bluuebunny/crossref_metadata_2025 huggingface dataset:
Timelapse viability assay to detect division and death of primary multiple myeloma cells in response to drug treatments with single cell resolution
Authors: Mark Christina , Callander Natalie S. , Chng Kenny , Miyamoto Shigeki , Warrick Jay
Published: November 2021

Heterogeneity among cancer cells and in the tumor microenvironment (TME) is thought to be a significant contributor to the heterogeneity of clinical therapy response observed between patients and can evolve over time. A primary example of this is multiple myeloma (MM), a generally incurable cancer where such heterogeneity contributes to the persistent evolution of drug resistance. However, there is a paucity of functional assays for studying this heterogeneity in patient samples or for assessing the influence of the patient TME on therapy response. Indeed, the population-averaged data provided by traditional drug response assays and the large number of cells required for screening remain significant hurdles to advancement. To address these hurdles, we developed a suite of accessible technologies for quantifying functional drug response to a panel of therapies inex vivothree-dimensional culture using small quantities of a patient’s own cancer and TME components. This suite includes tools for label-free single-cell identification and quantification of both cell division and death events with a standard brightfield microscope, an open-source software package for objective image analysis and feasible data management of multi-day timelapse experiments, and a new approach to fluorescent detection of cell death that is compatible with long-term imaging of primary cells. These new tools and capabilities are used to enable sensitive, objective, functional characterization of primary MM cell therapy response in the presence of TME components, laying the foundation for future studies and efforts to enable predictive assessment drug efficacy for individual patients.

Insight Box

The new tools and capabilities described here allow new insights into functional primary cell therapy response by (i) enabling more drugs to be tested on precious and limited numbers of patient cellsex vivo(ii) providing the ability to discriminate both cell division and death events over multiple days with single-cell resolution, and (iii) by incorporating the influences of a patient’s own cancer cells and TME components on drug response.
A quantitative synthesis of soil microbial effects on plant species coexistence
Authors: Yan Xinyi , Levine Jonathan M. , Kandlikar Gaurav S.
Published: November 2021

Soil microorganisms play a major role in shaping plant diversity, not only through their direct effects as pathogens, mutualists, and decomposers, but also by altering interactions between plants. In particular, previous research has shown that the soil community often generates frequency-dependent feedback loops among plants that can either destabilize species interactions, or generate stabilizing niche differences that promote species coexistence. However, recent insights from modern coexistence theory have shown that microbial effects on plant coexistence depend not only on these stabilizing or destabilizing effects, but also on the degree to which they generate competitive fitness differences. While many previous experiments have generated the data necessary for evaluating microbially mediated fitness differences, these effects have rarely been quantified in the literature. Here we present a meta-analysis of data from 50 studies, which we used to quantify the microbially mediated (de)stabilization and fitness differences derived from a classic plant-soil feedback model. Across 518 pairwise comparisons, we found that soil microbes generated both stabilization (or destabilization) and fitness differences, but also that the microbially mediated fitness differences dominated. As a consequence, if plants are otherwise equivalent competitors, the balance of soil microbe-generated (de)stabilization and fitness differences drives species exclusion much more frequently than coexistence or priority effects. Our work shows that microbially mediated fitness differences are an important but overlooked effect of soil microbes on plant coexistence. This finding paves the way for a more complete understanding of the processes that maintain plant biodiversity.
Site-Specific Synthesis of N4-Acetylcytidine in RNA Reveals Physiological Duplex Stabilization
Authors: Bartee David , Nance Kellie D. , Meier Jordan L.
Published: November 2021

N4-acetylcytidine (ac4C) is a post-transcriptional modification of RNA that is conserved across all domains of life. All characterized sites of ac4C in eukaryotic RNA occur in the central nucleotide of a 5’-CCG-3’ consensus sequence. However, the thermodynamic consequences of cytidine acetylation in this context have never been assessed due to its challenging synthesis. Here we report the synthesis and biophysical characterization of ac4C in its endogenous eukaryotic sequence context. First, we develop a synthetic route to homogenous RNAs containing electrophilic acetyl groups. Next, we use thermal denaturation to interrogate the effects of ac4C on duplex stability and mismatch discrimination in a native sequence found in human ribosomal RNA. Finally, we demonstrate the ability of this chemistry to incorporate ac4C into the complex modification landscape of human tRNA, and use duplex melting combined with sequence analysis to highlight a potentially unique enforcing role for ac4C in this setting. By enabling the analysis of nucleic acid acetylation in its physiological sequence context, these studies establish a chemical foundation for understanding the function of a universally-conserved nucleobase in biology and disease.
Human NOP2/NSUN1 Regulates Ribosome Biogenesis Through Non-Catalytic Complex Formation with Box C/D snoRNPs
Authors: Liao Han , Gaur Anushri , McConie Hunter , Shekar Amirtha , Wang Karen , Chang Jeffrey T. , Breton Ghislain , Denicourt Catherine
Published: November 2021

