bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluating the utility of REVEL and CADD for interpreting variants in amyotrophic lateral sclerosis genes
Authors: Fiorini Michael R. , Dilliott Allison A. , Farhan Sali M.K.
Published: August 2023

Amyotrophic lateral sclerosis (ALS) is a debilitating neurodegenerative disease affecting approximately two per 100,000 individuals globally. While there are many benefits to offering early genetic testing to people with ALS, this has also led to an increase in the yield of novel variants of uncertain significance in ALS-associated genes. Computational (in silico) predictors, including REVEL and CADD, are widely employed to provide supporting evidence of pathogenicity for variants in conjunction with clinical, molecular, and other genetic evidence. However, in silico predictors are developed to be broadly applied across the human genome; thus, their ability to evaluate the consequences of variation in ALS-associated genes remains unclear. To resolve this ambiguity, we surveyed 20 definitive and moderate ClinGen defined ALS-associated genes from two large, open access ALS sequencing datasets (total people with ALS = 8,230; controls = 9,671) to investigate REVEL and CADD’s ability to predict which variants are most likely to be disease-causing in ALS. While our results indicate a predetermined pathogenicity threshold for REVEL that could be of clinical value for classifying variants in ALS-associated genes, an accurate threshold was not evident for CADD, and both in silico predictors were of limited value for resolving which variants of uncertain significance (VUS) may be likely pathogenic in ALS. Our findings allow us to provide important recommendations of the use of REVEL and CADD scores for variants, and indicate that both tools should be used with caution when attempting to evaluate the pathogenicity of VUSs in ALS genetic testing.
WITHDRAWN: What Risk Factors to Cause Long COVID and Its Impact on Patient Survival Outcomes when Combined with the Effect from Organ Transplantation in the Acute COVID
Authors: Nyandemoh Ahmed N , Anzalone Jerrod , Dai Hongyin , Mannon Roslyn , Dong Jianghu (James)
Published: August 2023

Withdrawal statement

medRxiv has withdrawn this preprint owing to lack of consent from all listed authors. Therefore, this work should not be cited as a reference for this project.
Effectiveness of a mobile health app on initiated antihypertensive medications in patients with untreated hypertension
Authors: Matsumura Koichiro , Nakagomi Atsushi , Yagi Eijiro , Yamada Nobuhiro , Funauchi Yohei , Kakehi Kazuyoshi , Yoshida Ayano , Kawamura Takayuki , Ueno Masafumi , Nakazawa Gaku , Tabuchi Takahiro
Published: August 2023

Background

Few effective tools have been identified that facilitate the initiation of antihypertensive medications in patients with untreated hypertension. To determine whether a mobile health (mHealth) application facilitates the initiation of antihypertensive medications in patients with untreated hypertension.

Methods

We analyzed a large, longitudinal, integrated database mainly comprising middle-aged, working people and their families. The database contained health checkup data, health insurance claims data, and mHealth app data. The mHealth app, kencom, is used to manage daily life logs (i.e. weight, number of steps) and to provide health information tailored to customers. Patients with untreated hypertension were defined using the baseline health checkup data. A multivariable logistic regression analysis was performed to examine the association between use of the mHealth app and the initiation of antihypertensive medications.

Results

Among 50 803 eligible patients (mean age, 49 years; 78 % male) with a median follow-up period of 3.0 years. The rate of initiation of antihypertensive medication was 23.4 % vs. 18.5 % (p < 0.0001), which was significantly higher in the mHealth application group (n = 14 879) than in the non-user group (n = 35 924). Multivariable analysis revealed that usage of the mHealth app was associated with initiated antihypertensive medications (odds ratio 1.43, 95 % confidence interval 1.36–1.50).

Conclusion

In patients with untreated hypertension, the use of the mHealth app, which was not dedicated to hypertension treatment, was associated with the initiation of antihypertensive medications.
Prediction of Daily New COVID-19 Cases - Difficulties and Possible Solutions
Authors: Liu Xiaoping
Published: August 2023

