bluuebunny/crossref_metadata_2025 huggingface dataset:
Circulating Small Extracellular Vesicle RNA Profiling for the Detection of T1a stage Colorectal Cancer and Precancerous Advanced Adenoma
Authors: Min Li , Bu Fanqin , Meng Jingxin , Liu Xiang , Guo Qingdong , Zhao Libo , Li Zhi , Li Xiangji , Zhu Shengtao , Zhang Shutian
Published: June 2023

It takes more than twenty years for normal colorectal mucosa to develop into metastatic carcinoma. The long time window provides a golden opportunity for early detection to terminate the malignant progression. Here we aim to enable liquid biopsy of T1a stage colorectal cancer (CRC) and precancerous advanced adenoma (AA) by profiling circulating small extracellular vesicle (sEV)-derived RNAs. We exhibited a full RNA landscape for the circulating sEVs isolated from 60 participants. A total of 58,333 annotated RNAs were detected from plasma sEVs, among which 1,615 and 888 sEV-RNAs were found differentially expressed in plasma from T1a stage CRC and AA compared to normal controls (NC). Then we further categorized these sEV-RNAs into 6 modules by a weighted gene coexpression network analysis and constructed a 60-gene t-SNE model consisting of the top 10 RNAs of each module that could well distinguish T1a stage CRC/AA from NC samples. Some sEV-RNAs were also identified as indicators of specific endoscopic and morphological features of different colorectal lesions. The top-ranked biomarkers were further verified by RT-qPCR, proving that these candidate sEV-RNAs successfully identified T1a stage CRC/AA from NC in another cohort of 124 participants. Finally, we adopted different algorithms to improve the performance of RT-qPCR-based models and successfully constructed an optimized classifier with 79.3% specificity and 99.0% sensitivity. In conclusion, circulating sEVs of T1a stage CRC and AA patients have distinct RNA profiles, which successfully enable the detection of both T1a stage CRC and AA via liquid biopsy.
Ryegrass mottle virus complete genome determination and development of infectious cDNA by combining two methods – 3′ RACE and 5′ RACE-seq
Authors: Balke Ina , Silamikelis Ivars , Radovica-Spalvina Ilze , Zeltina Vilija , Resevica Gunta , Fridmanis Davids , Zeltins Andris
Published: June 2023

Sobemovirus ryegrass mottle virus (RGMoV) is a single-stranded positive virus with a 30 nm viral particle size. It exhibitsT=3symmetry, with 180 coat protein (CP) subunits forming the virus structure. The RGMoV genome comprises five open reading frames, encoding P1, Px, a membrane-anchored 3C-like serine protease, a virus genome-linked protein, P16, an RNA-dependent RNA polymerase, and a coat protein. The RGMoV genome size varies, ranging from 4175 nt (MW411579.1) to 4253 nt (MW411579.1) in deposited sequences. An earlier deposited RGMoV complete genome sequence of 4212 nt length (EF091714.1) was utilized to develop an infectious complementary DNA (icDNA) construct forin vitrogRNA transcription from the T7 promoter. However, when the transcribed gRNA was introduced to oat plants, it failed to induce viral infection. This indicated the potential absence of certain sequences in either the 5’ or 3’ untranslated regions (UTR) or both. To resolve this, the complete sequence of the 3’ UTR was determined through 3’ end RACE, while the 5’ UTR was identified using high-throughput sequencing (HTS) - 5’ RACE-seq. Only the icDNA vector containing both newly identified UTR sequences proved infectious, resulting in classical viral infection symptoms and subsequent propagation of progeny viruses, exhibiting the ability to cause repeated infection in oat plants after at least one passage. The successful generation of the icDNA highlights the synergistic potential of utilizing both methods when one approach alone fails. Furthermore, this study demonstrates the reliability of HTS as a method for determining the complete genome sequence of viral genomes.
Structural basis of EHEP-mediated offense against phlorotannin-induced defense from brown algae to protectakuBGL activity
Authors: Sun Xiaomei , Ye Yuxin , Sakurai Naofumi , Wang Hang , Kato Koji , Yu Jian , Yuasa Keizo , Tsuji Akihiko , Yao Min
Published: June 2023

The defensive-offensive associations between algae and herbivores determine marine ecology. Brown algae utilize phlorotannin as their chemical defense against the predatorAplysia kurodai, which uses β-glucosidase (akuBGL) to digest the laminarin in algae to glucose. Moreover,A. kurodaiemploysEiseniahydrolysis-enhancing protein (EHEP) as an offense to protectakuBGL activity from phlorotannin inhibition by precipitating phlorotannin. To underpin the molecular mechanism of this digestive-defensive-offensive system, we determined the structures of apo and tannic-acid (TNA, a phlorotannin-analog) bound form of EHEP, as well as apoakuBGL. EHEP consisted of three peritrophin-A domains formed in a triangle and bound TNA in the center without significant conformational changes. Structural comparison between EHEP and EHEP–TNA led us to find that EHEP can be resolubilized from phlorotannin-precipitation at an alkaline pH, which reflects a requirement in the digestive tract.akuBGL contained two GH1 domains, only one of which conserved the active site. Combining docking analysis, we propose the mechanisms by which phlorotannin inhibitsakuBGL by occupying the substrate-binding pocket, and EHEP protectsakuBGL against the inhibition by binding with phlorotannin to free theakuBGL pocket.
Mitral Annular Disjunction in Out-of-Hospital Cardiac Arrest Patients – a Retrospective Cardiac MRI Study
Authors: Troger Felix , Klug Gert , Poskaite Paulina , Tiller Christina , Lechner Ivan , Reindl Martin , Holzknecht Magdalena , Fink Priscilla , Brunnauer Eva-Maria , Gizewski Elke R. , Metzler Bernhard , Reinstadler Sebastian , Mayr Agnes
Published: June 2023

Background

Mitral annular disjunction (MAD), defined as defective attachment of the mitral annulus to the ventricular myocardium, has recently been linked to malignant arrhythmias. However, its role and prognostic significance in patients requiring cardio-pulmonary resuscitation (CPR) remains unknown. This retrospective analysis aimed to describe prevalence and significance of MAD by cardiac magnetic resonance (CMR) imaging, in out-of-hospital cardiac arrest (OHCA) patients.

