bluuebunny/crossref_metadata_2025 huggingface dataset:
High-Density Lipoprotein Anti-Inflammatory Capacity and Acute Kidney Injury After Cardiac and Vascular Surgery: A Prospective Observational Study
Authors: Perkins Zoe M. , Smith Derek K. , Yancey Patricia G. , Linton MacRae F. , Smith Loren E.
Published: October 2024

OBJECTIVES:

Acute kidney injury (AKI) predicts death after cardiac and vascular surgery. Higher preoperative high-density lipoprotein (HDL) concentrations are associated with less postoperative AKI. In animals, HDL's anti-inflammatory capacity to suppress endothelial cell adhesion molecule expression reduces kidney damage due to ischemia and hemorrhagic shock. The objective of this study is to evaluate the statistical relationship between HDL anti-inflammatory capacity and AKI after major cardiac and vascular surgery.

DESIGN:

Prospective observational study.

SETTING:

Quaternary medical center.

PATIENTS:

One hundred adults with chronic kidney disease on long-term statin therapy undergoing major elective cardiac and vascular surgery.

INTERVENTIONS:

None.

MEASUREMENTS AND MAIN RESULTS:

Apolipoprotein B-depleted serum collected at anesthetic induction was incubated with tumor necrosis factor alpha stimulated human endothelial cells. Reverse transcriptase-polymerase chain reaction was used to measure intercellular adhesion molecule-1 (ICAM-1) messenger RNA. Enzyme-linked immunosorbent assay assays were used to measure apolipoprotein A-I and postoperative soluble ICAM-1 concentrations in patient plasma. HDL concentration did not correlate with HDL ICAM-1 suppression capacity (Spearman R = 0.05; p = 0.64). Twelve patients (12%) were found to have dysfunctional, pro-inflammatory HDL. Patients with pro-inflammatory HDL had a higher rate of postoperative AKI than patients with anti-inflammatory HDL (p = 0.046). After adjustment for AKI risk factors, a higher preoperative HDL capacity to suppress endothelial ICAM-1 was independently associated with lower odds of AKI (odds ratio, 0.88; 95% CI, 0.80–0.98; p = 0.016). The association between HDL anti-inflammatory capacity and postoperative AKI was independent of HDL concentration (p = 0.018). Further, a higher long-term statin dose was associated with higher HDL capacity to suppress endothelial ICAM-1 (p = 0.045).

CONCLUSIONS:

Patients with chronic kidney disease undergoing cardiac and vascular surgery who have dysfunctional, pro-inflammatory HDL have a higher risk of postoperative AKI compared with patients with anti-inflammatory HDL. Conversely, a higher HDL anti-inflammatory capacity is associated with a lower risk of postoperative AKI, independent of HDL concentration. Higher long-term statin dose is associated with higher HDL anti-inflammatory capacity.
Science of Learning Strategy Series: Article 6, Elaboration
Authors: Van Hoof Thomas J. , Sumeracki Megan A. , Madan Christopher R. , Meehan Thomas P.
Published: October 2024

The science of learning (learning science) is an interprofessional field that concerns itself with how the brain learns and remembers important information. Learning science has compiled a set of evidence-based strategies, such as distributed practice, retrieval practice, and interleaving, which are quite relevant to continuing professional development. Spreading out study and practice separated by cognitive breaks (distributed practice), testing oneself to check mastery and memory of previously learned information (retrieval practice), and mixing the learning of separate but associated information (interleaving) represent strategies that are underutilized in continuing professional development. Participants and planners alike can benefit from learning science recommendations to inform their decisions. Elaboration, the subject of this article, is another evidence-based strategy that relates to how one studies or practices priority information.
Investigating the Transfer of Lisinopril Into Human Milk: A Quantitative Analysis
Authors: Chugh Julie , Dai Jean , Datta Palika , Krutsch Kaytlin
Published: October 2024

Lisinopril is commonly prescribed to manage conditions such as hypertension and heart failure. Although concerns about fetal toxicity have traditionally limited the use of lisinopril in women of reproductive age, recent American College of Obstetricians and Gynecologists guidelines promote aggressive treatment of hypertension, which may require the use of pharmacologic agents not previously considered in the postpartum period. We aimed to estimate infant exposure to maternal lisinopril through breast milk and report the tolerance of the breastfed infant. Five volunteers taking lisinopril provided samples of their human milk and their associated health information for research through the InfantRisk Center Human Milk Biorepository. The milk pharmacokinetics of lisinopril were measured using liquid chromatography-mass spectrometry. The mean milk concentration of lisinopril was 0.49 ng/mL per 10 mg daily dose. The relative infant dose was 0.06% for lisinopril, more than 100 times lower than the standard 10% safety threshold. The minimal transfer of lisinopril into human milk in this study suggests the drug is unlikely to pose a clinically significant risk to healthy breastfed infants.
Natural menopause timing and menopause symptoms among lesbian, bisexual, and heterosexual women in the Nurses' Health Study II
Authors: VanKim Nicole A. , Charlton Brittany M. , Berzansky Isa , Fields Nicole D. , Whitcomb Brian W. , Hankinson Susan E. , Sievert Lynnette Leidy , Bertone-Johnson Elizabeth R.
Published: October 2024

Objective

This study aimed to examine sexual orientation differences in natural menopause timing and symptoms between lesbian and bisexual women compared with heterosexual women.