Epidemiological compartmental models, such asSEIR(Susceptible, Exposed, Infectious, and Recovered) models, have been generally used in analyzing epidemiological data and forecasting the trajectory of transmission of infectious diseases such as COVID-19. Experience shows that accurately forecasting the trajectory of COVID-19 transmission curve is a big challenge to researchers in the field of epidemiological modeling. Multiple factors (such as social distancing, vaccinations, public health interventions, and new COVID-19 variants) can affect the trajectory of COVID-19 transmission. In the past years, we used a new compartmental model,l-i SEIRmodel, to analyze the COVID-19 transmission trend in the United States. The letterslandiare two parameters in the model representing the average time length of the latent period and the average time length of infectious period. Thel-i SEIRmodel takes into account of the temporal heterogeneity of infected individuals and thus improves the accuracy in forecasting the trajectory of transmission of infectious diseases. This paper describes how these multiple factors mentioned above could significantly change COVID-19 transmission trends, why accurately forecasting COVID-19 transmission trend is difficult, what the strategies we have used to improve the forecast outcome, and some of successful examples that we have obtained.
A novel Bayesian fine-mapping model using a continuous global-local shrinkage prior with applications in prostate cancer analysis
Authors: Li Xiang , Sham Pak Chung , Zhang Yan Dora
Published: August 2023

The aim of fine-mapping is to identify genetic variants causally contributing to complex traits or diseases. Existing fine-mapping methods employ discrete Bayesian mixture priors and depend on a pre-specified maximum number of causal variants which may lead to sub-optimal solutions. In this work, we propose a novel fine-mapping method called h2-D2, utilizing a continuous global-local shrinkage prior. We also present an approach to define credible sets of causal variants in continuous prior settings. Simulation studies demonstrate that h2-D2 outperforms the state-of-art fine-mapping methods such as SuSiE and FINEMAP in accurately identifying causal variants and estimating their effect sizes. We further applied h2-D2 to prostate cancer analysis and discovered some previously unknown causal variants. In addition, we inferred 385 target genes associated with the detected causal variants and several pathways that were significantly over-represented by these genes, shedding light on their potential roles in prostate cancer development and progression.
Spatial Transcriptomic Analysis of Pituitary Corticotroph Tumors Unveils Intratumor Heterogeneity
Authors: Piña Jeremie Oliver , Faucz Fabio R. , Padilla Cameron , Floudas Charalampos S. , Chittiboina Prashant , Quezado Martha , Tatsi Christina
Published: August 2023

Spatial transcriptomic (ST) analysis of tumors provides a novel approach on studying gene expression along with the localization of tumor cells in their environment to uncover spatial interactions. Herein, we present ST analysis of corticotroph pituitary neuroendocrine tumors (PitNETs) from formalin-fixed, paraffin-embedded (FFPE) tissues. We report that the in situ annotation of tumor tissue can be inferred from the gene expression profiles and is in concordance with the annotation made by a pathologist. Furthermore, relative gene expression in the tumor corresponds to common protein staining used in the evaluation of PitNETs, such as reticulin and Ki-67 index. Finally, we identify intratumor heterogeneity; clusters within the same tumor may present with different secretory capacity and transcriptomic profiles, unveiling potential intratumor cell variability with possible therapeutic interest. Together, our results provide the first attempt to clarify the spatial cell profile in PitNETs.
Recombinant SARS-CoV-2 lacking initiating and internal methionine codons within ORF10 is attenuatedin vivo
Authors: Gu Shichun , Bentley Eleanor G , Milligan Rachel I , Almuqrin Abdulaziz M. , Sharma Parul , Kirby Adam , Mega Daniele F , Kipar Anja , Erdmann Max , Bazire James , Heesom Kate J. , Lewis Philip A , Donovan-Banfield I’ah , Reston Charlotte , Webb Isobel , Neck Simon De , Dong Xaiofeng , Hiscox Julian A , Davidson Andrew D , Stewart James P , Matthews David A.
Published: August 2023