Methods

Eighty-six patients with OHCA and a CMR scan 5 days after CPR (interquartile range (IQR): 49 days before – 9 days after) were consecutively enrolled. MAD was defined as disjunction-extent ≥1mm in CMR long-axis cine-images. Medical records were screened for laboratory parameters, comorbidities and prior arrhythmias.

Results

In 34 patients (40%), no underlying cause for OHCA was found during hospitalization despite profound diagnostics. Unknown-cause OHCA patients showed a higher prevalence of MAD compared to definite-cause patients (56% vs. 10%, p<0.001) and had a MAD-extent of 6.3mm (IQR: 4.4-10.3); moreover, these patients were significantly younger (43 years vs. 61 years, p<0.001), more often female (74% vs. 21%, p<0.001) and had fewer comorbidities (hypertension, hypercholesterolemia, coronary artery disease, all p<0.005). By logistic regression analysis, presence of MAD remained significantly associated with OHCA of unknown cause (odds ratio: 8.49, 95% confidence interval: 2.37-30.41, p=0.001) after adjustment for age, presence of hypertension and hypercholesterolemia.

Conclusions

MAD is rather common in OHCA patients without definitive aetiology undergoing CMR. Presence of MAD remains independently associated to OHCA without identifiable trigger. Further research is needed to understand the exact role of MAD in OHCA patients.

Clinical Perspectives

This study showed that MAD is apparently a common finding in cardiac arrest patients without underlying trigger and was associated with it independently of age, concomitant hypertension and hypercholesterolemia.

In clinical routine, MAD should be considered as potential arrhythmogenic substrate especially in those cardiac arrest patients, in which eventually no clear etiology can be found.

However, future studies need to further explore the role of MAD in these patients and investigate the true arrhythmogenic potential of this anatomical variant.
Investigating the genetic relationship between depression symptoms and Alzheimer’s Disease in clinically diagnosed and proxy cases
Authors: Gilchrist Lachlan , Spargo Thomas P. , Green Rebecca E. , Coleman Jonathan R.I. , Howard David M. , Thorp Jackson G. , Adey Brett , Lord Jodie , Davies Helena L. , Mundy Jessica , ter Kuile Abigail , Davies Molly R. , Hübel Christopher , Bristow Shannon , Lee Sang Hyuck , Rogers Henry , Curtis Charles , Kalsi Gursharan , Arathimos Ryan , Corbett Anne , Ballard Clive , Brooker Helen , Creese Byron , Aarsland Dag , Hampshire Adam , Velayudhan Latha , Eley Thalia C. , Breen Gerome , Iacoangeli Alfredo , Koks Sulev , Lewis Cathryn M. , Proitsi Petroula
Published: June 2023

Depression is a risk factor for Alzheimer’s disease (AD), but evidence for their genetic relationship is mixed. Assessing depression symptom specific genetic associations may better clarify this relationship.

Using data from the UK Biobank, the GLAD Study and PROTECT, we performed the largest genome-wide meta-analyses (GWAS) of the nine depression symptom items, plus their sum score, on the Patient Health Questionnaire (PHQ-9) (GWAS equivalent N: 224,535—308,421). We assessed global/local genetic correlations and statistical colocalisation between depression phenotypes and AD across six AD GWAS with varying proportions of clinical and proxy (family history) case ascertainment. We assessed bi-directional causal associations using Mendelian randomisation (MR) and the predictiveness of depression phenotype polygenic risk scores (PRS) for AD case/control status in three clinical AD cohorts.

Our GWAS meta-analyses identified 37 genomic risk loci across the ten depression symptom phenotypes. Of the 72 global genetic correlation tests conducted between depression/depression symptoms and AD, 20 were significant at pFDR≤ 0.05. Only one significant genetic correlation was identified with AD GWAS containing clinical-only cases. Colocalisation was not identified at loci contains local genetic correlation but was identified in the region of transmembrane protein 106B (TMEM106B) between multiple depression phenotypes and both clinical-only and clinical+proxy AD. MR and PRS analyses did not yield statistically significant results.

Our findings do not demonstrate a causal role of depression/depression symptoms on AD and suggest that previous evidence of their genetic overlap may be driven by the inclusion of proxy cases/controls. However, the identification of colocalisation at TMEM106B warrants further investigation.
Pulmonary Radioaerosol Mucociliary Clearance Parameters as Potential Outcomes in Primary Ciliary Dyskinesia Trials
Authors: Marthin June K. , Holgersen Mathias G. , Nielsen Kim G. , Mortensen Jann
Published: June 2023

Background

Pulmonary Radioaerosol Mucociliary Clearance (PRMC) is anin vivowhole lung ciliary function test reliable for assessing mucociliary clearance for diagnostic purposes in individuals suspected of primary ciliary dyskinesia (PCD). We aimed to evaluate expanded use of PRMC by providing advantages and limitations for its potential use in providing outcome parameters in future trials aiming to restore ciliary activity.