Methods

We used longitudinal questionnaire data (1989-2015) from 92,314 women (858 lesbian, 375 bisexual) in the Nurses' Health Study II cohort. Women were 24-44 yr old at baseline and biennially reported their menopause status, including reasons for cessation of menstrual periods. In 2009 and 2013, women reported on their experience of hot flashes and night sweats. Covariates included age, age at menarche, body mass index, smoking, and parity. Sexual orientation was reported in 1995 and 2009.

Results

Age-adjusted Cox models and logistic regression models suggest that there were no statistically significant differences in menopause timing between lesbian or bisexual women when compared with heterosexual women. When examining menopause symptoms, lesbian women consistently had a statistically significantly (P < 0.05) higher odds of experiencing hot flashes or night sweats (odds ratio range: 1.17 to 1.72) and moderate/severe symptoms (OR range: 1.26 to 1.77) than heterosexual women, even after adjusting for covariates such as smoking and obesity. There were no statistically significant differences in menopause symptoms between bisexual and heterosexual women.

Conclusions

Our findings suggest no meaningful difference in menopause timing between sexual minority and heterosexual women. Additionally, menopause symptoms were more likely among lesbian women and warrants additional study.
Hiding in Plain Sight: Radiologic and Pathologic Findings Can Identify Beckwith-Wiedemann Syndrome in Patients With Wilms Tumor
Authors: Molina Laura M. , Rao Akhila , Meade Julia , Squires Judy H. , Yatsenko Svetlana A. , Salgado Claudia M. , Reyes-Múgica Miguel
Published: October 2024

Most pediatric specialists, including hematologists/oncologists, surgeons, radiologists, and pathologists, are familiar with the diagnosis and management of Wilms tumor (WT). However, it may be challenging to identify the underlying conditions causing cancer predisposition, which can change the management for the patient and potentially their entire family. In this paper, we present 3 cases of clinically suspected WT associated with Beckwith-Wiedemann syndrome (BWS). We review the radiologic and histologic findings to diagnose BWS. We also discuss the implications of a BWS diagnosis on the clinical management of WT and follow-up guidelines for BWS patients.
Management of chronic septic nonunion of the femur associated with an extended drug resistant nosocomial germ: a case report
Authors: Kalantar Seyed Hadi , Bagheri Nima , Khabiri Seyyed Saeed , Oghbaee Behnam , Hoveidaei Amir Human
Published: October 2024

Introduction and Importance:

Septic nonunion of long bones poses significant treatment challenges, often necessitating extended hospital stays and multiple surgical interventions. This report details a chronic septic nonunion of the femur, complicated by drug-resistant pathogens, managed through an integrated surgical approach.

Case Presentation:

A 32-year-old male with a history of infected nonunion and multiple failed surgeries presented with a vast bone defect and osteomyelitis. Treatment involved single-session debridement, bone resection, intramedullary fixation, and trifocal segment bone transport using the Limb Reconstruction System (LRS) and Masquelet technique. After eight months, the patient achieved radiologic and clinical bone healing, regained full ambulation, and resumed work without re-infection over an 18-months follow-up.

Clinical Discussion:

The management of septic nonunion, particularly with drug-resistant infections, requires a multifaceted approach. This case exemplifies the efficacy of combining radical bone resection, targeted debridement, and advanced reconstructive techniques. The use of antibiotic-impregnated cement beads and bone transport procedures played pivotal roles in the successful outcome. While such procedures can be complex and necessitate vigilant monitoring, they are crucial for improving treatment success and functional recovery.

Conclusion:

Septic nonunion caused by extended drug-resistant germs of the lower extremity long bones can be managed in a single-session bone resection and debridement, local antibiotic administration, and later bone defect reconstruction.
Impact of an Educational Deprescribing Intervention on Provider Confidence, Knowledge and Polypharmacy in the Nursing Home Setting
Authors: Murthi Julianne , Langford Molly , Abdallah Lisa
Published: October 2024

Polypharmacy is commonly encountered by providers caring for patients with medically complex and palliative care needs in many settings. The purpose of this quality improvement project was to measure the impact of an evidence-based educational deprescribing intervention on polypharmacy rate and provider confidence and knowledge in the nursing home. We invited providers working in 52 nursing homes to attend a 1-hour-long educational deprescribing session. Twenty-one nurse practitioners and 1 physician assistant across 11 states participated in the intervention. Provider confidence level related to deprescribing improved in all categories, with statistical significance demonstrated with both paired t test and Wilcoxon signed rank test (P < .001). The polypharmacy rate 3 months after the intervention decreased more in centers where a provider had attended the training. Additional open-ended data about experiences with and barriers to deprescribing were collected and analyzed. The findings from this quality improvement project demonstrate that an educational intervention focused on providers practicing in the nursing home setting can improve deprescribing confidence and reduce polypharmacy rates. These findings may be used to implement similar deprescribing education programs for palliative care nurses and providers that prioritize goals of care for patients living with serious illness.
Understanding Behavior Change in Clinical Practice Guideline Implementation: A Qualitative Study
Authors: Tilson Julie K. , Martinez Clarisa , Mickan Sharon , D’Silva Linda J. , Howard Robbin , MacDowell Sara , Roth Heidi R. , Skop Karen M. , Dannenbaum Elizabeth , Farrell Lisa
Published: October 2024

Background and Purpose:

Growing numbers of clinical practice guidelines (CPGs) are available to neurologic physical therapists to guide and inform evidence-based patient care. Adherence to CPG recommendations often necessitates behavior change for therapists and patients. The purpose of this qualitative study was to gain insight into the experiences, perspectives, and drivers of behavioral change for therapists working to improve adherence to a CPG. We also sought to understand the perspectives of patients impacted by this work.

