bluuebunny/crossref_metadata_2025 huggingface dataset:
Dynamic alterations in gene co-expression networks and gene-transcript associations characterize co-morbidities in cocaine use disorder
Authors: Nwaneshiudu Chinwe , Girdhar Kiran , Kleopoulos Steven P. , Fullard John F. , Butelman Eduardo R. , Parvaz Muhammad A. , Goldstein Rita Z. , Alia-Klein Nelly , Roussos Panos
Published: July 2024

Background

Individuals with cocaine use disorder (CUD) who attempt abstinence experience craving and relapse, which poses challenges in treatment. Longitudinal studies linking behavioral manifestations in CUD to the blood transcriptome in living individuals are limited. Therefore, we investigated the connection between drug use behaviors during abstinence with blood transcriptomics.

Methods

We conducted a comprehensive longitudinal study involving 12 subjects (9 males, 3 females) with CUD and RNA sequencing on blood collected at a drug-free baseline, and 3, 6 & 9 months thereafter. We categorized subjects into 2 responder groups (high-low) based on scores of drug use variables, and 3 responder groups (low-intermediate-high) on days of abstinence. We investigated differential expression and gene-transcript associations across responder groups at each time point. Lastly, we examined genes that are both co-expressed and showed dynamic expression with time.

Results

Genes with significant transcript associations between high and. intermediate days of abstinence at 9 months were notably enriched for cannabis use disorder, drinks weekly, and coronary artery disease risk genes. Time-specific gene co-expression analysis prioritized transcripts related to immune processes, cell cycle, RNA-protein synthesis, and second messenger signaling for days of abstinence.

Conclusion

We demonstrate that abstinence reflects robust changes in drug use behaviors and the blood transcriptome in CUD. We also highlight the importance of longitudinal studies to capture complex biological processes during abstinence in CUD.
A long non-coding RNA LINC00094 regulates the transcriptional expression of lipid metabolism-related genes as a new member of core regulatory circuitry in esophageal squamous cell carcinoma
Authors: Peng Liu , Wang Qiu-Yu , Chen Jia-Xin , Chen Yang , Li Rong-Yao , Liao Lian-Di , Lin Wan , Li Chun-Quan , Li En-Min , Xu Li-Yan
Published: July 2024

LINC00094 as a new supper-enhancer (SE)-related long non-coding RNA is associated with poor overall survival of patients with esophageal squamous cell carcinoma (ESCC). However, the transcriptional regulatory mechanism of LINC00094 and the molecular mechanisms by which LINC00094 affects the phenotype of ESCC remains unclear. Here, we found that LINC00094 promoted the proliferation of ESCC cells both in vitro and in vivo. LINC00094 knockdown significantly reduced the expression profiles of transcription activators including transcription factor 3 (TCF3) and Kruppel like factor 5 (KLF5) and lipid metabolism-related genes. Mechanically, TCF3 and KLF5 formed a core regulatory circuitry (CRC) that bound to the SEs of LINC00094 and to their own SEs to regulate the transcriptional expression in a positive feedback loop. LINC00094 recruited TCF3 and KLF5 to form a ternary complex, which forms a new CRC with TCF3 and KLF5 that regulated its own transcription as well as lipid metabolism-related genes. Knockdown of any or all three genes inhibited the expression of genes related to lipid synthesis and consistently reduced total lipid droplet levels. Treatment with SEs inhibitors (THZ1 and JQ1) effectively inhibited the formation of this CRC and the production of lipid droplets in ESCC cells. The high-risk group of CRC-associated signatures were closely associated with poor prognosis in patients with ESCC. Our findings suggest that LINC00094 is involved in the CRC by forming a complex with TCF3 and KLF5, and this regulation model can affect the phenotype of ESCC cells by controlling the expression of lipid metabolism-related genes.

Highlights

We identified a novel functional lncRNA-LINC00094 for esophageal squamous cell carcinoma.

LINC00094 forms a complex with the core transcription factors TCF3 and KLF5, thereby forming a core regulatory circuitry to participate in transcriptional regulation in ESCC.

A core regulatory circuitry mediated by LINC00094 regulates lipid metabolism in ESCC.
CryoJAM: Automating Protein Homolog Fitting in Medium Resolution Cryo-EM Density Maps
Authors: Carrion Jackson , Manjrekar Mrunali , Mikulevica Anna
Published: July 2024

Obtaining atomic structures of large protein complexes from medium-resolution cryogenic electron-microscopy (cryo-EM) density maps is a critical bottleneck in the cryo-EM workflow. CryoJAM aims to automate this process by using a 3D Convolutional Neural Network model within a U-Net architecture. This model is trained on a novel loss function that leverages Fourier-Shell Correlation (FSC), as a proxy for quality of fit, and Root Mean Squared Error (RMSE) to help optimize fits within real space. Capitalizing on the gold-standard status of FSC in cryo-EM, this method introduces an innovative implementation of FSC into cryo-EM model fitting software, enhancing the precision and efficiency of structural analysis. After 25 epochs, CryoJAM successfully reduced the RMSE in 21 out of 26 of the test cases, effectively fitting homologous protein structures into medium-resolution cryo-EM densities.
Contrasting Kinetics of Highly Similar Chloroalkane Reductive Dehalogenases
Authors: Picott Katherine J. , Edwards Elizabeth A.
Published: July 2024