SARS-CoV-2 has been proposed to encode ORF10 as the 3’ terminal gene in the viral genome. However, the potential role and even existence of a functional ORF10 product has been the subject of debate. There are significant structural features in the viral genomic RNA that could, by themselves, explain the retention of the ORF10 nucleotide sequences without the need for a functional protein product. To explore this question further we made two recombinant viruses, firstly a control virus (WT) based on the genome sequence of the original Wuhan isolate and with the inclusion of the early D614G mutation in the Spike protein. We also made a second virus, identical to WT except for two additional changes that replaced the initiating ORF10 start codon and an internal methionine codon for stop codons (ORF10KO). Here we show that the two viruses have apparently identical growth kinetics in a VeroE6 cell line that over expresses TMPRSS2 (VTN cells). However, in A549 cells over expressing ACE2 and TMPRSS2 (A549-AT cells) the ORF10KO virus appears to have a small growth rate advantage. Growth competition experiments were used whereby the two viruses were mixed, passaged in either VTN or A549-AT cells and the resulting output virus was sequenced. We found that in VTN cells the WT virus quickly dominated whereas in the A549-AT cells the ORF10KO virus dominated. We then used a hamster model of SARS-CoV-2 infection and determined that the ORF10KO virus has attenuated pathogenicity (as measured by weight loss). We found an almost 10-fold reduction in viral titre in the lower respiratory tract for ORF10KO vs WT. In contrast, the WT and ORF10KO viruses had similar titres in the upper respiratory tract. Sequencing of viral RNA in the lungs of hamsters infected with ORF10KO virus revealed that this virus frequently reverts to WT. Our data suggests that the retention of a functional ORF10 sequence is highly desirable for SARS-CoV-2 infection of hamsters and affects the virus’s ability to propagate in the lower respiratory tract.
DESIGN, DEVELOPMENT, AND VALIDATION OF A NANOBIOSENSOR TO DETECT CIRCULATING MICROBIOME FOR CARDIOVASCULAR DISEASE RISK ASSESSMENT
Authors: Nazeer Nazim , Dewangan Rakhi , Zaidi Kaniz , Gurjar Vikas , Tiwari Rajnarayan , Mishra Pradyumna Kumar
Published: August 2023

Cardiovascular disease (CVD) is a serious worldwide health concern that necessitates the development of novel diagnostic techniques for early identification and personalized healthcare management. Even before the insights provided by gut microbiota, current research has demonstrated the importance of circulating microbiome (CMB) in the evolution of cardiometabolic illness risk and progression. We developed a nanobiosensor that uses specific labeled capture probes with perovskite quantum dots (PQDs) to detect the targeted 16S rRNA sequences in the peripheral milieu. With ideal applicability, specificity, and sensitivity, this sensor delivers unique insights into the presence and characterization of circulating microbiota signatures. Developing a nanophotonic microbiome detection method in body fluids may pave the way for creating a distinctive tool for CVD risk prediction for population-based screening programs in low and middle-income countries.
The intersection between migration, HIV, and family planning in Uganda: a cross-sectional population-based study
Authors: Namusisi Prossy , Yeh Ping Teresa , Ssekubugu Robert , Chang Larry William , Lutalo Tom , Zimmerman Linnea A. , Grabowski Mary Kathryn
Published: August 2023

Background

Low use of modern methods of contraception has been linked to HIV seropositivity and to migration, but few studies have evaluated the intersection of both risk factors with contraceptive use.

Methods

We analyzed cross-sectional data from sexually active female participants aged 15 to 49 years in the Rakai Community Cohort Study (RCCS) between 2011 and 2013. The RCCS is an open population-based census and individual survey in south-central Uganda. Recent in-migrants (arrival within approximately 1.5 years) into RCCS communities were identified at time of household census. The primary outcome was unsatisfied demand for a modern contraceptive method (injectable, oral pill, implant, or condom), which was defined as non-use of a modern contraceptive method among female participants who did not want to become pregnant in the next 12 months. Poisson regression models with robust variance estimators were used to identify associations and interactions between recent migration and HIV serostatus on unsatisfied contraceptive demand.

Results

There were 3,417 sexually active participants with no intention of becoming pregnant in the next year. The mean age was 30 (±8) years, and 17.3% (n=591) were living with HIV. Overall, 43.9% (n=1,500) were not using any modern contraceptive method. Recent in-migrants were somewhat more likely to have unsatisfied contraceptive demand as compared to long-term residents (adjusted prevalence risk ratio [adjPRR]=1.14; 95% confidence interval [95%CI]: 1.02–1.27), whereas participants living with HIV were less likely to have unsatisfied contraceptive demand relative to HIV-seronegative participants (adjPRR=0.80; 95%CI=0.70-0.90). When stratifying on migration and HIV serostatus, we observed the highest levels of unsatisfied contraceptive demand among in-migrants living with HIV (48.7%); however, in regression analyses, interaction terms between migration and HIV serostatus were not statistically significant.