Methods:

Five sites participated in a 6-month implementation study integrating a CPG into local practice using the Knowledge to Action model. At the conclusion of the intervention, therapists and patients were recruited to participate in semi-structured interviews or focus groups. An inductive phenomenological approach was used for data analysis. Two authors coded data to generate primary themes. A secondary analysis used the Capability, Opportunity, Motivation, Behavior (COM-B) model to explain the drivers of behavior change for therapists and patients.

Results:

Perspectives from 16 therapists generated 6 themes around feedback/accountability, teamwork/belonging, complexity/adaptability, leadership/prioritization, engagement/benefit, and motivation/growth. Twelve patients’ perspectives generated 2 themes around communication/personalization and support/recovery. Drivers for behavior change associated with the COM-B model are highlighted.

Discussion and Conclusions:

Therapist adherence to CPG recommendations was supported by inclusive and goal-directed teams, regular quantitative audit and feedback, opportunities for learning, and a sense of accountability to their coworkers, patients, and themselves. Patients’ engagement in rehabilitation was supported by personalized education, objective measures of progress, and a strong therapeutic relationship.

Video Abstract available

for more insights from the authors (see the video, Supplemental Digital Content 1, available at: http://links.lww.com/JNPT/A491).
Whole transcription analysis identified the regulation of hypoxia-inducible factors in monocytes with immune suppression: implications for clinical outcomes
Authors: Shuai Zhao , Hui Li , Wei Luo , Zhaolan Hu , Yulu Wang , Tao Liu , Yanling Zhang , RuPing Dai
Published: October 2024

Aims

Sepsis progression is marked by a complex immune response, where the involvement of hypoxia-inducible factors (HIF) plays an uncertain role. The study aims to elucidate the involvement of HIF-1α in monocyte function during sepsis and its potential as a prognostic indicator.

Methods and Results

Transcriptomic data from healthy individuals and septic patients in datasets GSE54514, GSE167363, and GSE46955 were analyzed. Additionally, human monocytes were employed to elucidate how HIF regulates immune responses in the context of sepsis. Septic non-survivors exhibited sustained upregulation of HIF-1α expression alongside compromised inflammatory response and antigen presentation, with downregulation of NF-κB and HLADRB1 genes associated with poor sepsis prognosis. Conversely, septic survivors displayed an increased proportion of classical monocytes and enhanced inflammation and expression of antigen presentation-related genes. During the recovery phase of sepsis, monocytes continued to demonstrate elevated HIF-1α expression. In cultured THP1 cells and septic CD14+ monocytes, HIF hindered inflammatory responses and antigen presentation, while also suppressing the proportion of classical monocytes after LPS stimulation. Mechanistically, HIF significantly attenuated LPS-induced immune responses in monocytes by inhibiting the phosphorylation of IKK.

Conclusions

HIF in monocytes acts as a suppressor of immune-inflammatory responses and antigen presentation, and may serve as a negative molecular marker for sepsis development.
The Current State of Surgical Ergonomics
Authors: Park Amy J.
Published: October 2024

Objective

Work-related musculoskeletal disorders are widespread among surgeons, causing significant disability and career modification. Gender plays a role, as female surgeons have a two-fold greater risk for discomfort during certain operations due to generally smaller hand sizes and height as compared to male surgeons. There is a lack of appropriate surgical instrumentation geared toward smaller hand sizes. Maintaining a neutral position while operating, frequent changes in position, and increased awareness can contribute to decreasing the detrimental impact on surgeons' bodies from performing operations.
Perspectives on Surgical Leadership: Panel Discussion at the Society for Clinical Vascular Surgery
Authors: Bismuth Jean , Shames Murray , Duncan Audra , Mitchell Erica L , Howard David , Slaw Kenneth M , DeAugustine Perry , Wise Nadia , Teixeira Pedro G R , Lee Jason T
Published: October 2024

Leadership is a skill that all surgeons are confronted with in some capacity. Surprisingly, in the US most training programs do not offer a structured program in leadership and there certainly are no metrics used to assess leadership competency. As a response to this, at the Society for Clinical Vascular Surgery, a panel of leaders in vascular surgery both national and international along with leadership experts discussed some of the salient issues in this space. This article is the result of this discussion and serves as a good framework for understanding needs and current shortcomings of leadership training.
A demographic assessment of the Lansing Effect in duckweed (
Lemna turionifera
Landolt)
Authors: Dutt Priyanka , Laird Robert A.
Published: October 2024