5-Methylcytosine (m5C) is a base modification broadly found on various RNAs in the human transcriptome. In eukaryotes, m5C is catalyzed by enzymes of the NSUN family composed of seven human members (NSUN1-7). NOP2/NSUN1 has been primarily characterized in budding yeast as an essential ribosome biogenesis factor required for the deposition of m5C on the 25S ribosomal RNA (rRNA). Although human NOP2/NSUN1 has been known to be an oncogene overexpressed in several types of cancer, its functions and substrates remain poorly characterized. Here we used a miCLIP-seq approach to identify human NOP2/NSUN1 RNA substrates. Our analysis revealed that NOP2/NSUN1 catalyzes the deposition of m5C at position 4447 on the 28S rRNA. We also find that NOP2/NSUN1 binds to the 5’ETS region of the pre-rRNA transcript and regulates pre-rRNA processing through non-catalytic complex formation with box C/D snoRNAs. We provide evidence that NOP2/NSUN1 facilitates the recruitment of U3 and U8 snoRNAs to pre-90S ribosomal particles and their stable assembly into snoRNP complexes. Remarkably, expression of both WT and catalytically inactive NOP2/NSUN1 in knockdown background rescues the rRNA processing defects and the stable assembly of box C/D snoRNP complexes, suggesting that NOP2/NSUN1-mediated deposition of m5C on rRNA is not required for ribosome synthesis.
Stage-specific disruption of X chromosome expression during spermatogenesis in sterile house mouse hybrids
Authors: Larson Erica L. , Kopania Emily E. K. , Hunnicutt Kelsie E. , Vanderpool Dan , Keeble Sara , Good Jeffrey M.
Published: November 2021

Hybrid sterility is a complex phenotype that can result from the breakdown of spermatogenesis at multiple developmental stages. Here, we disentangle two proposed hybrid male sterility mechanisms in the house mice,Mus musculus domesticusandM. m. musculus, by comparing patterns of gene expression in sterile F1 hybrids from a reciprocal cross. We found that hybrid males from both cross directions showed disrupted X chromosome expression during prophase of meiosis I consistent with a loss of Meiotic Sex Chromosome Inactivation (MSCI) andPrdm9-associated sterility, but that the degree of disruption was greater in mice with anM. m. musculusX chromosome consistent with previous studies. During postmeiotic development, gene expression on the X chromosome was only disrupted in one cross direction, suggesting that misexpression at this later stage was genotype-specific and not a simple downstream consequence of MSCI disruption which was observed in both reciprocal crosses. Instead, disrupted postmeiotic expression may depend on the magnitude of earlier disrupted MSCI, or the disruption of particular X-linked genes or gene networks. Alternatively, only hybrids with a potential deficit ofSlycopies, a Y-linked ampliconic gene family, showed overexpression in postmeiotic cells, consistent with a previously proposed model of antagonistic coevolution between the X and Y-linked ampliconic genes contributing to disrupted expression late in spermatogenesis. The relative contributions of these two regulatory mechanisms and their impact on sterility phenotypes awaits further study. Our results further support the hypothesis that X-linked hybrid sterility in house mice has a variable genetic basis, and that genotype-specific disruption of gene regulation contributes to overexpression of the X chromosome at different stages of development. Overall, these findings underscore the critical role of epigenetic regulation of the X chromosome during spermatogenesis and suggest that these processes are prone to disruption in hybrids.
Persistent parental RNAi in the beetleTribolium castaneuminvolves maternal transmission of long double-stranded RNA
Authors: Horn Thorsten , Narov Kalin D. , Panfilio Kristen A.
Published: November 2021

Parental RNA interference (pRNAi) is a powerful and widely used method for gene-specific knockdown. Yet in insects its efficacy varies between species, and how the systemic RNAi response is transmitted from mother to offspring remains elusive. Using the flour beetleTribolium castaneum, we report an RT-qPCR strategy to unmask the presence of double-stranded RNA (dsRNA) distinct from endogenous mRNA. We find that the injected dsRNA is directly transmitted into the egg and persists throughout embryogenesis. Despite this depletion of dsRNA from the mother, we show that strong pRNAi can persist for months before waning at strain-specific rates. In seeking the receptor proteins for cellular uptake of long dsRNA into the egg, we lastly present a phylogenomics profiling approach to ascertain macroevolutionary distributions of candidate proteins. We demonstrate a visualization strategy based on taxonomically hierarchical assessment of orthology clustering data to rapidly assess gene age and copy number changes, refined by several lines of sequence-based evidence. We use this approach to document repeated losses of SID-1-like channel proteins in the arthropods, including wholesale loss in the Heteroptera (true bugs), which are nonetheless highly sensitive to pRNAi. Overall, we elucidate practical considerations for insect pRNAi against a backdrop of outstanding questions on the molecular mechanism of dsRNA transmission to achieve long-term, systemic knockdown.
Homology mediated end joining enables efficient non-viral targeted integration of large DNA templates in primary human T cells
Authors: Webber Beau R. , Johnson Matthew J. , Slipek Nicholas J. , Lahr Walker S. , DeFeo Anthony P. , Skeate Joseph G. , Qiu Xiaohong , Rathmann Blaine , Diers Miechaleen D. , Wick Bryce , Henley Tom , Choudhry Modassir , Scott McIvor R. , Moriarity Branden S.
Published: November 2021