Material and Methods

In this retrospective study, PRMC tests performed over a period of 24 years (1999-2022) were meticulously re-analyzed. Patients with genetically verified PCD and non-PCD controls were included. Originally, nebulized99mTc-albumin colloid was inhaled, and static and dynamic imaging acquired for 60 and 120 minutes, and 24 hours.

For the purpose of the present study 3 PRMC parameters were defined: 1 hour lung retention (LR1), tracheobronchial velocity (TBV), and cough clearance.

Results

Sixty-nine patients were included from the Danish PCD cohort. PRMC was overall completely absent regardless of PCD genotypes. In one patient with CCDC103 mutation, residual ciliary function and normal nasal NO, we found normal PRMC LR1 and measurable, however low, TBV.

Voluntary cough significantly increased clearance with a median (IQR) of 11 (4;24) %.

Conclusion

Absolute absence of PRMC would be the expected baseline result in by far the majority of patients with PCD regardless of genotype before introducing ciliary protein correctors in a clinical trial.

Measurable PRMC TBV and normal LR1 in one patient with residual ciliary function, indicated that PRMC parameters could potentially improve if ciliary function was to be restored during a clinical trial.

Involuntary cough and peripheral radioaerosol deposition were the main challenges of the PRMC method.
Association of Long COVID with housing insecurity in the United States, 2022-2023
Authors: Packard Samuel E. , Susser Ezra
Published: June 2023

Objectives

To assess the association of Long COVID with housing insecurity in the United States.

Methods

To compare the prevalence of 3 binary indicators of housing insecurity between people with Long COVID (symptoms > 3 months) and COVID-19 survivors who don’t report long-term symptoms, we used survey-weighted regression models on 203,807 responses from the Household Pulse Survey, a representative cross-sectional survey of US households collected September 2022 – April 2023. Among people with Long COVID, we assessed whether functional impairment, current COVID-19 related symptoms, and symptom impact on day-to- day life were associated with a higher prevalence of housing insecurity.

Results

During the study period, 54,446 (27.2%) respondents with COVID-19 experienced symptoms lasting 3 months or longer, representing an estimated 27 million US adults. People with Long COVID were nearly twice as likely to experience significant difficulty with household expenses (Prevalence ratio [PR] 1.85, 95% CI 1.74-1.96), be behind on housing payments (PR 1.76, 95% CI 1.57-1.99), and face likely eviction or foreclosure (PR 2.12, 95% CI 1.58-2.86). Functional limitation and current symptoms which impact day-to-day life were associated with higher prevalence of housing insecurity.

Conclusions

Compared with COVID-19 survivors who don’t experience long-term symptoms, people with Long COVID are more likely to report indicators housing insecurity, particularly those with functional limitations and long-term COVID-19 related symptoms impacting day-to- day life. Policies are needed to support people living with chronic illnesses following SARS- CoV-2 infection.
How could a pooled testing policy have performed in managing the early stages of the COVID-19 pandemic? Results from a simulation study
Authors: Heath Bethany , Villar Sofía S. , Robertson David S.
Published: June 2023

A coordinated testing policy is an essential tool for responding to emerging epidemics, as was seen with COVID-19. However, it is very difficult to agree on the best policy when there are multiple conflicting objectives. A key objective is minimising cost, which is why pooled testing (a method that involves pooling samples taken from multiple individuals and analysing this with a single diagnostic test) has been suggested. In this paper, we present results from an extensive and realistic simulation study comparing testing policies based on individually testing subjects with symptoms (a policy resembling the UK strategy at the start of the COVID-19 pandemic), individually testing subjects at random or pools of subjects randomly combined and tested. To compare these testing methods, a dynamic model compromised of a relationship network and an extended SEIR model is used. In contrast to most existing literature, testing capacity is considered as fixed and limited rather than unbounded. This paper then explores the impact of the proportion of symptomatic infections on the expected performance of testing policies. Only for less than 50% of infections being symptomatic does pooled testing outperform symptomatic testing in terms of metrics such as total infections and length of epidemic. Additionally, we present the novel feature for testing of non-compliance and perform a sensitivity analysis for different compliance assumptions. Our results suggest for the pooled testing scheme to be superior to testing symptomatic people individually, only a small proportion of the population (>2%) needs to not comply with the testing procedure.
More spice, less salt: how capsaicin affects liking for and perceived saltiness of foods in people with smell loss
Authors: Hunter Stephanie R. , Beatty Candelaria , Dalton Pamela H.
Published: June 2023

People who lose their sense of smell self-report consuming more salt to compensate for a lack of flavor and enhance eating enjoyment. However, this can contribute to excess sodium intake and a poor diet. Capsaicin may help increase salt taste intensity and eating enjoyment in this population, but this has not been studied. The purpose of this study was to determine 1) whether salt intake in those with smell loss differs from population averages, 2) whether capsaicin increases flavor and salt taste intensity, and 3) if adding spice to foods increases food liking in individuals with smell loss. Participants 18-65 years old with confirmed partial or total smell loss for at least 12 weeks completed two sets of replicate test sessions (four total). In two sessions participants rated overall flavor intensity, taste qualities’ intensities, spicy intensity, and liking for model tomato soups with low or regular sodium content and three levels of capsaicin (none, low, or moderate). In the other two sessions, participants rated the same sensory attributes for model food samples with three levels of added spice (none, low, or moderate). 24-hour urine samples were also collected to determine sodium intake. Results indicate that although sodium intake is higher than recommended in those with smell loss (2893 ± 258 mg/day), they do not consume more sodium than population averages. Adding low and moderate amounts of capsaicin to a model tomato soup increased the intensity of overall flavor and saltiness compared to a model tomato soup without capsaicin. However, the effect of capsaicin on liking differed by food type. In conclusion, the addition of capsaicin can improve flavor, salt taste intensity, and eating enjoyment in people with smell loss.
Efficacy of conventional rehabilitation, transcutaneous electrical nerve stimulation, or early mobilization to reverse acquired weakness in the intensive care unit. Randomized controlled pilot trial
Authors: Bueno-Ardariz César Alejandro , Cagide Sabrina Gimena , Gamarra Claudio Gabriel , Paz Darío Leonel , Rotela Eliana Verónica , Aranda-Castro Ricard , Bustos Fernando Daniel , Muller Sebastián , Settembrino Esteban Sebastián , Carrera Melina , Giongo Cecilia Elena , Nannini Gonzalo Martín , Nardelli Paula Gimena , Diaz-Ballve Ladislao Pablo
Published: June 2023

