These data provide chemical insight into the signaling function and metabolism of cells in the fibrotic border of abscesses, likely characteristic of lipid-laden macrophages.
Brain mineralization in postoperative delirium and cognitive decline
Authors: Lammers-Lietz Florian , Borchers Friedrich , Feinkohl Insa , Hetzer Stefan , Kanar Cicek , Lachmann Gunnar , Chien Claudia , Spies Claudia , Winterer Georg , Zaborszky Laszlo , Zacharias Norman , Paul Friedemann
Published: December 2023

Delirium is a severe postoperative complication associated with poor overall and especially neurocognitive prognosis. Altered brain mineralization is found in neurodegenerative disorders but has not been studied in postoperative delirium and postoperative cognitive decline. We hypothesized that mineralization-related hypointensity in susceptibility-weighted magnetic resonance imaging (SWI) is associated with postoperative delirium and cognitive decline.

We analyzed a subsample of cognitively healthy patients ≥65 years presenting for elective major surgery who underwent SWI before (N=65) and three months after surgery (N=33) as part of a subproject in the BioCog study. We measured relative SWI intensities in basal ganglia, hippocampus, and posterior basal forebrain cholinergic system (pBFCS). A post-hoc analysis of two pBFCS subregions (Ch4, Ch4p) was conducted. Patients were screened for delirium until the seventh postoperative day. Cognitive testing was performed before and three months after surgery.

Preoperative relative SWI hypointensities in the basal ganglia and pBFCS were associated with increased risk for postoperative delirium after adjustment for surgery duration. After additional adjustment for age, sex, preoperative MMSE and region volume, only the association of pBFCS hypointensity and postoperative delirium remained significant. Adjusted for surgery duration, perioperative change in relative SWI intensities of the pBFCS was associated with cognitive decline three months after surgery. This association remained at a trend level after adjustments for age, sex, and region volume, but a significant independent association especially with pBFCS-subregion Ch4p was found in a post-hoc analysis.

Brain mineralization, particularly in the cerebral cholinergic system, could be a pathomechanism in postoperative delirium and cognitive decline.
The small molecule inhibitor 3PO is a modulator of neutrophil metabolism, ROS production and NET release
Authors: Alarcon Michele Fresneda , Abdullah Genna Ali , Nolan Andy , Linford Christina , Meschis Maria Martina , Cross Andrew L , Sellin Andrew , Phelan Marie M , Wright Helen L
Published: December 2023

1.

Neutrophils are key effector leukocytes of the innate immune system and play a pivotal role in defending the host against microbial infections. Recent studies have identified a crucial link between glycolysis and neutrophil cellular functions. Using human neutrophils, we have investigated the intricate relationship between glycolysis, extracellular glucose availability, and the enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), in the regulation of reactive oxygen species (ROS) and neutrophil extracellular trap (NET) production. We have identified that PFKFB3 is elevated in rheumatoid arthritis (RA) neutrophils and that the small molecule PFKFB3 inhibitor 3PO is a key regulator of neutrophil ROS and NET production. 3PO blocked the production of ROS and NETs in a dose-dependent manner in both RA and healthy (HC) neutrophils (p<0.01), and RA neutrophils were more sensitive to lower concentrations of 3PO. Bacterial killing was only partially inhibited by 3PO, and the proportion of live neutrophils after 24h incubation was unchanged. Using NMR metabolomics, we identified that 3PO increases the concentration of lactate, phenylalanine and L-glutamine in neutrophils, as well as significantly decreasing intracellular glutathione (adj. p-value<0.05). We also demonstrated that RA neutrophils produce ROS and NETs in culture conditions which mimic the low glucose environments encountered in RA synovial joints. Our results also suggest 3PO may have molecular targets beyond PFKFB3. By dissecting the intricate interplay between metabolism and neutrophil effector functions, this study advances the understanding of the molecular mechanisms governing pro-inflammatory neutrophil responses and identifies 3PO as a potential therapeutic for conditions characterized by dysregulated neutrophil activation.
Specific cPRC1 complexes are co-opted to mediate oncogenic gene repression in diffuse midline glioma
Authors: Gannon Dáire , Lagan Eimear , Silva Ademar Jesus , Bibawi Peter , Doherty Anthony M. , Nimmo Darragh , McCole Rachel , Monger Craig , Genesi Giovani , Vanderlinden Dibekeme Aurelie , Innes James A. , Yang Lu , Chen Bryan , van Mierlo Guido , Jansen Pascal W.T.C , Wynne Keiran , Sánchez-Rivera Fran-cisco J. , Soto-Feliciano Yadira M. , Vermeulen Michiel , Oliverio Giorgio , Chen Chun-Wei , Philips Richard E. , Brackenand Adrian P. , Brien Gerard L.
Published: December 2023