Adoptive cellular therapy using genetically engineered immune cells holds tremendous promise for the treatment of advanced cancers. While the number of available receptors targeting tumor specific antigens continues to grow, the current reliance on viral vectors for clinical production of engineered immune cells remains a significant bottleneck limiting translation of promising new therapies. Here, we describe an optimized methodology for efficient CRISPR-Cas9 based, non-viral engineering of primary human T cells that overcomes key limitations of previous approaches. By synergizing temporal optimization of reagent delivery, reagent composition, and integration mechanism, we achieve targeted integration of large DNA cargo at efficiencies nearing those of viral vector platforms with minimal toxicity. CAR-T cells generated using our approach are highly functional and elicit potent anti-tumor cytotoxicity in vitro and in vivo. Importantly, our method is readily adaptable to cGMP compliant manufacturing and clinical scale-up, offering a near-term alternative to the use of viral vectors for production of genetically engineered T cells for cancer immunotherapy.
FoxP3 associates with enhancer-promoter loops to regulate Treg-specific gene expression
Authors: Ramirez Ricardo N. , Chowdhary Kaitavjeet , Leon Juliette , Mathis Diane , Benoist Christophe
Published: November 2021

Gene expression programs are specified by higher-order chromatin structure and enhancer-promoter loops (EPL). T regulatory cells (Treg) identity is dominantly specified by the transcription factor FoxP3, whose mechanism of action is unclear. We applied proximity-ligation with chromatin immunoprecipitation (HiChIP) in Treg and closely related conventional CD4+ T cells (Tconv). EPL identified by H3K27Ac HiChIP showed a range of connection intensity, with some super-connected genes. TF-specific HiChIP showed that FoxP3 interacts with EPLs at a large number of genes, including some not differentially expressed in Treg vs Tconv, but enriched at the core Treg signature loci that it upregulates. FoxP3 association correlates with heightened H3H27Ac looping, as ascertained by analysis of FoxP3-deficient Treg-like cells. There was marked asymmetry in the loci where FoxP3 associated at the enhancer- or the promoter-side of EPLs, with enrichment for different transcriptional cofactors. FoxP3 EPL intensity distinguished gene clusters identified by single-cell ATAC-seq as co-varying between individual Tregs, supporting a direct transactivation model for FoxP3 in determining Treg identity.

One Sentence Summary

FoxP3 is associated with enhancer-promoter loops in Treg cells, and correlates with heightened enhancer-promoter cross-talk
Molecular basis for the regulation of human glycogen synthase by phosphorylation and glucose-6-phosphate
Authors: McCorvie Thomas J. , Loria Paula M. , Tu Meihua , Han Seungil , Shrestha Leela , Froese D. Sean , Ferreira Igor M. , Berg Allison P. , Yue Wyatt W.
Published: November 2021

Glycogen synthase (GYS1), in complex with glycogenin (GYG1), is the central enzyme of muscle glycogen biosynthesis, and its inhibition has been proposed as a therapeutic avenue for various glycogen storage diseases (GSDs). GYS1 activity is inhibited by phosphorylation of its N- and C-termini, which can be relieved by allosteric activation of glucose-6-phosphate. However, the structural basis of GYS1 regulation is unclear. Here, we present the first cryo-EM structures of phosphorylated human GYS1 complexed with a minimal interacting region of GYG1 in the inhibited, activated, and catalytically competent states at resolutions of 3.0-4.0 Å. These structures reveal how phosphorylations of specific N- and C- terminal residues are sensed by different arginine clusters that lock the GYS1 tetramer complex in an inhibited state via inter-subunit interactions. The allosteric activator, glucose-6-phopshate, promotes a conformational change by disrupting these interactions and increases flexibility of GYS1 allowing for a catalytically competent state to occur when bound to the sugar donor UDP-glucose. We also identify an inhibited-like conformation that has not transitioned into the activated state, whereby the locking interaction of phosphorylation with the arginine cluster impedes the subsequent conformational changes due to glucose-6-phosphate binding. Finally, we show that the PP1 phosphatase regulatory subunit PPP1R3C (PTG) is recruited to the GYS1:GYG1 complex through direct interaction with glycogen. Our results address long-standing questions into the mechanism of human glycogen synthase regulation.
A Multi-Omics Longitudinal Aging Dataset in Primary Human Fibroblasts with Mitochondrial Perturbations
Authors: Sturm Gabriel , Monzel Anna S , Karan Kalpita R , Michelson Jeremy , Ware Sarah A. , Cardenas Andres , Lin Jue , Bris Céline , Santhanam Balaji , Murphy Michael P , Levine Morgan E , Horvath Steve , Belsky Daniel W , Wang Shuang , Procaccio Vincent , Kaufman Brett A. , Hirano Michio , Picard Martin
Published: November 2021

Aging is a process of progressive change. In order to develop biological models of aging, longitudinal datasets with high temporal resolution are needed. Here we report a multi-omic longitudinal dataset for cultured primary human fibroblasts measured across their replicative lifespans. Fibroblasts were sourced from both healthy donors (n=6) and individuals with lifespan-shortening mitochondrial disease (n=3). The dataset includes cytological, bioenergetic, DNA methylation, gene expression, secreted proteins, mitochondrial DNA copy number and mutations, cell-free DNA, telomere length, and whole-genome sequencing data. This dataset enables the bridging of mechanistic processes of aging as outlined by the “hallmarks of aging”, with the descriptive characterization of aging such as epigenetic age clocks. Here we focus on bridging the gap for the hallmark mitochondrial metabolism. Our dataset includes measurement of healthy cells, and cells subjected to over a dozen experimental manipulations targeting oxidative phosphorylation (OxPhos), glycolysis, and glucocorticoid signaling, among others. These experiments provide opportunities to test how cellular energetics affect the biology of cellular aging. All data are publicly available at our webtool:https://columbia-picard.shinyapps.io/shinyapp-Lifespan_Study/
A New Approach to Testing Mediation of the Microbiome using the LDM
Authors: Yue Ye , Hu Yi-Juan
Published: November 2021