Objective

To compare the efficacy of conventional rehabilitation, transcutaneous electrical nerve stimulation, and early mobilization in reducing the time needed to reverse intensive care unit-acquired weakness (ICUAW), as assessed by the Medical Research Council muscle strength scale (MRC-MSS) score, in patients clinically diagnosed with ICUAW.

Setting

Medical-surgical ICU of a general acute care hospital with 26 beds.

Design

Pilot trial with random assignment and a control group.

Interventions

Group 1: standard or routine rehabilitation, Group 2: transcutaneous electrical nerve stimulation, and Group 3: early mobilization protocol.

Subjects

Individuals over 18 years old admitted to the ICU with invasive mechanical ventilation for more than 24 hours and diagnosed with ICUAW defined as Score MRC-MSS<48 measured over two consecutive days.

Results

18 subjects were included. One-way ANOVA showed differences in the treatment days needed to achieve an MRC-MSS ≥48 (p = 0.01), as well as in the number of sessions required to reverse ICUAW (p = 0.01) when comparing the different interventions. Through post hoc analysis, the group treated with transcutaneous electrical nerve stimulation was identified as requiring significantly more days and sessions to reverse ICUAW. Differences were observed in the Barthel index at the end and in the variation of the Barthel index as a result of functional capacity among the different treatments.

Conclusion

The group treated with transcutaneous electrical nerve stimulation required a significantly greater number of days and sessions to achieve an MRC-SS ≥48. No differences in functional limitation were observed. These findings should be corroborated in similar studies. (Clinicaltrial.govNCT04613908)
Genetic Diagnosis of Facioscapulohumeral Muscular Dystrophy Type 1 Using Rare Variant Linkage Analysis and Long Read Genome Sequencing
Authors: Li Kun , Quiat Daniel , She Fei , Liu Yuanwei , He Rong , Haghighi Alireza , Liu Fang , Zhang Rui , DePalma Steven Robert , Yang Ying , Wang Wen , Seidman Christine E. , Zhang Ping , Seidman Jonathan G.
Published: June 2023

Facioscapulohumeral dystrophy type 1 (FSHD1) is a progressive, debilitating skeletal myopathy that requires a multimodal approach for complete molecular characterization of pathogenic genotypes. Here, we report genomic analyses of a family with suspected facioscapulohumeral dystrophy type 1 (FSHD1). We first performed short read genome sequencing, followed by parametric linkage analysis using rare variants to map the disease locus to a single 1.7 MB interval on chromosome 4q35.2 with a LOD score of 3.2. We then used ultra-long read genome sequencing as a single molecular test to genotype a pathogenic FSHD allele containing a 4qA permissive haplotype and 5 KpnI repeat units at the D4Z4 locus. These results demonstrate that genome-wide rare variant-based linkage analysis is a powerful tool for mapping disease loci in families and ultra-long read genome sequencing is capable of genotyping pathogenic FSHD1 alleles.
Single-molecule studies reveal the off-pathway elemental pause state as a target of streptolydigin inhibition of RNA polymerase and its dramatic enhancement by Gre factors
Authors: Arseniev Anatolii , Panfilov Mikhail , Pobegalov Georgii , Potyseva Alina , Pavlinova Polina , Yakunina Maria , Lee Jookyung , Borukhov Sergei , Severinov Konstantin , Khodorkovskii Mikhail
Published: June 2023

Antibiotic streptolydigin (Stl) inhibits bacterial transcription by blocking the trigger loop folding in the active center of RNA polymerase (RNAP), which is essential for catalysis. We use acoustic force spectroscopy to characterize the dynamics of transcription elongation in ternary elongation complexes of RNAP (ECs) in the presence of Stl at a single-molecule level. We found that Stl induces long-lived stochastic pauses while the instantaneous velocity of transcription between the pauses is unaffected. Stl enhances the short-lived pauses associated with an off-pathway elemental paused state of the RNAP nucleotide addition cycle. Unexpectedly, we found that transcript cleavage factors GreA and GreB, which were thought to be Stl competitors, do not alleviate the streptolydigin-induced pausing; instead, they synergistically increase transcription inhibition by Stl. This is the first known instance of a transcriptional factor enhancing antibiotic activity. We propose a structural model of the EC-Gre-Stl complex that explains the observed Stl activities and provides insight into possible cooperative action of secondary channel factors and other antibiotics binding at the Stl-pocket. These results offer a new strategy for high-throughput screening for prospective antibacterial agents.
Trading acyls and swapping sugars: metabolic innovations inSolanumtrichomes
Authors: Fiesel Paul D. , Kerwin Rachel E. , Jones A. Daniel , Last Robert L.
Published: June 2023