The Lansing Effect is the propensity for offspring of older parents to have shorter lifespans than offspring of younger parents. A recent review identified two demographic patterns that can produce the Lansing Effect: (i) a greater offspring mortality rate at all offspring ages in offspring of older versus younger parents (greater initial mortality parameter); and (ii) an offspring mortality rate that increases more rapidly with offspring age in offspring of older versus younger parents (greater mortality rate parameter). Here, we report on a longitudinal study designed to investigate these patterns, using the duckweed
Lemna turionifera
. We tracked asexually produced offspring that detached from parents that were comparatively young versus old (first versus fifth offspring, respectively). Offspring of older parents lived 15% shorter, on average, than offspring of younger parents, providing evidence of the Lansing Effect. A model-selection approach revealed that the difference between survival curves of first versus fifth offspring was mainly attributable to greater initial mortality in fifth compared to first offspring, though alternative models also received some support. Our study provides a demographic explanation for the Lansing Effect in
L. turionifera
in particular and provides a method for assessing the survival patterns underpinning the Lansing Effect in general.
Neural mechanisms of mutualistic fish cleaning behaviour: a study in the wild
Authors: Romeo Daniele , Ramirez-Calero Sandra , Ravasi Timothy , Rodolfo-Metalpa Riccardo , Schunter Celia
Published: October 2024

One crucial interaction for the health of fish communities in coral reefs is performed by cleaner fish by removing ectoparasites from the body of other fish, so-called clients. Studying the underlying mechanisms of this behaviour is essential to understanding how species react to social stimuli and defining the drivers of mutualistic social behaviour. Here, we pinpoint the neural molecular mechanisms in the cleaning behaviour of
Labroides dimidiatus
in the wild through an
in situ
interaction experiment at a coral reef in New Caledonia. Five cleaners and clients (
Abudefduf saxatilis
) were placed into underwater aquaria to interact, while five were not presented with a client. The brain transcriptomes revealed 233 differentially expressed genes in cleaners that were interacting with a client. Among these genes,
grin2d
,
npy
,
slc6a3
and immediate early genes (IEGs;
fosb
and
fosl1
) were related to learning and memory, glutamate and dopamine pathways, which confirm molecular pathways observed in laboratory studies. However, a new potential mechanism was found with
npy
(neuropeptide Y) as a driver of feeding behaviour. These results show the role of neurotransmitters and IEGs in mutualistic social behaviour, unveiling the mechanism behind the feeding stimulus that leads the cleaner fish to establish mutualistic interactions in coral reefs.
Modelling the mechanisms of antibody mixtures in viral infections: the cases of sequential homologous and heterologous dengue infections
Authors: Dugourd-Camus Charlotte , Ferreira Claudia P. , Adimy Mostafa
Published: October 2024

Antibodies play an essential role in the immune response to viral infections, vaccination or antibody therapy. Nevertheless, they can be either protective or harmful during the immune response. Moreover, competition or cooperation between mixed antibodies can enhance or reduce this protective or harmful effect. Using the laws of chemical reactions, we propose a new approach to modelling the antigen–antibody complex activity. The resulting expression covers not only purely competitive or purely independent binding but also synergistic binding which, depending on the antibodies, can promote either neutralization or enhancement of viral activity. We then integrate this expression of viral activity in a within-host model and investigate the existence of steady-states and their asymptotic stability. We complete our study with numerical simulations to illustrate different scenarios: firstly, where both antibodies are neutralizing and secondly, where one antibody is neutralizing and the other enhancing. The results indicate that efficient viral neutralization is associated with purely independent antibody binding, whereas strong viral activity enhancement is expected in the case of purely competitive antibody binding. Finally, data collected during a secondary dengue infection were used to validate the model. The dataset includes sequential measurements of virus and antibody titres during viremia in patients. Data fitting shows that the two antibodies are in strong competition, as the synergistic binding is low. This contributes to the high levels of virus titres and may explain the antibody-dependent enhancement phenomenon. Besides, the mortality of infected cells is almost twice as high as that of susceptible cells, and the heterogeneity of viral kinetics in patients is associated with variability in antibody responses between individuals. Other applications of the model may be considered, such as the efficacy of vaccines and antibody-based therapies.
External and internal driving forces of community and functional group stability in the forest mammal community
Authors: She Wen , Gu Jiayin , Qi Jinzhe , Liu Shuyan , Yang Xifu , Zhang Zhibin , Jiang Guangshun
Published: October 2024

Conserving mammal community stability in forest ecosystems is an urgent challenge in the face of global biodiversity threats and complex trophic interaction. However, understanding of the stability of mammal communities and targeted functional groups is still limited, hindering adaptive strategy development. We collected 210 120 camera trap photos across 5897 km
2
within the Amur tiger (
Panthera tigris altaica
) range in China’s natural forests. We decomposed stability into compensatory effects, statistical-averaging effects and population stability. Using structural equation modelling, we explored the impact of external factors, species diversity and vertical effects on the stability of the community and of four functional groups. As external factors, primary productivity increased the stability of top predators, while precipitation had inconsistent effects. Horizontal diversity (richness and evenness) positively impacted community stability. However, the vertical structure of the community, including trophic-level richness and functional groups’ compensatory dynamics, were the largest drivers for maintaining community stability. Horizontal diversity within functional groups and the diversity or biomass of their food resources/consumers both influenced the stability of functional groups. Our results recognize the importance of conserving and even re-establishing vertical trophic structures in forest communities. Acknowledging multiple levels of stability, from functional groups to communities, is an indispensable step for effective conservation strategies.
Statistically principled feature selection for single cell transcriptomics
Authors: Dollinger Emmanuel , Silkwood Kai , Atwood Scott , Nie Qing , Lander Arthur D.
Published: October 2024