Background

Understanding whether and which microbes played a mediating role between an exposure and a disease outcome are essential for researchers to develop clinical interventions to treat the disease by modulating the microbes. Existing methods for mediation analysis of the microbiome are often limited to a global test of community-level mediation or selection of mediating microbes without control of the false discovery rate (FDR). Further, while the null hypothesis of no mediation at each microbe is a composite null that consists of three types of null (no exposure-microbe association, no microbe-outcome association given the exposure, or neither), most existing methods for the global test such as MedTest and MODIMA treat the microbes as if they are all under the same type of null.

Methods

We propose a new approach based on inverse regression that regresses the (possibly transformed) relative abundance of each taxon on the exposure and the exposure-adjusted outcome to assess the exposure-taxon and taxon-outcome associations simultaneously. Then the association p-values are used to test mediation at both the community and individual taxon levels. This approach fits nicely into our Linear Decomposition Model (LDM) frame-work, so our new method is implemented in the LDM and enjoys all the features of the LDM, i.e., allowing an arbitrary number of taxa to be tested, supporting continuous, discrete, or multivariate exposures and outcomes as well as adjustment of confounding covariates, accom-modating clustered data, and offering analysis at the relative abundance or presence-absence scale. We refer to this new method as LDM-med.

Results

Using extensive simulations, we showed that LDM-med always controlled the type I error of the global test and had compelling power over existing methods; LDM-med always preserved the FDR of testing individual taxa and had much better sensitivity than alternative approaches. In contrast, MedTest and MODIMA had severely inflated type I error when different taxa were under different types of null. The flexibility of LDM-med for a variety of mediation analyses is illustrated by the application to a murine microbiome dataset.

Availability and Implementation

Our new method has been added to our R package LDM, which is available on GitHub at https://github.com/yijuanhu/LDM.
A deep learning framework for quantitative analysis of actin microridges
Authors: Bhavna Rajasekaran , Sonawane Mahendra
Published: November 2021

Microridges are evolutionarily conserved actin-rich protrusions present on the apical surface of the squamous epithelial cells. In zebrafish epidermal cells, microridges form self-evolving patterns due to the underlying actomyosin network dynamics. However, their morphological and dynamic characteristics have remained poorly understood owing to lack of automated segmentation methods. We achieved ~97% pixel-level accuracy with the deep learning microridge segmentation strategy enabling quantitative insights into their bio-physical-mechanical characteristics. From the segmented images, we estimated an effective microridge persistence length as ~0.61μm. We discovered the presence of mechanical fluctuations and found relatively greater stresses stored within patterns of yolk than flank, indicating distinct regulation of their actomyosin networks. Furthermore, spontaneous formations and positional fluctuations of actin clusters within microridge influenced pattern rearrangements over short length/time-scales. Our framework allows large-scale spatiotemporal analysis of microridges during epithelial development and probing of their responses to chemical and genetic perturbations to unravel the underlying patterning mechanisms.
The Stickland fermentation precursor trans-4-hydroxyproline differentially impacts the metabolism of Clostridioides difficile and commensal Clostridia
Authors: Reed A.D. , Fletcher J.R. , Huang Y.Y. , Thanissery R. , Rivera A.J. , Parsons R.J. , Stewart A. , Kountz D.J. , Shen A. , Balskus E.P. , Theriot C.M.
Published: November 2021

An intact gut microbiota confers colonization resistance against Clostridioides difficile through a variety of mechanisms, likely including competition for nutrients. Recently, proline was identified as an important environmental amino acid that C. difficile uses to support growth and cause significant disease. A post-translationally modified form, trans-4-hydroxyproline, is highly abundant in collagen, which is degraded by host proteases in response to C. difficile toxin activity. The ability to dehydrate trans-4-hydroxyproline via the HypD glycyl radical enzyme is wide-spread amongst gut microbiota, including C. difficile and members of the commensal Clostridia, suggesting that this amino acid is an important nutrient in the host environment. Therefore, we constructed a C. difficile ΔhypD mutant and found that it was modestly impaired in fitness in a mouse model of infection, and was associated with an altered microbiota when compared to mice challenged with the wild type strain. Changes in the microbiota between the two groups were largely driven by members of the Lachnospiraceae family and the Clostridium genus. We found that C. difficile and type strains of three commensal Clostridia had significant alterations to their metabolic gene expression in the presence of trans-4-hydroxyproline in vitro. The proline reductase (prd) genes were elevated in C. difficile, consistent with the hypothesis that trans-4-hydroxyproline is used by C. difficile to supply proline for fermentation. Similar transcripts were also elevated in some commensal Clostridia tested, although each strain responded differently. This suggests that the uptake and utilization of other nutrients by the commensal Clostridia may be affected by trans-4-hydroxyproline metabolism, highlighting how a common nutrient may be a signal to each organism to adapt to a unique niche. Further elucidation of the differences between them in the presence of hydroxyproline and other key nutrients will be important to determining their role in nutrient competition against C. difficile.
Ferric Citrate Uptake is a Virulence Factor in Uropathogenic Escherichia coli
Authors: Frick-Cheng Arwen E. , Sintsova Anna , Smith Sara N. , Pirani Ali , Snitkin Evan S. , Mobley Harry L.T
Published: November 2021