Solanaceae (nightshade family) species synthesize a remarkable array of clade- and tissue-specific specialized metabolites. Protective acylsugars, one such class of structurally diverse metabolites, are produced by AcylSugar AcylTransferases from sugars and acyl-coenzyme A esters. Published research revealed trichome acylsugars composed of glucose and sucrose cores in species across the family. In addition, acylsugars were analyzed across a small fraction of the >1200 species in the phenotypically megadiverseSolanumgenus, with a handful containing inositol and glycosylated inositol cores. The current study sampled several dozen species across subclades of theSolanumto get a more detailed view of acylsugar chemodiversity. In depth characterization of acylsugars from the Clade II speciesSolanum melongena(brinjal eggplant) led to the identification of eight unusual structures with inositol or inositol glycoside cores, and hydroxyacyl chains. Liquid chromatography-mass spectrometry analysis of 31 additional species in theSolanumgenus revealed striking acylsugar diversity with some traits restricted to specific clades and species. Acylinositols and inositol-based acyldisaccharides were detected throughout much of the genus. In contrast, acylglucoses and acylsucroses were more restricted in distribution. Analysis of tissue-specific transcriptomes and interspecific acylsugar acetylation differences led to the identification of theS. melongenaAcylSugar AcylTransferase 3-Like 1 (SmASAT3-L1; SMEL4.1_12g015780) enzyme. This enzyme is distinct from previously characterized acylsugar acetyltransferases, which are in the ASAT4 clade, and appears to be a functionally divergent ASAT3. This study provides a foundation for investigating the evolution and function of diverseSolanumacylsugar structures and harnessing this diversity in breeding and synthetic biology.
Engineering the Maize Root Microbiome: A Rapid MoClo Toolkit and Identification of Potential Bacterial Chassis for studying Plant-Microbe Interactions
Authors: Schaik John van , Li Zidan , Cheadle John , Crook Nathan
Published: June 2023

Sustainably enhancing crop production is a necessity given the increasing demands for staple crops and their associated carbon/nitrogen inputs. Plant-associated microbiomes offer one avenue for addressing this demand; however, studying these communities and engineering them has remained a challenge due to limited genetic tools and methods. In this work, we detail the development of the Maize Root ToolKit (MRTK); a rapid Modular Cloning (MoClo) toolkit that only takes 2.5 hours to generate desired constructs (5400 potential plasmids) that replicate and express heterologous genes inEnterobacter ludwigiistrain AA4 (Elu),Pseudomonas putidaAA7 (Ppu),Herbaspirillum robiniaestrain AA6 (Hro),Stenotrophomonas maltophiliastrain AA1 (Sma) andBrucella pituitosastrain AA2 (Bpi) which comprise a model maize root synthetic community (SynCom). In addition to these genetic tools, we describe a highly efficient transformation protocol (10^7-10^9 transformants/µg of DNA) for each of these strains. Utilizing this highly efficient transformation protocol, we identified endogenous expression sequences for each strain (ES; promoter and ribosomal binding sites) via genomic promoter trapping. Overall, the MRTK is a scalable platform that expands the genetic engineering toolbox while providing a standardized, high efficiency transformation method that can be implemented across a diverse group of root commensals. These results unlock the ability to elucidate and engineer plant-microbe interactions promoting plant growth for each of the 5 bacterial strains in this study.
Illuminating Dark Proteins using Reactome Pathways
Authors: Brunson Timothy , Sanati Nasim , Matthews Lisa , Haw Robin , Beavers Deidre , Shorser Solomon , Sevilla Cristoffer , Viteri Guilherme , Conley Patrick , Rothfels Karen , Hermjakob Henning , Stein Lincoln , D’Eustachio Peter , Wu Guanming
Published: June 2023

Limited knowledge about a substantial portion of protein coding genes, known as “dark” proteins, hinders our understanding of their functions and potential therapeutic applications. To address this, we leveraged Reactome, the most comprehensive, open source, open-access pathway knowledgebase, to contextualize dark proteins within biological pathways. By integrating multiple resources and employing a random forest classifier trained on 106 protein/gene pairwise features, we predicted functional interactions between dark proteins and Reactome-annotated proteins. We then developed three scores to measure the interactions between dark proteins and Reactome pathways, utilizing enrichment analysis and fuzzy logic simulations. Correlation analysis of these scores with an independent single-cell RNA sequencing dataset provided supporting evidence for this approach. Furthermore, systematic natural language processing (NLP) analysis of over 22 million PubMed abstracts and manual checking of the literature associated with 20 randomly selected dark proteins reinforced the predicted interactions between proteins and pathways. To enhance the visualization and exploration of dark proteins within Reactome pathways, we developed the Reactome IDG portal, deployed athttps://idg.reactome.org, a web application featuring tissue-specific protein and gene expression overlay, as well as drug interactions. Our integrated computational approach, together with the user-friendly web platform, offers a valuable resource for uncovering potential biological functions and therapeutic implications of dark proteins.
Systematic annotation of a complete adult maleDrosophilanerve cord connectome reveals principles of functional organisation
Authors: Marin Elizabeth C. , Morris Billy J. , Stürner Tomke , Champion Andrew S. , Krzeminski Dominik , Badalamente Griffin , Gkantia Marina , Dunne Christopher R. , Eichler Katharina , Takemura Shin-ya , Tamimi Imaan F. M. , Fang Siqi , Moon Sung Soo , Cheong Han S. J. , Li Feng , Schlegel Philipp , Ahnert Sebastian E. , Berg Stuart ,   Janelia FlyEM Project Team, Card Gwyneth M. , Costa Marta , Shepherd David , Jefferis Gregory S.X.E.
Published: June 2023