The high dimensionality of data in single cell transcriptomics (scRNAseq) requires investigators to choose subsets of genes (feature selection) for downstream analysis (e.g., unsupervised cell clustering). The evaluation of different approaches to feature selection is hampered by the fact that, as we show here, the performance of feature selection methods varies greatly with the task being performed. For routine cell type identification, even randomly chosen features can perform well, but for cell type differences that are subtle, both number of features and selection strategy can matter strongly. Here we present a simple feature selection method grounded in an analytical model that, without resorting to arbitrary thresholds or user-defined parameters, allows for interpretable delineation of both how many and which features to choose, facilitating identification of biologically meaningful rare cell types. We compare this method to default methods in scanpy and Seurat, as well as SCTransform, showing how greater accuracy can often be achieved with surprisingly few, well-chosen features.
Integrated Isoform-Resolved Transcriptomic Analysis ofGossypium barbadenseandGossypium hirsutumFibers
Authors: Tang Jiwei , Gu Xinxin , Ma Yue , Meng Qingying , Xie Peihao , Jiang Shihe , Hui Liuyang , Lang Yiyang , Tang Mengqin , Zheng Ying , Yuan Daojun
Published: October 2024

A comprehensive understanding of RNA expression and splicing during cotton fiber development plays a crucial role in explaining the differences in fiber quality between two different cotton species. To date, most cotton transcriptomic studies have utilized short-read sequencing data, which limits the ability to capture full-length mRNAs. In this study, we compiled long-read RNA sequencing data from the PacBio platform, as well as short-read RNA sequencing data from 10 fiber developmental stages, for bothGossypium barbadenseandGossypium hirsutum. We identified 183,767 and 178,994 isoforms inGossypium barbadenseandGossypium hirsutum, respectively, generating the most comprehensive transcriptomic atlas of cotton to date. Alternative splicing events exhibited stage-specific variations during fiber development, and co-expression network analysis, combined with LASSO regression, identified isoforms highly correlated with each developmental stage. These findings reveal previously uncharacterized aspects of isoform regulation during fiber development and provide valuable resources for further research into the molecular mechanisms underlying fiber quality differences between cotton species.
Deep Learning Enhances Precision of Citrullination Identification in Human and Plant Tissue Proteomes
Authors: Gabriel Wassim , Meelker Gonzalez Rebecca , Laposchan Sophia , Riedel Erik , Dündar Gönül , Poppenberger Brigitte , Wilhelm Mathias , Lee Chien-Yun
Published: October 2024

Citrullination is a critical yet understudied post-translational modification (PTM) implicated in various biological processes. Exploring its role in health and disease requires a comprehensive understanding of the prevalence of this PTM at a proteome-wide scale. Although mass spectrometry has enabled the identification of citrullination sites in complex biological samples, it faces significant challenges, including limited enrichment tools and a high rate of false positives due to the identical mass with deamidation (+0.9840 Da) and errors in monoisotopic ion selection. These issues often necessitate manual spectrum inspection, reducing throughput in large-scale studies. In this work, we present a novel data analysis pipeline that incorporates the deep learning model Prosit-Cit into the MS database search workflow to improve both the sensitivity and precision of citrullination site identification. Prosit-Cit, an extension of the existing Prosit model, has been trained on ∼53,000 spectra from ∼2,500 synthetic citrullinated peptides and provides precise predictions for chromatographic retention time and fragment ion intensities of both citrullinated and deamidated peptides. This enhances the accuracy of identification and reduces false positives. Our pipeline demonstrated high precision on the evaluation dataset, recovering the majority of known citrullination sites in human tissue proteomes and improving sensitivity by identifying up to 14 times more citrullinated sites. Sequence motif analysis revealed consistency with previously reported findings, validating the reliability of our approach. Furthermore, extending the pipeline to a tissue proteome dataset of the model plantArabidopsis thalianaenabled the identification of ∼200 citrullination sites across 169 proteins from 30 tissues, representing the first large-scale citrullination mapping in plants. This pipeline can be seamlessly applied to existing proteomics datasets, offering a robust tool for advancing biological discoveries and deepening our understanding of protein citrullination across species.
Regulation of replication timing inSaccharomyces cerevisiae
Authors: Berners-Lee Rosie , Gilmore Eamonn , Berkemeier Francisco , Boemo Michael A.
Published: October 2024