Conclusions

Unsatisfied contraceptive demand was high in this rural Ugandan setting. Being an in-migrant, particularly among those living with HIV, was associated with higher unsatisfied contraceptive demand.
Spatial and temporal control of 3D hydrogel viscoelasticity through phototuning
Authors: Crandell Philip , Stowers Ryan
Published: August 2023

The mechanical properties of the extracellular environment can regulate a variety of cellular functions, such as spreading, migration, proliferation, and even differentiation and phenotypic determination. Much effort has been directed at understanding the effects of the extracellular matrix (ECM) elastic modulus and more recently, stress relaxation, on cellular processes. In physiological contexts like development, wound healing, and fibrotic disease progression, ECM mechanical properties change substantially over time or space. Dynamically tunable hydrogel platforms have been developed to spatiotemporally modulate a gel’s elastic modulus. However, dynamically altering the stress relaxation rate of a hydrogel remains a challenge. Here, we present a strategy to tune hydrogel stress relaxation rates in time or space using a light-triggered tethering of poly(ethylene glycol) (PEG) to alginate. We show that stress relaxation rate can be tuned without altering the elastic modulus of the hydrogel. We found that cells are capable of sensing and responding to dynamic stress relaxation rate changes, both morphologically and through differences in proliferation rates. We also exploited the light-based technique to generate spatial patterns of stress relaxation rates in 3D hydrogels. We anticipate that user-directed control of 3D hydrogel stress relaxation rate will be a powerful tool that enables studies that mimic dynamic ECM contexts, or as a means to guide cell fate in space and time for tissue engineering applications.
A model to analyze age-related differences in human-generated head-tail sequences
Authors: Baena-Mirabete Sergio , García-Viedma Rosario , Fernández-Guinea Sara , Puig Pere
Published: August 2023

The generation of random-like sequences is a common task for assessing high-level cognitive abilities, such as inhibition, sustained attention and working memory. In general, many studies have shown a detrimental effect of aging on pseudo-random productions. The performance of participants in random generation tasks has typically been assessed by measures of randomness such as, among others, entropy and algorithmic complexity that are calculated from the series of responses produced by the subject. We focus on analyzing the mental model of randomness that people implicitly use when producing random series. We propose a novel latent class model based on Markov chains that aims to classify individuals into homogeneous classes according to the way they generate head-tail series. Our results reveal that there are significant age-related differences in the way individuals produce random-like sequences. Specifically, the group of healthy adults implicitly uses a simpler mental mechanism, in terms of memory requirements, compared to the group of younger participants.

Author summary

It is well known that, in general, people deviate from randomness as they attempt to mentally generate head-tail sequences as randomly as possible. The extensive literature on this topic has shown that human-generated head-tail series tend to have more alternations than would be expected by chance. However, it seems unrealistic to suppose that all individuals generate sequences based on the same random mental model. We conducted an experiment in which 331 individuals were asked to mentally simulate a fair coin: 69 healthy older adults with an age ≥ 60 and 262 Biology students with an age between 18 and 20. We found that the way in which random sequences are generated varies between subjects. A similar approach could be used to analyze differences in random generation tasks between subjects with different disorders and healthy subjects.
EagleC Explorer: A desktop application for interactively detecting and visualizing SVs and enhancer hijacking on Hi-C contact maps
Authors: Fu Yihao , Wang Xiaotao , Yue Feng
Published: August 2023

It has been shown that Hi-C can be used as a powerful tool to detect structural variations (SVs) and enhancer hijacking events. However, there has been no existing programs that can directly visualize and detect such events on a personal computer, which hinders the broad adaption of the technology for intuitive discovery in cancer studies. Here, we introduce the EagleC Explorer, a desktop software that is specifically designed for exploring Hi-C and other chromatin contact data in cancer genomes. EagleC Explorer has a set of unique features, including 1) conveniently visualizing global and local Hi-C data; 2) interactively detecting SVs on a Hi-C map for any user-selected region on screen within seconds, using a deep-learning model; 3) reconstructing local Hi-C map surrounding user-provided SVs and generating publication-quality figures; 4) detecting enhancer hijacking events for any user-suggested regions on screen. In addition, EagleC Explorer can also incorporate other genomic tracks such as RNA-Seq or ChIP-Seq to facilitate scientists for integrative data analysis and making novel discoveries.
FRET-guided modeling of nucleic acids
Authors: Steffen Fabio D. , Cunha Richard A. , Sigel Roland K.O. , Börner Richard
Published: August 2023