Nervous systems function as ensembles of neurons communicating via synaptic connections, and a functional understanding of nervous systems requires extensive knowledge of their connectomes. In a companion paper (Takemura et al., 2023), we describe the acquisition of a complete fruit fly nerve cord connectome, the first for an animal that can walk or fly. Here, to efficiently navigate and to appreciate the biological significance of this connectome, we categorise and name nearly all neurons systematically and link them to the experimental literature. We employ a system of hierarchical coarse annotations and group similar neurons across the midline and across segments, then define systematic cell types for sensory neurons, intrinsic neurons, ascending neurons, and non-motor efferent neurons. Stereotyped arrays of neuroblasts generate related neuron populations called hemilineages that repeat across the segments of the nerve cord. We confirm that larval-born neurons from a given hemilineage generally express the same neurotransmitter but find that earlier born neurons often express a different one. We match over 35% of intrinsic, ascending, and non-motor efferent neurons across segments, defining serial sets which were crucial for systematic typing of motor neurons and sensory neurons. We assign a sensory modality to over 5000 sensory neurons, cluster them by connectivity, and identify serially homologous cell types and a layered organisation likely corresponding to peripheral topography. Finally, we present selected examples of sensory circuits predicated on programmatic analysis of a complete VNC connectome. Our annotations are critical for analysing the structure of descending input to the nerve cord and of motor output, both described in a third companion paper (Cheong et al., 2023). These annotations are being released as part of theneuprint.janelia.organdclio.janelia.orgweb applications and also serve as the basis for programmatic analysis of the connectome through dedicated tools that we describe in this paper.
Natural Silencing of Quorum-Sensing Activity ProtectsVibrio parahaemolyticusfrom Lysis by an Autoinducer-Detecting Phage
Authors: Duddy Olivia P. , Silpe Justin E. , Fei Chenyi , Bassler Bonnie L.
Published: June 2023

Quorum sensing (QS) is a chemical communication process that bacteria use to track population density and orchestrate collective behaviors. QS relies on the production, accumulation, and group-wide detection of extracellular signal molecules called autoinducers. Vibriophage 882 (phage VP882), a bacterial virus, encodes a homolog of theVibrioQS receptor-transcription factor, called VqmA, that monitors theVibrioQS autoinducer DPO. Phage VqmA binds DPO at high host-cell density and activates transcription of the phage geneqtip. Qtip, an antirepressor, launches the phage lysis program. Phage-encoded VqmA when bound to DPO also manipulates host QS by activating transcription of the host genevqmR. VqmR is a small RNA that controls downstream QS target genes. Here, we sequenceVibrio parahaemolyticusstrain O3:K6 882, the strain from which phage VP882 was initially isolated. The chromosomal region normally encodingvqmRandvqmAharbors a deletion encompassingvqmRand a portion of thevqmApromoter, inactivating that QS system. We discover thatV. parahaemolyticusstrain O3:K6 882 is also defective in its other QS systems, due to a mutation inluxO, encoding the central QS transcriptional regulator LuxO. Both thevqmR-vqmAandluxOmutations lockV. parahaemolyticusstrain O3:K6 882 into the low-cell density QS state. Reparation of the QS defects inV. parahaemolyticusstrain O3:K6 882 promotes activation of phage VP882 lytic gene expression and LuxO is primarily responsible for this effect. Phage VP882-infected QS-competentV. parahaemolyticusstrain O3:K6 882 cells lyse more rapidly and produce more viral particles than the QS-deficient parent strain. We propose that, inV. parahaemolyticusstrain O3:K6 882, constitutive maintenance of the low-cell density QS state suppresses the launch of the phage VP882 lytic cascade, thereby protecting the bacterial host from phage-mediated lysis.
A mutation-mediated host shift drives Avian paramyxovirus type 1 evolution
Authors: Chang Han , Feng Shengyong , Tang Xiaolu , Wang Ziming , Su Wen , Han Shuyi , Li Guanghao , Hu Bin , Jing Shengfan , Wang Bo , Xiang Chen , Xing Yanan , Wang Ye , Lu Jian , He Hongxuan
Published: June 2023

PPMV-1, an antigenic variant of APMV-1, associated with specific pigeon host species. However, its evolutionary strategy and underlying drivers of host specificity remain unknown. In this study, we collect the outbreak data on a global scale to investigate its evolutionary dynamics, and provide an evidence-supported analysis the host shift of PPMV-1 from chickens to pigeons, and this shift is driven by the P protein. Our data indicated that the viruses in the United States and China have undergone convergent evolution. We find that three mutations of P protein, especially R163G, can significantly affect the adaptation of APMV-1 in pigeons. Mechanistically, sensor LSm14A inhibits the replication APMV-1 in DF-1 cells, and R163G substitutionon P protein increase LSm14A degradation. We propose the host shift drive the evolution of PPMV-1 and the underlying mechanism, offering new insights into the adaptive evolutionary process of the virus.
Broaden the application ofYarrowia Lipolyticasynthetic biology tools to explore the potential of Yarrowia clade biodiversity
Authors: Park Young-Kyoung , Rossignol Tristan
Published: June 2023

Yeasts have established themselves as prominent microbial cell factories, and the availability of synthetic biology tools has led to breakthroughs in the rapid development of industrial chassis strains. The selection of a suitable microbial host is critical in metabolic engineering applications, but it has been largely limited to a few well-defined strains. However, there is growing consideration for evaluating strain diversity, as a wide range of specific traits and phenotypes have been reported even within a specific yeast genus or species. Moreover, with the advent of synthetic biology tools, non-type strains can now be easily and swiftly reshaped.