Chloroform and trichloroethanes are pervasive groundwater contaminants for which bioremediation has been an effective treatment strategy. Reductive dehalogenase (RDase) enzymes from organohalide-respiring bacteria are essential for their remediation under anaerobic conditions. RDases are responsible for dehalogenating these chlorinated solvents, leading to their removal. This work explores the kinetic characteristics of three closely relatedDehalobacterchloroalkane-reductases—TmrA, CfrA, and AcdA—and identifies differences between their activity on chloroform (CF), 1,1,1-trichloroethane (TCA), and 1,1,2-TCA. The side-by-side comparison of these enzymes has emphasized that TmrA and AcdA are specialized toward CF with both having 4-fold higher maximum specific activity (Vmax) on CF than 1,1,1-TCA, whereas, CfrA has very similar rates on both CF and 1,1,1-TCA. AcdA is the most sensitive to substrate inhibition by CF and 1,1,2-TCA, and inhibition by a common co-contaminant trichloroethene. Finally, the reduction of 1,1,2-TCA, which can produce both 1,2-dichloroethane and vinyl chloride, was assessed for each enzyme. Interestingly, each enzyme has a distinct preference for the major product it produces, indicating a favoured reaction pathway. Despite over 95% sequence identity, TmrA, CfrA, and AcdA exhibit substantial differences in kinetic behaviour, highlighting the importance of understanding such nuances for informed bioremediation strategies.

SYNOPSIS

Three similar dechlorinating enzymes were found to have distinct reaction rates and levels of sensitivity to inhibition. These differences have implications for the enzymes’ use in bioremediation.
Proteostasis Landscapes of Cystic Fibrosis Variants Reveals Drug Response Vulnerability
Authors: McDonald Eli Fritz , Kim Minsoo , Olson John A. , Meiler Jens , Plate Lars
Published: July 2024

Cystic Fibrosis (CF) is a lethal genetic disorder caused by variants in CF transmembrane conductance regulator (CFTR). Many variants are treatable with correctors, which enhance the folding and trafficking of CFTR. However, approximately 3% of persons with CF harbor poorly responsive variants. Here, we used affinity purification mass spectrometry proteomics to profile the protein homeostasis (proteostasis) changes of CFTR variants during correction to assess modulated interactions with protein folding and maturation pathways. Responsive variant interactions converged on similar proteostasis pathways during correction. In contrast, poorly responsive variants subtly diverged, revealing a partial restoration of protein quality control surveillance and partial correction. Computational structural modeling showed that corrector VX-445 failed to confer enough NBD1 stability to poor responders. NBD1 secondary stabilizing mutations rescued poorly responsive variants, revealing structural vulnerabilities in NBD1 required for treating poor responders. Our study provides a framework for discerning the underlying protein quality control and structural defects of CFTR variants not reached with existing drugs to expand therapeutics to all susceptible CFTR variants.

SIGNIFICANCE STATEMENT

Cystic Fibrosis (CF) is a lethal genetic disease with variants leading to misfolding of an anion channel protein. Enhancing productive channel folding using a novel class of small molecules calledcorrectorshas emerged as the current CF treatment paradigm. However, correctors fail to reach all patient variants. Using high throughput interactomics, Rosetta simulations, and biochemical trafficking assays, this study demonstrates poorly responsive CF variants experience diverse misfolding pathways caused by structural defects in the core of a nucleotide-binding domain. Stabilizing secondary mutations in this domain rescues poorly responsive variants, paving the way for mechanistic-based therapeutic development for untreatable CF variants and future protein misfolding corrector drugs.

COMPETING INTERESTS

The authors declare no competing interests.
A shared threat-anticipation circuit is dynamically engaged at different moments by certain and uncertain threat
Authors: Cornwell Brian R. , Didier Paige R. , Grogans Shannon E. , Anderson Allegra S. , Islam Samiha , Kim Hyung Cho , Kuhn Manuel , Tillman Rachael M. , Hur Juyoen , Scott Zachary S. , Fox Andrew S. , DeYoung Kathryn A. , Smith Jason F. , Shackman Alexander Joseph
Published: July 2024

Temporal dynamics play a central role in models of emotion: "fear" is widely conceptualized as a phasic response to certain-and-imminent danger, whereas "anxiety" is a sustained response to uncertain-or-distal harm. Yet the underlying neurobiology remains contentious. Leveraging a translationally relevant fMRI paradigm and theory-driven modeling approach in 220 adult humans, we demonstrate that certain- and uncertain-threat anticipation recruit a shared circuit that encompasses the central extended amygdala (EAc), periaqueductal gray, midcingulate, and anterior insula. This circuit exhibits persistently elevated activation when threat is uncertain and distal, and transient bursts of activation just before certain encounters with threat. Although there is agreement that the EAc plays a critical role in orchestrating responses to threat, confusion persists about the respective contributions of its major subdivisions, the bed nucleus of the stria terminalis (BST) and central nucleus of the amygdala (Ce). Here we used anatomical regions-of-interest to demonstrate that the BST and Ce exhibit statistically indistinguishable threat dynamics. Both regions exhibited activation dynamics that run counter to popular models, with the Ce showing sustained responses to uncertain-and-distal threat and the BST showing phasic responses to certain-and-imminent threat. For many scientists, feelings are the hallmark of fear and anxiety. Here we used an independently validated multivoxel brain signature to covertly probe the moment-by-moment dynamics of anticipatory distress for the first time. Results mirrored the dynamics of neural activation. These observations provide fresh insights into the neurobiology of threat-elicited emotions and set the stage for more ambitious clinical and mechanistic research.
GeneticSNUPNvariants cause spinocerebellar atrophy by disrupting global splicing in Purkinje cells
Authors: Okubo Mariko , Ogawa Megumu , Eura Nobuyuki , Inoue Yukiko U. , Dewa Ken-ichi , Owa Tomohiro , Miyashita Satoshi , Murakami Terumi , Nakamura Hisayoshi , Hayashi Shinichiro , Nonaka Ikuya , Ogata Katsuhisa , Hoshino Mikio , Inoue Takayoshi , Nishino Ichizo , Noguchi Satoru
Published: July 2024