The functional diversity of RNA is encoded in their innate conformational heterogeneity. The combination of single-molecule spectroscopy and computational modeling offers new, attractive opportunities to map structural transitions within nucleic acid ensembles. Here, we describe a framework to harmonize single-molecule FRET measurements with molecular dynamics simulations andde novostructure prediction. Using either all-atom or implicit fluorophore modeling we recreate FRET experimentsin silico, visualize the underlying structural dynamics and quantify the simulated reaction coordinates. Using multiple accessible-contact volumes (multi-ACV) as apost-hocscoring method for fragment-assembly in Rosetta, we demonstrate that FRET effectively refinesde novoRNA structure prediction without the need of explicit dye labeling. We benchmark our FRET-assisted modeling approach on double-labeled DNA strands and validate it against an intrinsically dynamic manganese(II)-binding riboswitch. We show that a FRET coordinate describing the assembly of a four-way junction allows our pipeline to recapitulate the global fold of the riboswitch with sub-helical accuracy to the crystal structure. We conclude that computational fluorescence spectroscopy facilitates the interpretability of dynamic structural ensembles and improves the mechanistic understanding of nucleic acid interactions.

Graphical abstract

Schematic workflow of integrative FRET modeling using all-atom fluorophores or an accessible-contact volume dye model. All-atom molecular dynamics track the dye coordinate explicitly as part of the simulation while multi-ACV infer mean dye positionspost hoc.
Development of neonatal connectome dynamics and its prediction for cognitive and language outcomes at age 2
Authors: Xu Yuehua , Liao Xuhong , Lei Tianyuan , Cao Miao , Zhao Jianlong , Zhang Jiaying , Zhao Tengda , Li Qiongling , Jeon Tina , Ouyang Minhui , Chalak Lina , Rollins Nancy , Huang Hao , He Yong
Published: August 2023

The functional connectome of the human brain comprises time-varying network structure that facilitates efficient inter-module communication and support flexible cognitive functions. However, little is known about how the connectome dynamics of the brain emerges and develops at very early stages of human life and whether this dynamics is predictive of neurocognitive outcomes later in life. Here, we employed resting-state functional MRI data from 39 infants (31 to 42 postmenstrual weeks) and a multilayer network model to characterize the development of connectome dynamics during the third trimester and its critical role in predicting future neurocognitive outcomes at 2 years of age. We observed that the modular architecture of baby functional connectomes spontaneously reconfigures over time, with lower network module switching across time primarily in the primary regions and higher module switching mainly in the association areas. With development, the dynamic switching between the brain modules was significantly decreased, primarily located in the lateral precentral gyrus, medial temporal lobe, and subcortical areas. The clustering analysis further revealed that the primary areas displayed a higher developmental rate than the higher-order systems. Using the support vector regression approach, we found that brain connectome dynamics at birth significantly predicted cognitive and language performance at 2 years of age. Our findings highlight the emergence and spatially inhomogeneous maturation of the neonate connectome dynamics, laying a critical neural foundation for the development of cognitive and language skills later in life.
Nuclear Factor Kappa B Over-Activation in the Intervertebral Disc Leads to Macrophage Recruitment and Severe Disc Degeneration
Authors: Burt Kevin G. , Kim Min Kyu M. , Viola Dan C. , Abraham Adam C. , Chahine Nadeen O.
Published: August 2023

Objective

Low back pain (LBP) is the leading cause of global disability and is thought to be driven primarily by intervertebral disc (IVD) degeneration (DD). Persistent upregulation of catabolic enzymes and inflammatory mediators have been associated with severe cases of DD. Nuclear factor kappa B (NF-κB) is a master transcription regulator of immune responses and is over expressed during inflammatory-driven musculoskeletal diseases, including DD. However, its role in triggering DD is unknown. Therefore, this study investigated the effect of NF-κB pathway over-activation on IVD integrity and DD pathology.

Methods

Using skeletally mature mouse model, we genetically targeted IVD cells for canonical NF-κB pathway activation via expression of a constitutively active form of inhibitor of κB kinase B (IKKβ), and assessed changes in IVD cellularity, structural integrity including histology, disc height, and extracellular matrix (ECM) biochemistry, biomechanics, expression of inflammatory, catabolic, and neurotropic mediators, and changes in macrophage subsets, longitudinally up to 6-months post activation.