In order to maintain genomic integrity, DNA replication must be highly coordinated. Disruptions in this process can cause replication stress which is aberrant in many pathologies including cancer. Despite this, little is known about the mechanisms governing the temporal regulation of DNA replication initiation, thought to be related to the limited copy number of firing factors. Here, we present a high (1-kilobase) resolution stochastic model ofSaccharomyces cerevisiaewhole-genome replication in which origins compete to associate with limited firing factors. After developing an algorithm to fit this model to replication timing data, we validated the model by reproducing experimental inter-origin distances, origin efficiencies, and replication fork directionality. This suggests the model accurately simulates the aspects of DNA replication most important for determining its dynamics. We also use the model to predict measures of DNA replication dynamics which are yet to be determined experimentally and investigate the potential impacts of variations in firing factor concentrations on DNA replication.
AlphaFold2’s training set powers its predictions of fold-switched conformations
Authors: Schafer Joseph W. , Porter Lauren L.
Published: October 2024

AlphaFold2 (AF2), a deep-learning based model that predicts protein structures from their amino acid sequences, has recently been used to predict multiple protein conformations. In some cases, AF2 has successfully predicted both dominant and alternative conformations of fold-switching proteins, which remodel their secondary and tertiary structures in response to cellular stimuli. Whether AF2 has learned enough protein folding principles to reliably predict alternative conformations outside of its training set is unclear. Here, we address this question by assessing whether CFold–an implementation of the AF2 network trained on a more limited subset of experimentally determined protein structures– predicts alternative conformations of eight fold switchers from six protein families. Previous work suggests that AF2 predicted these alternative conformations by memorizing them during training. Unlike AF2, CFold’s training set contains only one of these alternative conformations. Despite sampling 1300-4400 structures/protein with various sequence sampling techniques, CFold predicted only one alternative structure outside of its training set accurately and with high confidence while also generating experimentally inconsistent structures with higher confidence. Though these results indicate that AF2’s current success in predicting alternative conformations of fold switchers stems largely from its training data, results from a sequence pruning technique suggest developments that could lead to a more reliable generative model in the future.
Anticancer Target Combinations: Network-Informed Signaling-Based Approach to Discovery
Authors: Yavuz Bengi Ruken , Jang Hyunbum , Nussinov Ruth
Published: October 2024

While anticancer drug discovery has seen dramatic innovations and successes, sequential single therapies are time-limited by resistance, and combinatorial strategies have been lagging. The number of possible drug combinations is vast. To select drug combinations the oncologist requires knowledge of the optimal combination of proteins to co-target. Currently, combinations that the oncologist considers are primarily from empirical observations and clinical praxis. Our aim is to develop a signaling-based method to discover optimal proteins for the oncologist to co-target with drug combinations, and test it on available, patient-derived data. To temper the expected resistance to single drug regimen, we offer a concept-based stratified pipeline aimed at selecting co-targets for drug combinations. Our strategy is unique in its co-target selection being based on signaling pathways. This is significant since in cancer, drug resistance commonly bypasses blocked proteins by wielding alternative, or complementary, routes to execute cell proliferation. Our network-informed signaling-based approach harnesses advanced network concepts and metrics, and our compiled, tissue-specific co-existing mutations. Co-existing driver mutations are common in resistance. Thus, to mimic cancer and counter drug resistance scenarios, our pipeline seeks co-targets that when targeted by drug combinations, can shut off cancer’smodus operandi. That is, its parallel or complementary signaling pathways would be blocked. Rotating through combinations could further lessen emerging resistance. We applied it to patient-derived breast and colorectalESR1|PIK3CAandBRAF|PIK3CAsubnetworks. Consistently, in breast cancer, our results suggest co-targeting proteins from theESR1|PIK3CAsubnetwork with an alpelisib-LJM716 combination. In colorectal cancer, they co-targetBRAF|PIK3CAwith alpelisib, cetuximab, and encorafenib combination. Collectively, our pipeline’s results are promising, and validated by patient-based xenografts.

GRAPHICAL ABSTRACT
IVNS1ABP Deficiency Disrupts Actin Filament Organization and Leads to Cellular Senescence in a Newly Identified Progeroid Neuropathy Syndrome
Authors: Yuan Fang , Tan Ye Sing , Wang Haofei , Ali Ain Nur , Yuan Qiang , Chou Shu-Min , Yen Yu-Hsin , Narayanan Gunaseelan , Zhou Lei , Shboul Mohammad , Bonnard Carine , Reversade Bruno , Zhang Su-Chun
Published: October 2024

A homozygous variant inIVNS1ABPwas identified in three siblings, displaying progeroid features with severe neuropathy. By generating isogenic induced pluripotent stem cells (iPSCs) from the patients’ fibroblasts and differentiating the iPSCs into neural progenitor cells (NPCs), we found that mutant IVNS1ABP fibroblasts, iPSCs, and NPCs exhibited disrupted cytokinesis, DNA damage and cellular senescence. Correspondingly, cerebral organoids displayed premature differentiation of NPCs to neurons. Molecular profiling as well as biochemical and cellular analysis revealed altered binding of mutant IVNS1ABP to actin /actin-associated proteins and dysregulated actin dynamics during cytokinesis. Taken together, we propose that mutant IVNS1ABP dysregulates actin polymerization and organization which is at least partly responsible for the cellular senescence phenotypes in this progeroid neuropathy syndrome.
A novel toolbox of GATEWAY-compatible vectors for rapid functional gene analysis in soybean composite plants
Authors: Mejias Joffrey , Margets Alexandra , Bredow Melissa , Foster Jessica , Khwanbua Ekkachai , Goshon Jackson , Maier Thomas R. , Whitham Steven A. , Innes Roger W. , Baum Thomas J.
Published: October 2024