We identified genetic variants in theSNUPNgene, which encodes the adapter protein snurportin-1 for the nuclear import of U1 snRNPs, in two families affected by spinocerebellar ataxia. We have elucidated the pathogenicity of these variants and the molecular pathomechanisms underlying this disease by assessing mutant snurportin-1 propertiesin vitro, cerebella at the morphological and molecular levelsex vivo, and motor functions inSnupn-variant knocked-in micein vivo. Mutant snurportin-1 impaired nuclear-cytosol shuttling, leading to defective nuclear transport of U1 snRNPs in cerebellar Purkinje cells. This resulted in aberrant splicing and expression of genes essential for Purkinje cell development and impaired dendrite formation. The malformation of Purkinje cell dendrites resulted in hypoplasia and premature migration of granule cell precursors and interneurons, leading to abnormal lobe development and atrophy in the cerebellum.
Multimodal and Quantitative Analysis of the Epileptogenic Zone in the Pre-Surgical Evaluation of Drug-Resistant Focal Epilepsy
Authors: Karimi-Rouzbahani Hamid , Vogrin Simon , Cao Miao , Plummer Chris , McGonigal Aileen
Published: July 2024

Surgical resection for epilepsy often fails due to incomplete Epileptogenic Zone (EZ) localization from standard electroencephalography (EEG), stereo-EEG (SEEG), and Magnetic Resonance Imaging (MRI). Subjective interpretation based on interictal, or ictal recordings limits conventional EZ localization. This study employs multimodal analysis using high-density-EEG (HDEEG), Magnetoencephalography (MEG), functional-MRI (fMRI), and SEEG to overcome these limitations in a patient with drug-resistant MRI-negative focal epilepsy. A teenage boy with drug-resistant epilepsy underwent evaluation. HDEEG, MEG, fMRI, and SEEG were used, with a novel HDEEG-cap facilitating simultaneous EEG-MEG and EEG-fMRI recordings. Electrical and magnetic source imaging were performed, and fMRI data were analysed for homogenous regions. SEEG analysis involved spike detection, spike timing analysis, ictal fast activity quantification, and Granger-based connectivity analysis. Non-invasive sessions revealed consistent interictal source imaging results identifying the EZ in the right anterior cingulate cortex. EEG-fMRI highlighted broader activation in the right cingulate cortex. SEEG analysis localized spikes and fast activity in the right anterior and posterior cingulate gyri. Multi-modal analysis suggested the EZ in the right frontal lobe, primarily involving the anterior and mid-cingulate cortices. Multi-modal non-invasive analyses can optimise SEEG implantation and surgical decision-making. Invasive analyses corroborated non-invasive findings, emphasising the importance of individual-case quantitative analysis across modalities in complex epilepsy cases.
Even A Worm Will Turn: Appendicitis Malpractice Litigation Since 2020
Authors: Menshawey Rahma , Menshawey Esraa
Published: July 2024

Background

Appendicitis is the inflammation of the vermiform appendix, and it is the most common abdominal surgical emergency in the world. Its diagnosis, however, has many pitfalls including lack of a pathognomonic sign or symptoms, and the low predictive value of laboratory testing. Appendicitis is a leading cause of malpractice concerns.

Methods

Using Google Case Law, we used the search terms “appendicitis”, and “malpractice” to identify appendicitis litigation cases. We included cases published since 2020. We included any case where a confirmed diagnosis of appendicitis was made. We included cases filed for malpractice due to complications of appendicitis and its treatment. Outcomes of interest included: the state the case was published, the defendants, the date of when the patient first complained of abdominal pain to when they had had an appendectomy/treatment, diagnostics and consultations, medical and legal issues, and final verdict/opinion/decisions on the cases.

Results

A total of 44 cases were identified, which were screened for inclusion. A total of 14 cases met the inclusion criteria and were analyzed. Most cases did not present in an atypical way, the majority of patients presented with clear statements of abdominal pain of varying severity. The majority of the defendants were MD’s and hospitals. The average time from symptom to diagnosis was 2.4 ±2.1 days, while the longest time for diagnosis was 7 days. The leading medico-legal issues were failure to diagnose and delayed diagnosis, while among the cases, 35.7% had outcomes in favor of the plaintiff.