The yeastYarrowia lipolyticahas been extensively studied for various applications such as fuels, chemicals, and food. Additionally, other members of the Yarrowia clade are currently being evaluated for their industrial potential. In this study, we demonstrate the versatility of synthetic biology tools originally developed forY. lipolyticaby repurposing them for engineering other yeasts belonging to the Yarrowia clade. Leveraging the GoldenGateY. lipolyticatool kit, we successfully expressed fluorescent proteins as well as the carotenoid pathway in at least six members of the clade, serving as proof of concept.

This research lays the foundation for conducting more comprehensive investigations into the under-characterized strains within the Yarrowia clade and exploring their potential applications in biotechnology.
Stable Structures or poly(A)-binding protein loading protect cellular and viral RNAs against ISG20-mediated decay
Authors: Louvat Camille , Deymier Séverine , Nguyen Xuan-Nhi , Labaronne Emmanuel , Noy Kodie , Cariou Marie , Corbin Antoine , Mateo Mathieu , Ricci Emiliano P. , Fiorini Francesca , Cimarelli Andrea
Published: June 2023

ISG20 is an interferon-induced 3’-to-5’ RNA exonuclease that acts as a broad antiviral factor. At present, the features that expose RNA to ISG20 remain unclear, although recent studies have pointed to the modulatory role of epitranscriptomic modifications in the susceptibility of target RNAs to ISG20. These findings raise the question as to how cellular RNAs, on which these modifications are abundant, cope with ISG20. To obtain an unbiased perspective on this topic, we used RNAseq and biochemical assays to identify elements that regulate the behavior of RNAs against ISG20. The results we have obtained indicate that poly(A)-binding protein (PABP1) loading on the RNA 3’ tail provides a primal protection against ISG20, easily explaining the overall protection of cellular mRNAs observed by RNAseq. The second element we uncovered is provided by terminal stem-loop RNA structures, that have been associated to ISG20 protection before, but that we re-examine here systematically to define the stabilities that tilt the balance between resistance and susceptibility to ISG20. Given that these elements are present on cellular mRNAs, but can be co-opted by viruses as well, these results shed new light on the complex interplay that regulates the susceptibility of different classes of viruses against ISG20.
Enhanced fungal specificity andin vivotherapeutic efficacy of a C-22 modified FK520 analog againstC. neoformans
Authors: Rivera Angela , Lim Won Young , Park Eunchong , Dome Patrick A. , Hoy Michael J. , Spasojevic Ivan , Sun Sheng , Averette Anna Floyd , Pina-Oviedo Sergio , Juvvadi Praveen R. , Steinbach William J. , Ciofani Maria , Hong Jiyong , Heitman Joseph
Published: June 2023

Fungal infections are of mounting global concern, and the current limited treatment arsenal poses challenges when treating such infections. In particular, infections byCryptococcus neoformansare associated with high mortality, emphasizing the need for novel therapeutic options. Calcineurin is a protein phosphatase that mediates fungal stress responses, and calcineurin inhibition by the natural product FK506 blocksC. neoformansgrowth at 37°C. Calcineurin is also required for pathogenesis. However, because calcineurin is conserved in humans, and inhibition with FK506 results in immunosuppression, the use of FK506 as an anti-infective agent is precluded. We previously elucidated the structures of multiple fungal calcineurin-FK506-FKBP12 complexes and implicated the C-22 position on FK506 as a key point for differential modification of ligand inhibition of the mammalian versus fungal target proteins. Throughin vitroantifungal and immunosuppressive testing of FK520 (a natural analog of FK506) derivatives, we identified JH-FK-08 as a lead candidate for further antifungal development. JH-FK-08 exhibited significantly reduced immunosuppressive activity and both reduced fungal burden and prolonged survival of infected animals. JH-FK-08 exhibited additive activity in combination with fluconazolein vivo. These findings further advance calcineurin inhibition as an antifungal therapeutic approach.

Importance

Fungal infections cause significant morbidity and mortality globally. The therapeutic armamentarium against these infections is limited and development of antifungal drugs has been hindered by the evolutionary conservation between fungi and the human host. With rising resistance to the current antifungal arsenal and an increasing at-risk population, there is an urgent need for the development of new antifungal compounds. The FK520 analogs described in this study display potent antifungal activity as a novel class of antifungals centered on modifying an existing orally-active FDA approved therapy. This research advances the development of much needed newer antifungal treatment options with novel mechanisms of action.
A structural competition involving WDR5 times circadian oscillations
Authors: Börgel Arne , Rawleigh Audrey , Conrady Marcel , Hof Florian , Ricci Kyra , Brown Steven , Wolf Eva
Published: June 2023

In the mammalian circadian clock, the transcription factor BMAL1/CLOCK cycles between an active state recruiting co-activators like MLL1, and repressed states associated with the clock proteins Period1/2 (PER1/2) and Cryptochrome1/2 (CRY1/2). The MLL1 complex component WDR5 was also found in a repressive PER complex, but the roles of WDR5-PER interactions are unknown. Here we show that WDR5 directly binds to the C-terminal CRY binding domain (CBD) regions of PER1 and PER2. PER2 binds WDR5 via a WDR5 binding (WBM) motif within the PER2/CRY interface, imposing a molecular choice between PER2/WDR5- and PER2/CRY complexes. PER1 predominantly binds WDR5 via a WDR5 interacting (WIN) motif outside the CBD and exhibits a 15-fold higher WDR5 affinity than PER2. Thereby PER1 can form trimeric PER1WIN/WDR5/RbBP5WBM- and PER1/WDR5/CRY complexes as potential transition states between activating MLL1WIN/WDR5/RbBP5WBM- and repressive PER/CRY complexes. Overexpressing WDR5 in mammalian cells increases the circadian amplitude, whereas a compound targeting the WIN motif binding site of WDR5 weakens PER1-WDR5 interactions and shortens the circadian oscillation period by 2 to 3 hours. Together, our studies uncover WDR5 as direct PER interaction partner at the interface between active- and repressed states of BMAL1/CLOCK and suggest a functional role of WDR5 and its WIN site interactions in the mammalian clock by both enhancing circadian oscillations and creating a temporal delay.
Functional characterization of helminth-associated Clostridiales reveals covariates of Treg differentiation
Authors: Sargsian Shushan , Lejeune Alannah , Ercelen Defne , Jin Wen-Bing , Varghese Alan , Loke P’ng , Lim Yvonne A. L. , Guo Chun-Jun , Cadwell Ken
Published: June 2023