The generation of transgenic plants is essential for plant biology research to investigate plant physiology, pathogen interactions and gene function. However, producing stable transgenic plants for plants such as soybean is a laborious and time-consuming process, which can impede research progress. Composite plants consisting of wild-type shoots and transgenic roots are an alternative method for generating transgenic plant tissues that can facilitate functional analysis of genes-of-interest involved in root development or root-microbe interactions. In this report, we introduce a novel set of GATEWAY-compatible vectors that enable a wide range of molecular biology uses in roots of soybean composite plants. These vectors incorporate in-frame epitope fusions of green fluorescent protein, 3x-HA, or miniTurbo-ID, which can be easily fused to a gene-of-interest using the GATEWAY cloning system. Moreover, these vectors allow for the identification of transgenic roots using either mCherry fluorescence or the RUBY marker. We demonstrate the functionality of these vectors by expressing subcellular markers in soybean, providing evidence of their effectiveness in generating protein fusions in composite soybean plants. Furthermore, we show how these vectors can be used for gene function analysis by expressing the bacterial effector, AvrPphB in composite roots, enabling the identification of soybean targets via immunoprecipitation followed by mass spectrometry. Additionally, we demonstrate the successful expression of stable miniTurbo-ID fusion proteins in composite roots. Overall, this new set of vectors is a powerful tool that can be used to assess subcellular localization and perform gene function analyses in soybean roots without the need to generate stable transgenic plants.

Key Message

We developed a set of GATEWAY vectors to accelerate gene function analysis in soybean composite plants to rapidly screen transgenic roots and investigate subcellular localization, protein-protein interactions, and root-pathogen interactions.
Efficient indexing and querying of annotations in a pangenome graph
Authors: Novak Adam M. , Chung Dickson , Hickey Glenn , Djebali Sarah , Yokoyama Toshiyuki T. , Garrison Erik , Narzisi Giuseppe , Paten Benedict , Monlong Jean
Published: October 2024

The current reference genome is the backbone of diverse and rich annotations. Simple text formats, like VCF or BED, have been widely adopted and helped the critical exchange of genomic information. There is a dire need for tools and formats enabling pangenomic annotation to facilitate such enrichment of pangenomic references. The Graph Alignment Format (GAF) is a text format, tab-delimited like BED/VCF files, which was proposed to represent alignments. GAF could also be used to store paths representing annotations in a pangenome graph, but there are no tools to index and query them efficiently.

Here, we present extensions to vg and HTSlib that provide efficient sorting, indexing, and querying for GAF files. With this approach, annotations overlapping a subgraph can be extracted quickly. Paths are sorted based on the IDs of traversed nodes, compressed with BGZIP, and indexed with HTSlib/tabix via our extensions for the GAF format. Compared to the binary GAM format, GAF files are easier to edit or inspect because they are plain text, and we show that they are twice as fast to sort and half as large on disk. In addition, we updatedvg annotate, which takes BED or GFF3 annotation files relative to linear sequences and projects them into the pangenome. It can now produce GAF files representing these annotations’ paths through the pangenome.

We showcase these new tools on several applications. We projected annotations for all Human Pangenome Reference Consortium Year 1 haplotypes, including genes, segmental duplications, tandem repeats and repeats annotations, into the Minigraph-Cactus pangenome (GRCh38-based v1.1). We also projected known variants from the GWAS Catalog and expression QTLs from the GTEx project into the pangenome. Finally, we reanalyzed ATAC-seq data from ENCODE to demonstrate what a coverage track could look like in a pangenome graph. These rich annotations can be quickly queried with vg and visualized using existing tools like the Sequence Tube Map or Bandage.
mTOR variants activation discovers PI3K-like cryptic pocket, expanding allosteric, mutant-selective inhibitor designs
Authors: Liu Yonglan , Zhang Wengang , Jang Hyunbum , Nussinov Ruth
Published: October 2024

mTOR plays a crucial role in PI3K/AKT/mTOR signaling. We hypothesized that mTOR activation mechanisms driving oncogenesis can advise effective therapeutic designs. To test this, we combined cancer genomic analysis with extensive molecular dynamics simulations of mTOR oncogenic variants. We observed that conformational changes within mTOR kinase domain are associated with multiple mutational activation events. The mutations disturb the α-packing formed by the kαAL, kα3, kα9, kα9b, and kα10 helices in the kinase domain creating cryptic pocket. Its opening correlates with opening of the catalytic cleft, including active site residues realignment, favoring catalysis. The cryptic pocket created by disrupted α-packing coincides with the allosteric pocket in PI3Kα can be harmoniously fitted by the PI3Kα allosteric inhibitor RLY-2608, suggesting that analogous drugs designed based on RLY-2608 can restore the packed α-structure, resulting in mTOR inactive conformation. Our results exemplify that knowledge of detailed kinase activation mechanisms can inform innovative allosteric inhibitor development.
Machine vision based frailty assessment for genetically diverse mice
Authors: Sabnis Gautam S. , Churchill Gary A. , Kumar Vivek
Published: October 2024