Conclusions

Appendicitis remains an area of high risk of litigation. Malpractice suits are often due to failure to diagnose and failure to treat, but there maybe proactive measures to address the modern pitfalls to promote a decreased litigation risk and patient safety
PodGPT: An audio-augmented large language model for research and education
Authors: Jia Shuyue , Bit Subhrangshu , Searls Edward , Lauber Meagan V. , Claus Lindsey A. , Fan Pengrui , Jasodanand Varuna H. , Veerapaneni Divya , Wang William M. , Au Rhoda , Kolachalama Vijaya B.
Published: July 2024

The proliferation of scientific podcasts has generated an extensive repository of audio content, rich in specialized terminology, diverse topics, and expert dialogues. Here, we introduce a computational framework designed to enhance large language models (LLMs) by leveraging this informational content from publicly accessible podcast data across science, technology, engineering, mathematics and medical (STEMM) disciplines. This dataset, comprising over 3, 700 hours of audio content, was transcribed to generate over 42 million text tokens. Our model, PodGPT, integrates this wealth of complex dialogue found in audio podcasts to improve understanding of natural language nuances, cultural contexts, as well as scientific and medical knowledge. PodGPT also employs retrieval augmented generation (RAG) on a vector database built from articles in Creative Commons PubMed Central andThe New England Journal of Medicine, enhancing STEMM research and education by providing real-time access to emerging scientific literature. Evaluated across multiple benchmarks, PodGPT demonstrated an average improvement of 3.51 percentage points over standard open-source benchmarks and 3.81 percentage points when augmented with evidence from the RAG pipeline. Moreover, it showcased an average improvement of 4.06 percentage points in its zero-shot multi-lingual transfer ability, effectively generalizing to different linguistic contexts. By harnessing the untapped potential of podcast content, PodGPT advances natural language processing and conversational AI, offering enhanced capabilities for STEMM research and education.
Screening and treatment time in school-based caries prevention: A randomized clinical trial
Authors: Godín Tamarinda J. Barry , Hawthorne Gabriel , Shah Radhika , Ruff Ryan Richard
Published: July 2024

Background

School-based caries prevention can increase access to dental services for underrepresented children and reduce the risk of tooth decay.

Methods

The CariedAway study was a longitudinal pragmatic randomized trial of silver diamine fluoride (SDF), fluoride varnish, dental sealants, and atraumatic restorations (ART) provided as part of a school caries prevention program. Using electronic health record software and reproducible procedures, we estimated the total time required to screen and treat program participants. Differences at initial treatment between interventions, provider (registered nurse and dental hygienist), dentition mix, and caries burden were determined using linear regression with cluster standard error estimation, and longitudinal effects were estimated using linear mixed effects models.

Results

A total of 7418 children were enrolled in the CariedAway trial, of which 7176 (97%) had viable data recorded for screening and treatment time. Overall treatment time for children receiving SDF and fluoride varnish was 283 seconds (SD=739), compared to 753 seconds (SD=2166) for children receiving dental sealants and ART. At the initial program visit, treatment time using SDF was significantly shorter than sealants and ART (B = −458.8, 95% CI = −650.1, −266.8) and treatment time decreased with each subsequent observation (B = −51.9, 95% CI = −68.4, −35.4). Treatment time significantly increased as the number of carious teeth per child increased, and there were no differences in treatment time using SDF between registered nurses and dental hygienists.

Conclusions

The sustainability of school-based caries prevention can be supported by robust data on program logistics and treatment time. These results can be leveraged by future school-based sealant and SDF programs to estimate the total reach and effectiveness of intended treatments.
Bacterial Protein Signatures Identified inPorphyromonas gingivalisContaining-Autophagic Vacuoles Reveal Co-Evolution Between Oral Red/Orange Complex Bacteria and Gut Bacteria
Authors: Paul Ayana , Wellslager Bridgette , Williamson Maddie , Yilmaz Özlem
Published: July 2024

Modern oral bacterial species present as a concoction of commensal and opportunistic pathogens originating from their evolution in humans. Due to the intricate colonization mechanisms shared amongst oral and gut bacteria, these bacteria have likely evolved together to establish and adapt in the human oro-digestive tract, resulting in the transfer of genetic information. Our liquid chromatography-with-tandem-mass-spectrometry (LC-MS-MS) analyses have revealed protein signatures, Elongation Factor Tu, RagB/SusD nutrient uptake outer membrane protein and DnaK, specifically fromPorphyromonas gingivalis-containing autophagic vacuoles isolated from the infected human primary gingival epithelial cells. Interestingly, our Mass-Spectrometry analysis reported similar proteins from closely related oral bacteria,Tannerella forsythiaandPrevotella intermedia. In our phylogenetic study of these key protein signatures, we have established that pathogenic oral bacteria share extensive relatedness to each other and gut resident bacteria. We show that in the virulence factors identified from gut bacteria, Elongation Factor Tu and DnaK, there are several structural similarities and conservations with proteins from oral pathogenic bacteria. There are also major similarities in the RagB/SusD proteins of oral bacteria to prominent gut bacteria. These findings not only highlight the shared virulence mechanisms amongst oral bacterial pathogens/pathobionts but also gut bacteria and elucidate their co-evolutions in the human host.
RABGAP1 acts as a sensor to facilitate sorting and processing of amyloid precursor protein
Authors: Eden Jessica , Kaufman Jonathan G.G. , Cattin-Ortola Jerome , Benedetti Lorena , Nieuwenhuis Bart , Owen David J , Lippincott-Schwartz Jennifer , Munro Sean , Gershlick David C
Published: July 2024