More than half of women will experience a urinary tract infection (UTI) with uropathogenic Escherichia coli (UPEC) causing ∼80% of uncomplicated cases. Iron acquisition systems are essential for uropathogenesis, and UPEC encode functionally redundant iron acquisition systems, underlining their importance. However, a recent UPEC clinical isolate, HM7 lacks this functional redundancy and instead encodes a sole siderophore, enterobactin. To determine if E. coli HM7 possesses unidentified iron acquisition systems, we performed RNA-sequencing under iron-limiting conditions and demonstrated that the ferric citrate uptake system (fecABCDE and fecIR) was highly upregulated. Importantly, there are high levels of citrate within urine, some of which is bound to iron, and the fec system is highly enriched in UPEC isolates compared to environmental or fecal strains. Therefore, we hypothesized that HM7 and other similar strains use the fec system to acquire iron in the host. Deletion of both enterobactin biosynthesis and ferric citrate uptake (ΔentB/ΔfecA) abrogates use of ferric citrate as an iron source and fecA provides an advantage in human urine in absence of enterobactin. However, in a UTI mouse model, fecA is a fitness factor independent of enterobactin production, likely due to the action of host Lipocalin-2 chelating ferrienterobactin. These findings indicate that ferric citrate uptake is used as an iron source when siderophore efficacy is limited, such as in the host during UTI. Defining these novel compensatory mechanisms and understanding the nutritional hierarchy of preferred iron sources within the urinary tract are important in the search for new approaches to combat UTI.

Importance

UPEC, the primary causative agent of uncomplicated UTI is responsible for five billion dollars in healthcare costs in the US each year. Rates of antibiotic resistance are on the rise therefore it is vital to understand the mechanisms of UPEC pathogenesis to uncover potential targets for novel therapeutics. Iron acquisition systems used to obtain iron from sequestered host sources are essential for UPEC survival during UTI and have been used as vaccine targets to prevent infection. This study reveals the ferric citrate uptake system is another important iron acquisition system that is highly enriched in UPEC strains. Ferric citrate uptake has not previously been associated with these pathogenic isolates, underlining the importance of the continued study of these strains to fully understand their mechanisms of pathogenesis.
Protection of mice against experimental cryptococcosis by synthesized peptides delivered in glucan particles
Authors: Specht Charles A. , Homan E. Jane , Lee Chrono K. , Mou Zhongming , Gomez Christina L. , Hester Maureen M. , Abraham Ambily , Rus Florentina , Ostroff Gary R. , Levitz Stuart M.
Published: November 2021

The high global burden of cryptococcosis has made development of a protective vaccine a public health priority. We previously demonstrated that a vaccine composed of recombinantCryptococcus neoformanschitin deacetylase 2 (Cda2) delivered in glucan particles (GPs) protects BALB/c and C57BL/6 mice from an otherwise lethal challenge with a highly virulentC. neoformansstrain. An immunoinformatic analysis of Cda2 revealed a peptide sequence predicted to have strong binding to the MHC Class II (MHC II) H2-IAd allele found in BALB/c mice. BALB/c mice vaccinated with GPs containing a 32 amino acid peptide (Cda2-Pep1) that included this strong binding region were protected from cryptococcosis. Protection was lost with GP-based vaccines containing versions of recombinant Cda2 protein and Cda2-Pep1 with mutations predicted to greatly diminish MHC II binding. Cda2 has homology to the three otherC. neoformanschitin deacetylases, Cda1, Cda3 and Fpd1, in the high MHC II binding region. GPs loaded with homologous peptides of Cda1, Cda3 and Fpd1 protected BALB/c mice from experimental cryptococcosis, albeit not as robustly as the Cda2-Pep1 vaccine. Finally, seven other peptides were synthesized based on regions in Cda2 predicted to contain promising CD4+T cell epitopes in BALB/c or C57BL/6 mice. While five peptide vaccines significantly protected BALB/c mice, only one protected C57BL/6 mice. Thus, GP-based vaccines containing a single peptide can protect mice against cryptococcosis. However, given the diversity of human MHC II alleles, a peptide-basedCryptococcusvaccine for use in humans would be challenging and likely need to contain multiple peptide sequences.

Importance

Cryptococcosis, due to infection by fungi of theCryptococcus neoformansspecies complex, is responsible for substantial morbidity and mortality in immunocompromised persons, particularly those with AIDS. Cryptococcal vaccines are a public health priority yet are not available for human use. We previously demonstrated mice could be protected from experimental cryptococcosis with vaccines composed of recombinant cryptococcal proteins encased in hollow highly purified yeast cell walls (glucan particles). Here, we examined one such protective protein, Cda2, and using bioinformatics, identified a region predicted to stimulate strong T cell responses. A peptide containing this region formulated in glucan particle-based vaccines protected mice as well as the recombinant protein. Other peptide vaccines also protected, including peptides containing sequences from proteins homologous to Cda2. These preclinical mouse studies provide a proof of principle that peptides can be effective as vaccines to protect against cryptococcosis and that bioinformatic approaches can guide peptide selection.
Polygenic risk scores based on European GWAS correlate to disease prevalence differences around the world
Authors: Jain Pritesh R , Burch Myson , Martinez Melanie , Mir Pablo , Fichna Jakub , Zekanowski Cezary , Rizzo Renata , Tümer Zeynep , Barta Csaba , Yannaki Evangelia , Stamatoyannopoulos John , Drineas Petros , Paschou Peristera
Published: November 2021

Background

Complex disorders are caused by a combination of genetic, environmental and lifestyle factors, and their prevalence can vary greatly across different populations. Genome wide association studies (GWAS) can help identify common variants that underlie disease risk. However, despite their increasing number, the vast majority of studies focuses on European populations, leading to questions regarding the transferability of findings to non-Europeans. Here, we investigated whether polygenic risk scores (PRS) based on European GWAS correlate to disease prevalence within Europe and around the world.