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Frailty indexes (FIs) capture health status in humans and model organisms. To accelerate our understanding of biological aging and carry out scalable interventional studies, high-throughput approaches are necessary. We previously introduced a machine vision-based visual frailty index (vFI) that uses mouse behavior in the open field to assess frailty using C57BL/6J (B6J) data. Aging trajectories are highly genetic and are frequently modeled in genetically diverse animals. In order to extend the vFI to genetically diverse mouse populations, we collect frailty and behavior data on a large cohort of aged Diversity Outbred (DO) mice. Combined with previous data, this represents one of the largest video-based aging behavior datasets to date. Using these data, we build accurate predictive models of frailty, chronological age, and even the proportion of life lived. The extension of automated and objective frailty assessment tools to genetically diverse mice will enable better modeling of aging mechanisms and enable high-throughput interventional aging studies.
In vitro models to mimic tumor endothelial cell-mediated immune cell reprogramming in lung adenocarcinoma
Authors: Krejbich Morgane , Fresquet Judith , Navarro Emilie , Forest Virginie , Cotinat Marine , Isen Valentin , Perdrieau Hortense , Chatelain Camille , Dochez Vincent , Roulois David , Boisgerault Nicolas , Redon Richard , Corre Isabelle , Blanquart Christophe , Treps Lucas
Published: October 2024

Tumor endothelial (TECs) cells play a critical role in regulating immune responses within the tumor microenvironment (TME). However, the mechanisms by which TECs modulate immune cell population remain unclear, particularly in non-small cell lung cancer (NSCLC). Here, we investigated how NSCLC cells tweak normal endothelial cells (NECs) into TECs and the subsequent effects on immune regulation. NECs were cocultured with various NSCLC cell lines, using 2D and 3D coculture models to evaluate TEC-mediated effects on immune cells. We show that direct coculture led to significant transcriptomic, proteomic and kinomic alterations in TECs, especially in pro-inflammatory pathways. We identified a downregulation of the co-stimulatory molecule OX40L in TECs compared to NECs, suggesting impaired T-cell proliferation support. While TECs showed a limited effect on CD8+ T-cell activation, TECs supported CD4 T-cells polarization into Treg and Th22 subsets. Moreover, TECs also promoted M2-like macrophages polarization, thereby potentially contributing to the TME immunosuppression. State-of-the-art single-cell RNA sequencing of 3D multicellular tumor spheroids (MCTS) revealed distinct TEC subpopulations, including an inflammatory subset with UPR signature. The latter was absent in 2D-cultured NECs but present in freshly isolated and 2D-cultured TECs from NSCLC patients. Importantly, we also identified within MCTS a perivascular M2-like macrophage subset, predicted to interact with TECs with MIF and Midkine signaling. In conclusion, TECs in NSCLC tumors play a pivotal role in remodeling the TME immune landscape by promoting immune suppression. This study highlights the complex immunoregulatory functions of TECs within our differentin vitromodels that mimic aspects of the TME. Our data may provide new insights into potential therapeutic strategies targeting TECs or regulatory signaling to improve the efficacy of immunotherapy in NSCLC.
Mutant p53 regulates cancer cell invasion in complex three-dimensional environments through mevalonate pathway-dependent Rho/ROCK signaling
Authors: Guzman Asja , Kawase Tatsuya , Devanny Alexander J. , Efe Gizem , Navaridas Raúl , Yu Karen , Regunath Kausik , Mercer Iris G. , Avard Rachel C. , Muniz de Queiroz Rafaela , Rustgi Anil K. , Kaufman Laura J. , Prives Carol
Published: October 2024

Certain mutations can confer neomorphic gain of function (GOF) activities to the p53 protein that affect cancer progression. Yet the concept of mutant p53 GOF has been challenged. Here, using various strategies to alter the status of mutant versions of p53 in different cell lines, we demonstrate that mutant p53 stimulates cancer cell invasion in three-dimensional environments. Mechanistically, mutant p53 enhances RhoA/ROCK-dependent cell contractility and cell-mediated extracellular matrix (ECM) re-organization via increasing mevalonate pathway-dependent RhoA localization to the membrane. In line with this, RhoA-dependent pro-invasive activity is also mediated by IDI-1, a mevalonate pathway product. Further, the invasion-enhancing effect of mutant p53 is dictated by the biomechanical properties of the surrounding ECM, thereby adding a cell-independent layer of regulation to mutant p53 GOF activity that is mediated by dynamic reciprocal cell-ECM interactions. Together our findings link mutant p53 metabolic GOF activity with an invasive cellular phenotype in physiologically relevant and context-dependent settings.

Significance

This study addresses the contribution of mutant p53 to the process of cancer cell dissemination in physiologically relevant three-dimensional environments – a key characteristic of metastatic disease. Several mutant p53 proteins display pro-oncogenic activity with respect to cancer cell invasion in 3D environments via mevalonate pathway-dependent Rho/ROCK signaling axis.