A key hallmark of Alzheimer's disease (AD) is the accumulation of extracellular amyloid-beta plaques in the brains of patients. Amyloid-beta is a 40-42 amino acid peptide produced by the proteolytic processing of amyloid precursor protein (APP) by a series of membrane-bound proteases. APP is a type-I transmembrane protein and thus its trafficking to encounter the proteases represents a rate-limiting step in the progression of AD. Although there has been a focused research effort to understand APP processing, its trafficking itinerary and machinery is incompletely understood. To address this we have performed an unbiased interaction screen for interactors of the cytosolic tail of APP. We identified previously characterised APP binders, as well as novel interactors. We have mapped the binding of APP to multiple new machineries, including RABGAP1. We have demonstrated that RABGAP1 partially colocalises with APP and directly interacts with a YENPTY motif in the APP cytosolic tail. Depletion or overexpression of RABGAP1 caused mistrafficking and misprocessing of endogenous APP in both human and rodent neurons. Interestingly, this effect was dependent on the GAP activity of RABGAP1, demonstrating that RABGAP1 affects the trafficking of APP by modulating the RAB activity on endosomal subdomains. This novel trafficking mechanism has implications for other NPXY cargoes and also presents a possible therapeutic avenue to explore.
Systematic characterization of indel variants using a yeast-based protein folding sensor
Authors: Larsen-Ledet Sven , Lindemose Søren , Panfilova Aleksandra , Gersing Sarah , Suhr Caroline H. , Genzor Aitana Victoria , Lanters Heleen , Nielsen Sofie V. , Lindorff-Larsen Kresten , Winther Jakob R. , Stein Amelie , Hartmann-Petersen Rasmus
Published: July 2024

Gene variants resulting in insertions or deletions of amino acid residues (indels) have important consequences for evolution and are often linked to disease, yet compared to missense variants the effects of indels are poorly understood and predicted. To approach this issue, we developed a sensitive protein folding sensor based on complementation of uracil auxotrophy in yeast by circular permutated orotate phosphoribosyltransferase (CPOP). The sensor accurately reports on the folding of disease-linked missense variants andde novodesigned proteins. Applying the folding sensor to a saturated library of single amino acid indel variants in human DHFR revealed that most regions which tolerate indels are confined to internal loops and the N- and C-termini. Surprisingly, indels are also allowed at a central α-helix. Several indels are temperature-sensitive and the folding of most of these indels is rescued upon binding to the competitive DHFR inhibitor methotrexate. Rosetta and AlphaFold2 predictions correlate with the observed effects, suggesting that most indels operate by destabilizing the native fold and that these computational tools may be useful for classification of indels observed in population sequencing.
The fate of secretory cells during intestinal homeostasis, regeneration, and tumor formation is regulated by Tcf4
Authors: Janeckova Lucie , Stastna Monika , Hrckulak Dusan , Berkova Linda , Kubovciak Jan , Onhajzer Jakub , Kriz Vitezslav , Dostalikova Stela , Mullerova Tereza , Vecerkova Katerina , Tenglerova Marketa , Coufal Stepan , Kostovcikova Klara , Blumberg Richard S , Filipp Dominik , Basler Konrad , Valenta Tomas , Kolar Michal , Korinek Vladimir
Published: July 2024

The single-layer epithelium of the gastrointestinal tract is a dynamically renewing tissue that ensures nutrient absorption, secretory and barrier functions and is involved in immune responses. The basis for this homeostatic renewal is the Wnt signaling pathway. Blocking this pathway can lead to epithelial damage, while its abnormal activation can result in the development of intestinal tumors. In this study, we investigated the dynamics of intestinal epithelial cells and tumorigenesis using a conditional mouse model. Using single-cell and bulk RNA sequencing and histological analysis, we elucidated the cellular responses following the loss of specific cell types. We focused on the fate of cells in the lower parts of the intestinal crypts and the development of colon adenomas. By partially inactivating the transcription factor Tcf4, a key effector of the Wnt signaling pathway, we analyzed the regeneration of isolated hyperproliferative foci (crypts). Our results suggest that the damaged epithelium is not restored by a specific regeneration program associated with oncofetal gene production, but rather by a standard homeostatic renewal pathway. Moreover, disruption of Tcf4 in secretory progenitors resulted in a significant shift in the cell lineage from Paneth cells to goblet cells, characterized by morphological changes and loss of Paneth cell-specific genes. We also found that hyperactivation of the Wnt signaling pathway in colonic adenomas correlated with the upregulation of genes typical of Paneth cells in the intestine, followed by the emergence of secretory tumor cells producing the Wnt3 ligand. The absence of Tcf4 led to a phenotypic shift of the tumor cells towards goblet cells. Our study presents a new model of epithelial regeneration based on the genetically driven partial elimination of intestinal crypts. We highlight the critical role of Tcf4 in the control of cell lineage decisions in the intestinal epithelium and colon tumors.
Olfactory basis for essential amino acid perception during foraging in Caenorhabditis elegans
Authors: Siddiqui Ritika , Mehta Nikita , Ranjith Gopika , Felix Marie-Anne , Chen Changchun , Singh Varsha
Published: July 2024