Diffuse midline glioma (DMG) is a fatal childhood brain tumour characterised primarily by mutant histone H3 (H3K27M). H3K27M causes a global reduction in Polycomb Repressive Complex 2 (PRC2)-mediated H3K27me3 by inhibiting PRC2 enzymatic activity. Paradoxically, PRC2 is essential in DMG tumour cells where residual complex activity is required for oncogenic gene repression, although the molecular mechanisms acting downstream of PRC2 in this context are poorly understood. Here, we’ve discovered this oncogenic gene repression is mediated by specific canonical PRC1 (cPRC1) formations. By combining CRISPR screening, biochemical and chromatin mapping approaches with functional perturbations we show that cPRC1 complexes containing CBX4 and PCGF4 drive oncogenic gene repression downstream of H3K27me3 in DMG cells. Remarkably, the altered H3K27me3 modification landscape characteristic of these tumours rewires the distribution of cPRC1 complexes on chromatin. CBX4 and PCGF4 containing cPRC1 accumulate at sites of H3K27me3 while other cPRC1 formations are displaced. Despite accounting for <5% of cPRC1 complexes in DMG, CBX4/PCGF4-containing complexes predominate as gene repressors. Our findings link the altered distribution of H3K27me3 with imbalanced cPRC1 function, promoting oncogenic gene repression in DMG cells, revealing new disease mechanisms and highlighting potential therapeutic opportunities in this incurable childhood brain tumour.
The Parkinson’s Disease related mutant VPS35 (D620N) amplifies the LRRK2 response to endolysosomal stress
Authors: McCarron Katy R. , Elcocks Hannah , Mortiboys Heather , Urbé Sylvie , Clague Michael J.
Published: December 2023

The identification of multiple genes linked to Parkinson’s Disease invites the question as to how they may cooperate. We have generated isogenic cell lines that inducibly express either wild-type or a mutant form of the retromer component VPS35 (D620N), which has been linked to Parkinson’s Disease. This has enabled us to test proposed effects of this mutation in a setting where the relative expression reflects the physiological occurrence. We confirm that this mutation compromises VPS35 association with the WASH complex, but find no defect in WASH recruitment to endosomes, nor in the distribution of lysosomal receptors, cation-independent mannose-6-phosphate receptor and Sortilin. We show VPS35 (D620N) enhances the activity of the Parkinson’s associated kinase LRRK2 towards RAB12 under basal conditions. Furthermore, VPS35 (D620N) amplifies the LRRK2 response to endolysosomal stress resulting in enhanced phosphorylation of RABs 10 and 12. By comparing different types of endolysosomal stresses such as the ionophore nigericin and the membranolytic agent LLOMe, we are able to dissociate phospho-RAB accumulation from membrane rupture.
Demonstration of Bioplastic Production from CO2and Formate using the Reductive Glycine Pathway inE. coli
Authors: Fedorova Daria , Ben-Nissan Roee , Milshtein Eliya , Jona Ghil , Dezorella Nili , Feiguelman Gil , Fedorov Rostislav , Gomaa Aya , Lindner Ariel B. , Noor Elad , Milo Ron
Published: December 2023

There is a strong need to develop technologies that reduce anthropogenic pollution and the dependence on nonrenewable Earth resources. One way of doing so is by harnessing biological systems for replacing the production of fossil-fuel based goods with low-environmental-impact alternatives. Recently, progress was made in engineering the model organismE. colito grow using CO2and formate as its only carbon and energy sources using the reductive glycine pathway (rGlyP). Here, we use this engineered strain ofE. colias a host system for the production of polyhydroxybutyrate (PHB), a biologically derived and biodegradable plastic. We confirmed the production of PHB in this strain using Nile red fluorescent microscopy, transmission electron microscopy, and GC measurements. Since formate can be efficiently generated from CO2by electrochemical reduction using renewable energy sources, this study serves as a proof of concept for the emerging field of electro-bioproduction.