Results

GWAS summary statistics of 20 different disorders were used to estimate PRS in nine European and 24 worldwide reference populations. We estimated the correlation between average genetic risk for each of the 20 disorders and their prevalence in Europe and around the world. A clear variation in genetic risk was observed based on ancestry and we identified populations that have a higher genetic liability for developing certain disorders both within European and global regions. We also found significant correlations between worldwide disease prevalence and PRS for 13 of the studied disorders with obesity genetic risk having the highest correlation to disease prevalence. For these 13 disorders we also found that the loci used in PRS are significantly more conserved across the different populations compared to randomly selected SNPs as revealed by Fst and linkage disequilibrium structure.

Conclusion

Our results show that PRS of world populations calculated based on European GWAS data can significantly capture differences in disease risk and identify populations with the highest genetic liability to develop various conditions. Our findings point to the potential transferability of European-based GWAS results to non-European populations and provide further support for the validity of GWAS.
Viral load and infectivity of SARS-CoV-2 in paired respiratory and oral specimens from symptomatic, asymptomatic or post-symptomatic individuals
Authors: Tallmadge Rebecca L. , Laverack Melissa , Cronk Brittany , Venugopalan Roopa , Martins Mathias , Zhang XiuLin , Elvinger François , Plocharczyk Elizabeth , Diel Diego G.
Published: November 2021

In the present study, we assessed the diagnostic sensitivity and determined the viral load and infectivity of SARS-CoV-2 in paired respiratory (nasopharyngeal and anterior nares) and oral samples (saliva and sublingual swab). Samples were collected from 77 individuals of which 75 were diagnosed with COVID-19 and classified as symptomatic (n=29), asymptomatic (n=31), or post-symptomatic (n=15). Specimens were collected at one time point from each individual, between day 1 to 23 after the initial COVID-19 diagnosis, and included self-collected saliva (S), or sublingual (SL) swab, and bilateral anterior nares (AN) swab, followed by healthcare provider collected nasopharyngeal (NP) swab. Sixty-three specimen sets were tested using five assay/platforms. The diagnostic sensitivity of each assay/platform and specimen type was determined. Of the 63 specimen sets, SARS-CoV-2 was detected in 62 NP specimens, 52 AN specimens, 59 saliva specimens, and 31 SL specimens by at least one platform. Infectious SARS-CoV-2 was isolated from 21 NP, 13 AN, 12 saliva, and one SL specimen out of 50 specimen sets. SARS-CoV-2 isolation was most successful up to 5 days after initial COVID-19 diagnosis using NP specimens from symptomatic patients (16 of 24 positives, 66.67%), followed by specimens from asymptomatic patients (5 of 17 positives, 29.41%), while it was not very successful with specimens from post-symptomatic patients. Benefits of self-collected saliva and AN specimens balance the loss of sensitivity relative to NP specimens. Therefore, saliva and AN specimens are acceptable alternatives for symptomatic SARS-CoV-2 diagnostic testing or surveillance with increased sampling frequency of asymptomatic individuals.

Importance

The dynamics of infection with SARS-CoV-2 has a significant impact on virus infectivity and in the diagnostic sensitivity of molecular and classic virus detection tests. In the present study we determined the diagnostic sensitivity of paired respiratory (nasopharyngeal and anterior nares swabs) and oral secretions (saliva and sublingual swab) and assessed infectious virus shedding patterns by symptomatic, asymptomatic or post-symptomatic individuals. Understanding the diagnostic performance of these specimens and the patterns of infectious virus shedding in these bodily secretions provides critical information to control COVID-19, and may help to refine guidelines on isolation and quarantine of positive individuals and their close contacts identified through epidemiological investigations.
Age Based Trends for Unplanned Pediatric Rehospitalizations in the United States
Authors: Amritphale Nupur , Amritphale Amod , Vasireddy Deepa , Batra Mansi , Sehgal Mukul , Gremse David
Published: November 2021

BACKGROUND AND OBJECTIVES

Hospital readmission rate helps to highlight the effectiveness of post-discharge care. There remains a paucity of plausible age based categorization especially for ages below one year for hospital readmission rates.

METHODS

Data from 2017 Healthcare cost and utilization project National readmissions database was analyzed for ages 0-18 years. Logistic regression analysis was performed to identify predictors for unplanned early readmissions.