Acquisition of essential nutrients through diet is crucial for the survival of animals. Dietary odors might enable foraging in animals for nutrient-rich diets. We asked if Caenorhabditis elegans, a bacterivorous nematode, uses olfactory cues to forage for essential amino acid-rich (EAA) diets. Using the native microbiome of C. elegans, we show that worms rely on olfaction to select leucine (EAA)-enriched bacteria. Using gas chromatography, we find that leucine-enriched bacteria produce isoamyl alcohol (IAA) odor in the highest abundance. Prior adaptation of worms to IAA diminishes the diet preference of worms. Several wild isolates of C. elegans display robust responses to IAA emphasizing its ecological relevance. We find that foraging for a leucine-enriched diet is mediated via the AWC olfactory neurons. Finally, we identify SRD-12, in AWC neurons, as a receptor for IAA and a mediator of dietary decisions in worms. Our study identifies a receptor-ligand module underpinning foraging behavior in C. elegans.
mettannotator: a comprehensive and scalable Nextflow annotation pipeline for prokaryotic assemblies
Authors: Gurbich Tatiana A. , Beracochea Martin , De Silva Nishadi H. , Finn Robert D.
Published: July 2024

Summary: In recent years there has been a surge in prokaryotic genome assemblies, coming from both isolated organisms and environmental samples. These assemblies often include novel species that are poorly represented in reference databases creating a need for a tool that can annotate both well-described and novel taxa, and can run at scale. Here, we present mettannotator - a comprehensive Nextflow pipeline for prokaryotic genome annotation that identifies coding and non-coding regions, predicts protein functions, including antimicrobial resistance, and delineates gene clusters. The pipeline summarises the results of these tools in a GFF (General Feature Format) file that can be easily utilised in downstream analysis or visualised using common genome browsers. Mettannotator has been tested on both novel and known taxa, and used across Ensembl Bacteria and MGnify. Here, we show how it works on 29 prokaryotic phyla, both isolate and metagenome-assembled genomes, and present metrics on its performance in comparison to other tools. Availability and implementation: The pipeline is written in Nextflow and Python and published under an open source Apache 2.0 licence. Instructions and source code can be accessed at https://github.com/EBI Metagenomics/mettannotator. The pipeline is also available on WorkflowHub: https://workflowhub.eu/workflows/1069.
Malassezia responds to environmental pH signals through the conserved Rim/Pal pathway
Authors: Pianalto Kaila M. , Telzrow Calla L. , Harding Hannah Brown , Brooks Jacob T. , Granek Joshua A. , Gushiken-Ibañez Eduardo , LeibundGut-Landmann Salomé , Heitman Joseph , Ianiri Giuseppe , Alspaugh J. Andrew
Published: July 2024

During mammalian colonization and infection, microorganisms must be able to rapidly sense and adapt to changing environmental conditions including alterations in extracellular pH. The fungus-specific Rim/Pal signaling pathway is one process that supports microbial adaptation to alkaline pH. This cascading series of interacting proteins terminates in the proteolytic activation of the highly conserved Rim101/PacC protein, a transcription factor that mediates microbial responses that favor survival in neutral/alkaline pH growth conditions, including many mammalian tissues. We identified the putative Rim pathway proteins Rim101 and Rra1 in the human skin colonizing fungusMalassezia sympodialis. Gene deletion by transconjugation and homologous recombination revealed that Rim101 and Rra1 are required forM. sympodialisgrowth at higher pH. Additionally, comparative transcriptional analysis of the mutant strains compared to wild-type suggested mechanisms for fungal adaptation to alkaline conditions. These pH-sensing signaling proteins are required for optimal growth in a murine model of atopic dermatitis, a pathological condition associated with increased skin pH. Together these data elucidate both conserved and phylum-specific features of microbial adaptation to extracellular stresses.

Importance

The ability to adapt to host pH has been previously associated with microbial virulence in several pathogenic fungal species. Here we demonstrate that a fungal-specific alkaline response pathway is conserved in the human skin commensal fungusMalassezia sympodialis(Ms). This pathway is characterized by the pH-dependent activation of the Rim101/PacC transcription factor that controls cell surface adaptations to changing environmental conditions. By disrupting genes encoding two predicted components of this pathway, we demonstrated that the Rim/Pal pathway is conserved in this fungal species as a facilitator of alkaline pH growth. Moreover, targeted gene mutation and comparative transcriptional analysis supports the role of theMsRra1 protein as a cell surface pH sensor conserved within the basidiomycete fungi, a group including plant and human pathogens. Using an animal model of atopic dermatitis, we demonstrate the importance ofMsRim/Pal signaling in this common inflammatory condition characterized by increased skin pH.
Rational Design of a Flavoenzyme for Aerobic Nicotine Catabolism
Authors: Xu Ping , Tang Hongzhi , Hu Haiyang , Xu Zhaoyong , Zhang Zhiyao , Stull Frederick , Song Peizhi
Published: July 2024

Enzymatic therapy with nicotine-degrading enzyme is a new strategy in treating nicotine addiction, which can reduce nicotine concentrations and weaken withdrawal in the rat model. However, when O2is used as the electron acceptor, no satisfactory performance has been achieved with one of the most commonly studied and efficient nicotine-catabolizing enzymes, NicA2. To obtain more efficient nicotine-degrading enzyme, we rationally designed and engineered a flavoenzyme Pnao, which shares high structural similarity with NicA2 (RMSD = 1.143 Å) and efficiently catalyze pseudooxynicotine into 3-succinoyl-semialdehyde pyridine using O2. Through amino acid alterations with NicA2, five Pnao mutants were generated, which can degrade nicotine in Tris-HCl buffer and retained catabolic activity on its natural substrate. Nicotine-1'-N-oxide was identified as one of the reaction products. Four of the derivative mutants showed activity in rat serum and Trp220 and Asn224 were found critical for enzyme specificity. Our findings offer a novel avenue for research into aerobic nicotine catabolism and provides a promising method of generating additional nicotine-catalytic enzymes.
Commensal-pathogen dynamics structure disease outcomes duringClostridioides difficilecolonization
Authors: Fishbein Skye R. S. , DeVeaux Anna L. , Khanna Sakshi , Ferreiro Aura L. , Liao James , Agee Wesley , Ning Jie , Mahmud Bejan , Wallace Miranda J. , Hink Tiffany , Reske Kimberly A. , Guruge Janaki , Leekha Sidh , Dubberke Erik R. , Dantas Gautam
Published: July 2024