Parasitic helminths influence the composition of the gut microbiome. However, the microbiomes of individuals living in helminth-endemic regions are understudied. The Orang Asli, an indigenous population in Malaysia with high burdens of the helminthTrichuris trichiura, displayed microbiotas enriched in Clostridiales, an order of spore-forming obligate anaerobes previously shown to have immunogenic properties. We previously isolated novel Clostridiales that were enriched in these individuals and found that a subset promoted theTrichurislife cycle. Here, we further characterized the functional properties of these bacteria. Enzymatic and metabolomic profiling revealed a range of activities associated with metabolism and host response. Consistent with this finding, monocolonization of mice with individual isolates identified bacteria that were potent inducers of regulatory T cell (Treg) differentiation in the colon. Comparisons between variables revealed by these studies identified enzymatic properties correlated with Treg induction andTrichurisegg hatching. These results provide functional insights into the microbiotas of an understudied population.
A Connectome of the MaleDrosophilaVentral Nerve Cord
Authors: Takemura Shin-ya , Hayworth Kenneth J , Huang Gary B , Januszewski Michal , Lu Zhiyuan , Marin Elizabeth C , Preibisch Stephan , Xu C Shan , Bogovic John , Champion Andrew S , Cheong Han SJ , Costa Marta , Eichler Katharina , Katz William , Knecht Christopher , Li Feng , Morris Billy J , Ordish Christopher , Rivlin Patricia K , Schlegel Philipp , Shinomiya Kazunori , Stürner Tomke , Zhao Ting , Badalamente Griffin , Bailey Dennis , Brooks Paul , Canino Brandon S , Clements Jody , Cook Michael , Duclos Octave , Dunne Christopher R , Fairbanks Kelli , Fang Siqi , Finley-May Samantha , Francis Audrey , George Reed , Gkantia Marina , Harrington Kyle , Hopkins Gary Patrick , Hsu Joseph , Hubbard Philip M , Javier Alexandre , Kainmueller Dagmar , Korff Wyatt , Kovalyak Julie , Krzemiński Dominik , Lauchie Shirley A , Lohff Alanna , Maldonado Charli , Manley Emily A , Mooney Caroline , Neace Erika , Nichols Matthew , Ogundeyi Omotara , Okeoma Nneoma , Paterson Tyler , Phillips Elliott , Phillips Emily M , Ribeiro Caitlin , Ryan Sean M , Rymer Jon Thomson , Scott Anne K , Scott Ashley L , Shepherd David , Shinomiya Aya , Smith Claire , Smith Natalie , Suleiman Alia , Takemura Satoko , Talebi Iris , Tamimi Imaan FM , Trautman Eric T , Umayam Lowell , Walsh John J , Yang Tansy , Rubin Gerald M , Scheffer Louis K , Funke Jan , Saalfeld Stephan , Hess Harald F , Plaza Stephen M , Card Gwyneth M , Jefferis Gregory SXE , Berg Stuart
Published: June 2023

Animal behavior is principally expressed through neural control of muscles. Therefore understanding how the brain controls behavior requires mapping neuronal circuits all the way to motor neurons. We have previously established technology to collect large-volume electron microscopy data sets of neural tissue and fully reconstruct the morphology of the neurons and their chemical synaptic connections throughout the volume. Using these tools we generated a dense wiring diagram, or connectome, for a large portion of theDrosophilacentral brain. However, in most animals, including the fly, the majority of motor neurons are located outside the brain in a neural center closer to the body, i.e. the mammalian spinal cord or insect ventral nerve cord (VNC). In this paper, we extend our effort to map full neural circuits for behavior by generating a connectome of the VNC of a male fly.
An obligate intracellular bacterial pathogen forms a direct, interkingdom membrane contact site
Authors: Acevedo-Sánchez Yamilex , Woida Patrick J. , Kraemer Stephan , Lamason Rebecca L.
Published: June 2023

Interorganelle communication regulates cellular homeostasis through the formation of tightly-associated membrane contact sites1–3. Prior work has identified several ways that intracellular pathogens alter contacts between eukaryotic membranes4–6, but there is no existing evidence for contact sites spanning eukaryotic and prokaryotic membranes. Here, using a combination of live-cell microscopy and transmission and focused-ion-beam scanning electron microscopy, we demonstrate that the intracellular bacterial pathogenRickettsia parkeriforms a direct membrane contact site between its bacterial outer membrane and the rough endoplasmic reticulum (ER), with tethers that are approximately 55 nm apart. Depletion of the ER-specific tethers VAPA and VAPB reduced the frequency of rickettsia-ER contacts, suggesting these interactions mimic organelle-ER contacts. Overall, our findings illuminate a direct, interkingdom membrane contact site uniquely mediated by rickettsia that seems to mimic traditional host MCSs.