RESULTS

We identified 5,529,389 inpatient pediatric encounters which were further divided into age group cohorts. The overall rate of readmissions was identified at 3.2%. Beyond infancy, the readmission rate was found to be 6.7%. Across all age groups, the major predictors of unplanned readmission were cancers, diseases affecting transplant recipients and sickle cell patients. It was determined that reflux, milk protein allergy, hepatitis and inflammatory bowel diseases were significant comorbidities leading to readmission. Anxiety, depression and suicidal ideation depicted higher readmission rates in those older than 13 years. Across ages 1-4 yrs, dehydration, asthma and bronchitis were negative predictors of unplanned readmission.

CONCLUSIONS

Thirty-day unplanned readmissions remain a problem leading to billions of tax-payer-dollars lost per annum. Effective strategies for mandatory outpatient follow-up may help the financial aspect of care while also enhancing the quality of care.
Implementation of Unassisted and Community-Based HIV Self-Testing (HIVST) during the COVID-19 pandemic among Men-who-have-sex-with-Men (MSM) and Transgender Women (TGW): A Demonstration Study in Metro Manila, Philippines
Authors: Rosadiño John Danvic T. , Pagtakhan Ronivin G. , Briñes Matthew T. , Dinglasan Jeanno Lorenz G. , Cruz Denis P. , Corciega John Oliver L. , Pagtakhan Aeronne B. , Regencia Zypher Jude G. , Baja Emmanuel S.
Published: November 2021

Objective

The study aimed to demonstrate the feasibility of an unassisted and community-based HIV self-testing (HIVST) distribution model and to evaluate its acceptability among men-having-sex-with-men (MSM) and transgender women (TGW).

Methods

Our observational study focused on implementing the HIVST service in Metro Manila, Philippines. Convenience sampling was done with the following inclusion criteria: MSM or TGW, at least 18 years old, and had no previous HIV diagnosis. Individuals taking HIV Pre-exposure Prophylaxis (PrEP), on Antiretroviral Therapy (ART), or female sex at birth were excluded. The implementation of the study was online using a virtual assistant and delivery system via courier due to COVID-19-related lockdowns. Feasibility was measured by the number of HIVST kits successfully delivered and utilized and the HIV point prevalence rate. Moreover, acceptability was evaluated by a 10-item system usability scale (SUS). HIV prevalence was estimated with linkage to care prioritized for reactive participants.

Results

Out of 1,690 kits distributed, only 953 (56.4%) participants reported their results. Overall HIV point prevalence was 9.8%, with 56 (60.2%) reactive participants linked to further testing. Furthermore, 27.4% of respondents self-reported, and 13.4% of the reactive participants were first-time testers. The HIVST service had an overall mean ± standard deviation SUS score of 81.0 ± 13.0, rendering the HIVST kits very acceptable.

Conclusions

HIVST is acceptable and feasible to MSM and TGW. Online platforms are an innovative and effective way to deliver HIVST service during a pandemic. However, messaging to entice people to use the kit must be differentiated based on their age, gender identity and expression, and previous HIVST experience to offer the service efficiently to the target populations.
Cryo-EM structure of the Inner Ring fromXenopus laevisNuclear Pore Complex
Authors: Huang Gaoxingyu , Zhan Xiechao , Zeng Chao , Liang Ke , Zhu Xuechen , Zhao Yanyu , Wang Pan , Wang Qifan , Zhou Qiang , Tao Qinghua , Liu Minhao , Lei Jianlin , Yan Chuangye , Shi Yigong
Published: November 2021

Nuclear pore complex (NPC) mediates nucleocytoplasmic shuttling. Here we present single-particle cryo-electron microscopy (cryo-EM) structure of the inner ring (IR) subunit fromXenopus laevisNPC at an average resolution of 4.4 Å. The symmetric IR subunit comprises a cytoplasmic half and a nuclear half. A homo-dimer of Nup205 resides at the center of the IR subunit, flanked by two molecules of Nup188. Four molecules of Nup93 each places an extended helix into the axial groove of Nup205 or Nup188, together constituting the central scaffold. The channel nucleoporin heterotrimer (CNT) of Nup54/58/62 is anchored on the central scaffold. Six Nup155 molecules interact with the central scaffold and together with the NDC1-ALADIN hetero-dimers anchor the IR subunit to the nuclear envelope and to outer rings. The scarce inter-subunit contacts may allow sufficient latitude in conformation and diameter of the IR. Our structure of vertebrate IR reveals key insights that are functionally important.
Reliable population signal of subjective economic value from unreliable neurons in primate orbitofrontal cortex
Authors: Ferrari-Toniolo Simone , Schultz Wolfram
Published: November 2021

Behavior-related neuronal signals often vary between neurons. Despite the unreliability of individual neurons, brains are able to accurately represent and drive behavior. The notion may also apply to economic (‘value-based’) choices and the underlying reward signals. Reward value is subjective and can be defined by nonlinear weighting of magnitude (utility) and probability. Using a wide variety of reward magnitude and probability, we assessed subjective reward value at choice indifference between safe and risky rewards as prescribed by the continuity axiom that provides stringent criteria for meaningful choice. We found that individual neurons in the orbitofrontal cortex (OFC) of monkeys carry unreliable and heterogeneous neuronal signals for subjective value that largely fails to match the animal’s choice. However, the averaged neuronal signals matched well the animals’ choices, suggesting reliable subjective economic value encoding by the observed population of unreliable neurons.

Highlights

Different from widely held views, reliable neuronal information processing may not require reliable processors.

Neurons in monkey orbitofrontal cortex (OFC) process reward magnitude and probability heterogeneously and unreliably.