Gastrointestinal colonization byClostridioides difficileis common in healthcare settings and ranges in clinical presentation from asymptomatic carriage to lethalC. difficileinfection (CDI). We used a systems biology approach to investigate why patients colonized withC. difficilehave a range of outcomes. Microbiota-humanization of germ-free mice with fecal samples from toxigenic C. difficile carriers revealed a spectrum of virulence among clade 1 lineages and identified commensalBlautiaassociated with markers of non-pathogenic colonization. Using gnotobiotic mice engrafted with defined human microbiota, we observed strain-specific CDI severity across clade 1 strains. Yet, mice engrafted with a higher diversity community were protected from severe disease across all strains without suppression ofC. difficilecolonization. These results indicate that when colonization resistance has been breached without overt infection, commensals can attenuate a diversity of virulent strains without inhibiting pathogen colonization, providing insight into determinants of stableC. difficilecarriage.
Direct testing of natural twister ribozymes from over a thousand organisms reveals a broad tolerance for structural imperfections
Authors: McKinley Lauren N. , Meyer McCauley O. , Sebastian Aswathy , Chang Benjamin K. , Messina Kyle J. , Albert Istvan , Bevilacqua Philip C.
Published: July 2024

Twister ribozymes are an extensively studied class of nucleolytic RNAs. Thousands of natural twisters have been proposed using sequence homology and structural descriptors. Yet, most of these candidates have not been validated experimentally. To address this gap, we developed CHiTA (Cleavage High-Throughput Assay), a high-throughput pipeline utilizing massively parallel oligonucleotide synthesis and next-generation sequencing to test putative ribozymesen massein a scarless fashion. As proof of principle, we applied CHiTA to a small set of known active and mutant ribozymes. We then used CHiTA to test two large sets of naturally occurring twister ribozymes: over 1, 600 previously reported putative twisters and ∼1, 000 new candidate twisters. The new candidates were identified computationally in ∼1, 000 organisms, representing a massive increase in the number of ribozyme-harboring organisms. Approximately 94% of the twisters we tested were active and cleaved site-specifically. Analysis of their structural features revealed that many substitutions and helical imperfections can be tolerated. We repeated our computational search with structural descriptors updated from this analysis, whereupon we identified and confirmed the first intrinsically active twister ribozyme in mammals. CHiTA broadly expands the number of active twister ribozymes found in nature and provides a powerful method for functional analyses of other RNAs.

GRAPHICAL ABSTRACT
Brain Segregation and Integration Relate to Word-Finding Abilities in Older and Younger Adults
Authors: Oosterhuis Elise J. , Bailey Neil , Slade Kate , May Patrick J. C. , Nuttall Helen Elizabeth
Published: July 2024

Previous research has shown that word-finding difficulties in older age are associated with functional and structural brain changes. However, the use of functional brain networks, measured through electroencephalography, to predict word-finding in older and younger adults has not yet been investigated. This study utilised resting-state electroencephalography data (61 channels) from the Leipzig Study for Mind-Body-Emotion Interactions dataset (Babayan et al., 2019) to investigate the relationship between functional brain networks and word-finding ability in healthy younger and older adults. Graph theory-based measures in individualised delta, theta, alpha, and beta bands were computed to assess brain segregation and integration of 53 older (aged 59-77) and 53 younger right-handed adults (aged 20-35). Word-finding ability was quantified as the number of orally produced words during a semantic and letter fluency task. Multiple linear regression revealed that, in older adults, greater functional connectedness in the delta band was associated with lower semantic fluency. Irrespective of age, greater modularity in the alpha band was related to lower semantic fluency. A greater small-world index in the delta band was related to better semantic fluency, irrespective of age. Increased brain integration in the delta band corresponded to greater semantic fluency in older adults. Hence, word-finding ability seems to be related to brain segregation and integration specific to the frequency band, possibly indicating alterations in cognitive control or compensatory shifts to less functionally specific frequency bands. The article further provides a discussion on neural dedifferentiation, hyper-synchronisation, study limitations, and directions for future research.
Sarm1-Dependent Metabolic Reprogramming of Schwann Cells Following Nerve Injury
Authors: Stepanova Ekaterina , Cho Carol , Lee John , Pavelec Caitlin M , Tripathi Anika , Vegiraju Tanvika , Hunter-Chang Sarah , Leitinger Norbert , Campbell John Nelson , Deppmann Christopher
Published: July 2024

Schwann cells (SCs) transition into a repair phenotype after peripheral nerve injury, which is crucial for supporting axon regeneration. However, the early SC injury response preceding the repair state remains poorly understood. Here, we demonstrate that Sarm1, a key regulator of axon degeneration, is expressed in SCs and has a critical role in the early SC injury response. Leveraging the fact that Sarm1 deletion impairs the SC transition to the repair state, we used single-nucleus RNA sequencing to compare the transcriptional responses of wild-type and Sarm1 knockout SCs 24 hours after nerve injury. Remarkably, Sarm1-deficient SCs, unlike wild-type SCs, showed increased expression of genes involved in oxidative phosphorylation and the TCA cycle. These findings were functionally validated, revealing that Sarm1 knockout SCs displayed increased mitochondrial respiration in response to injury. Intriguingly, Sarm1 knockout SCs also exhibited enhanced axon protection compared to wild-type SCs in an in vitro model of axon degeneration. We propose that Sarm1 gates the transition of SCs from a protective, oxidative phosphorylation-dependent state (which we term Protection Associated Schwann Cells or PASCs) to a glycolytic, pro-regenerative repair phenotype after injury. Our findings challenge the prevailing view of Sarm1 as an exclusively axon-autonomous regulator of degeneration and reveal a paradigm shift in understanding the role of Sarm1 in the SC injury response, with broad implications for the treatment of peripheral neuropathies and neurodegenerative diseases.
Beyond the Black Box: Avenues for Transparency in Regulating Radiological AI/ML-enabled SaMD via the FDA 510(k) Pathway
Authors: Youssef Alaa , Fronk David , Grimes John Nicholas , Cheuy Lina , Larson David B.
Published: July 2024

Background

The majority of AI/ML-enabled software as a medical device (SaMD) has been cleared through the FDA 510(k) pathway, but with limited transparency on algorithm development details. Because algorithm quality depends on the quality of the training data and algorithmic input, this study aimed to assess the availability of algorithm development details in the 510(k) summaries of AI/ML-enabled SaMD. Then, clinical and/or technical equivalence between predicate generations was assessed by mapping the predicate lineages of all cleared computer-assisted detection (CAD) devices, to ensure equivalence in diagnostic function.

Methods

The FDA’s public database was searched for CAD devices cleared through the 510(k) pathway. Details on algorithmic input, including annotation instructions and definition of ground truth, were extracted from summary statements, product webpages, and relevant publications. These findings were cross-referenced with the American College of Radiology–Data Science Institute AI Central database. Predicate lineages were also manually mapped through product numbers included within the 510(k) summaries.

Results

In total, 98 CAD devices had been cleared at the time of this study, with the majority being computer-assisted triage (CADt) devices (67/98). Notably, none of the cleared CAD devices provided image annotation instructions in their summaries, and only one provided access to its training data. Similarly, more than half of the devices did not disclose how the ground truth was defined. Only 13 CAD devices were reported in peer-reviewed publications, and only two were evaluated in prospective studies. Significant deviations in clinical function were seen between cleared devices and their claimed predicate.

Conclusion

The lack of imaging annotation instructions and signicant mismatches in clinical function between predicate generations raise concerns about whether substantial equivalence in the 510(k) pathway truly equates to equivalent diagnostic function. Avenues for greater transparency are needed to enable independent evaluations of safety and performance and promote trust in AI/ML-enabled devices.
Multimodal Prehabilitation in People Awaiting Acute Inpatient Cardiac Surgery: Study Protocol for a Pilot Feasibility Trial (PreP-ACe)
Authors: Raut Sarah , Hales Aaron , Twiddy Maureen , Dixon Lili , Ngaage Dumbor , Yates David , Danjoux Gerard , Ingle Lee
Published: July 2024

Background

The concept of “prehabilitation,” or optimising individual physical and mental well-being prior to surgery is well established in cancer and orthopaedic populations. However, amongst the cardiac surgery population, the concept is relatively new. Of the few studies available, all focus on the elective surgical population. This pilot feasibility trial is novel as it will focus on the impact of multimodal prehabilitation on the acute inpatient cardiac surgical population.

Methods

This single centre, prospective, single arm pilot feasibility trial will recruit 20 inpatients awaiting cardiac surgery. Measurements will be collected at the start of the trial (baseline), 7 days after intervention, and 14 days after the intervention or before the day of surgery. The primary outcome measure will be feasibility and practicality of the programme in an acute inpatient population. We will be looking into participant eligibility, acceptability, recruitment rates, completion rates and barriers to implementing a prehabilitation programme. Secondary outcomes include incidence of study-related adverse events, improvement in 6 minutes walk test (6MWT), hand grip strength, quality of life, anxiety scores and spirometry. At the end of the trial, we will be seeking the feedback of the participants on key components of the programme to help us inform future work. We hypothesise that light to moderate structured exercise training is low risk and feasible in patients awaiting inpatient cardiac surgery. The study was approved by Health Research Authority and Heath and Care Research Wales (Yorkshire & the Humber-Bradford Leeds Research Ethics Committee: REC reference 23/YH/0255) on the 8th November 2023.

Discussion

Multimodal prehabilitation could improve individual physical and mental wellbeing whilst awaiting inpatient cardiac surgery. Prehabilitation can provide individuals with a sense of ownership and control over their condition, improve their motivation and independence, and enhance their mental and physical recovery after surgery. Traditionally, patients waiting for cardiac surgery are discouraged from physical activity/ structured exercise training and receive limited information regarding their health. Appropriate physical and psychological support could improve their confidence to mobilise sooner after surgery. This may then facilitate earlier discharge leading to improved hospital bed utilisation and patient flow.