Despite unreliable neuronal processing, OFC population activity codes choices reliably.

Reliability systems performance from unreliable elements seems to be a broad feature of neuronal reward coding.

In brief

Using stringent concepts of behavioral choice, Ferrari-Toniolo and Schultz describe unreliable individual reward neurons in monkey orbitofrontal cortex whose activity combines to a reliable population code for economic choice.
Siderophores as an iron source for Prochlorococcus in deep chlorophyll maximum layers of the oligotrophic ocean
Authors: Hogle Shane L , Hackl Thomas , Bundy Randelle M , Park Jiwoon , Satinsky Brandon , Hiltunen Teppo , Biller Steven , Berube Paul M , Chisholm Sallie W
Published: November 2021

Prochlorococcus is one of the most abundant photosynthesizing organisms in the oligotrophic oceans. Gene content variation among Prochlorococcus populations in separate ocean basins often mirrors the selective pressures imposed by the region's distinct biogeochemistry. By pairing genomic datasets with trace metal concentrations from across the global ocean, we show that the genomic capacity for siderophore-mediated iron uptake is widespread in low-light adapted Prochlorococcus populations from iron-depleted regions of the oligotrophic Pacific and S. Atlantic oceans: Prochlorococcus siderophore consumers were absent in the N. Atlantic ocean (higher iron flux) but constituted up to half of all Prochlorococcus genomes from metagenomes in the N. Pacific (lower iron flux). Prochlorococcus siderophore consumers, like many other bacteria with this trait, also lack siderophore biosynthesis genes indicating that they scavenge exogenous siderophores from seawater. Statistical modeling suggests that the capacity for siderophore uptake is endemic to remote ocean regions where atmospheric iron fluxes are the smallest, particularly at deep chlorophyll maximum and primary nitrite maximum layers. We argue that abundant siderophore consumers at these two common oceanographic features could be a symptom of wider community iron stress, consistent with prior hypotheses. Our results provide a clear example of iron as a selective force driving the evolution of Prochlorococcus.
Uncovering natural variation in root system architecture and growth dynamics using a robotics-assisted phenomics platform
Authors: LaRue Therese , Lindner Heike , Srinivas Ankit , Exposito-Alonso Moises , Lobet Guillaume , Dinneny José R.
Published: November 2021

The plant kingdom contains a stunning array of complex morphologies easily observed above ground, but largely unexplored below-ground. Understanding the magnitude of diversity in root distribution within the soil, termed root system architecture (RSA), is fundamental to determining how this trait contributes to species adaptation in local environments. Roots are the interface between the soil environment and the shoot system and therefore play a key role in anchorage, resource uptake, and stress resilience. Previously, we presented the GLO-Roots (Growth and Luminescence Observatory for Roots) system to study the RSA of soil-grownArabidopsis thalianaplants from germination to maturity (Rellán-Álvarez et al. 2015). In this study, we present the automation of GLO-Roots using robotics and the development of image analysis pipelines in order to examine the natural variation of RSA in Arabidopsis over time. This dataset describes the developmental dynamics of 93 accessions and reveals highly complex and polygenic RSA traits that show significant correlation with climate variables.
Poly ADP-ribosylation of SET8 leads to aberrant H4K20 methylation in mammalian nuclear genome
Authors: Estève Pierre-Olivier , Sen Sagnik , Vishnu US , Ruse Cristian , Chin Hang Gyeong , Pradhan Sriharsa
Published: November 2021

In mammalian cells, SET8 mediated Histone H4 Lys 20 monomethylation (H4K20me1) has been implicated in regulating mitotic condensation, DNA replication, DNA damage response, and gene expression. Here we show SET8, the only known enzyme for H4K20me1 is post-translationally poly ADP-ribosylated by PARP1 on lysine residues. PARP1 interacts with SET8 in a cell cycle-dependent manner. Poly ADP-ribosylation on SET8 renders it catalytically compromised, and degradation via ubiquitylation pathway. Knockdown of PARP1 led to an increase of SET8 protein levels, leading to aberrant H4K20me1 and H4K20me3 domains genome-wide. H4K20me1 is associated with higher gene transcription levels while the increase of H4K20me3 levels was predominant in DNA repeat elements correlating with modulation of chromatin states. Hence, SET8 mediated chromatin remodeling in mammalian cells are modulated by poly ADP-ribosylation by PARP1.
Precision size and refractive index analysis of weakly scattering nanoparticles in polydispersions
Authors: Kashkanova Anna D. , Blessing Martin , Gemeinhardt André , Soulat Didier , Sandoghdar Vahid
Published: November 2021

Characterization of the size and material properties of particles in liquid suspensions is in very high demand, e.g., for the analysis of colloidal samples or of bodily fluids such as urine or blood plasma. However, the existing methods are limited in deciphering the constituents of realistic samples. Here, we introduce iNTA as a new method, which combines interferometric detection of scattering with nanoparticle tracking analysis, to reach an unprecedented sensitivity and precision in determining the size and refractive index distributions of nanoparticles in suspensions. After benchmarking iNTA with samples of colloidal gold, we present its remarkable ability to resolve the constituents of various multi-component and polydisperse samples of known origin. Furthermore, we showcase the method by elucidating the refractive index and size distributions of extracellular vesicles from Leishmania parasites and nanoparticles in human urine. The current performance of iNTA already enables advances in several important applications, but we also discuss possible improvements.