Results

Prolonged NF-κB activation led to severe structural degeneration, with a loss of glycosaminoglycan (GAG) content and complete loss of nucleus pulposus (NP) cellularity. Structural and compositional changes decreased IVD height and compressive mechanical properties with prolonged NF-κB activation. These alterations were accompanied by increases in gene expression of inflammatory molecules (Il1b, Il6, Nos2), chemokines (Mcp1,Mif), catabolic enzymes (Mmp3, Mmp9, Adamts4), and neurotrophic factors (Bdnf,Ngf) within IVD tissue. Increased recruitment of activatedF4/80+macrophages exhibited a greater abundance of pro-inflammatory (CD38+) over inflammatory-resolving (CD206+) macrophage subsets in the IVD, with temporal changes in the relative abundance of macrophage subsets over time, providing evidence for temporal regulation of macrophage polarization in DDin vivo,where macrophages participate in resolving the inflammatory cascade but promote fibrotic transformation of the IVD matrix. We further show that NF-κB driven secretory factors from IVD cells increase macrophage migration and inflammatory activation, and that the secretome of inflammatory-resolving macrophages mitigates effects of NF-κB overactivation.

Conclusion

Overall the observed results suggest prolonged NF-κB activation can induce severe DD, acting through increases in inflammatory cytokines, chemotactic proteins, catabolic enzymes, and the recruitment and inflammatory activation of a macrophage cell populations, that can be mitigated with inflammatory-resolving macrophage secretome.
Single-cell somatic copy number variants in brain using different amplification methods and reference genomes
Authors: Kalef-Ezra Ester , Turan Zeliha Gozde , Perez-Rodriguez Diego , Bomann Ida , Behera Sairam , Morley Caoimhe , Scholz Sonja W. , Jaunmuktane Zane , Demeulemeester Jonas , Sedlazeck Fritz J , Proukakis Christos
Published: August 2023

The presence of somatic mutations, including copy number variants (CNVs), in the brain is well recognized. Comprehensive study requires single-cell whole genome amplification, with several methods available, prior to sequencing. We compared PicoPLEX with two recent adaptations of multiple displacement amplification (MDA): primary template-directed amplification (PTA) and droplet MDA, across 93 human brain cortical nuclei. We demonstrated different properties for each, with PTA providing the broadest amplification, PicoPLEX the most even, and distinct chimeric profiles. Furthermore, we performed CNV calling on two brains with multiple system atrophy and one control brain using different reference genomes. We found that 38% of brain cells have at least one Mb-scale CNV, with some supported by bulk sequencing or single-cells from other brain regions. Our study highlights the importance of selecting whole genome amplification method and reference genome for CNV calling, while supporting the existence of somatic CNVs in healthy and diseased human brain.
Predictive Modeling of Evoked Intracranial EEG Response to Medial Temporal Lobe Stimulation in Patients with Epilepsy
Authors: Acharya Gagan , Davis Kathryn A. , Nozari Erfan
Published: August 2023

Despite promising advancements, closed-loop neurostimulation for drug-resistant epilepsy (DRE) still relies on manual tuning and produces variable outcomes, while automated predictable algorithms remain an aspiration. As a fundamental step towards addressing this gap, here we study predictive dynamical models of human intracranial EEG (iEEG) response under parametrically rich neurostimulation. Using data from n = 13 DRE patients, we find that stimulation-triggered switched-linear models with ∼300ms of causal historical dependence best explain evoked iEEG dynamics. These models are highly consistent across different stimulation amplitudes and frequencies, allowing for learning a generalizable model from abundant STIM OFF and limited STIM ON data. Further, evoked iEEG in nearly all subjects exhibited a distance-dependent pattern, whereby stimulationdirectlyimpacts the actuation site and nearby regions (≲ 20mm), affects medium-distance regions (20 ∼ 100mm) through network interactions, and hardly reaches more distal areas (≳ 100mm). Peak network interaction occurs at 60∼80mm from the stimulation site. Due to their predictive accuracy and mechanistic interpretability, these models hold significant potential for model-based seizure forecasting and closed-loop neurostimulation design.
Deciphering the orthorhombic crystal structure of a novel NEIL1 nanobody with pseudo-merohedral twinning
Authors: Thompson Marlo K. , Sharma Nidhi , Prakash Aishwarya
Published: August 2023

Nanobodies or VHHs (Variable Heavy domains of Heavy chain) are single domain antibodies that comprise three antigenic complementary determining regions (CDR). Nanobodies are used in numerous scientific applications including, bio-imaging, diagnosis, therapeutics, and macromolecular crystallography. We obtained crystals of a ∼14 kDa nanobody specific for the NEIL1 DNA glycosylase (hereafter called A5) in 0.5 M ammonium sulfate, 0.1 M sodium citrate tribasic dihydrate pH 5.6, and 1.0 M lithium sulfate monohydrate from the Crystal HT Hampton Research screen that were further optimized. Here, we describe the structure determination and refinement of the A5 crystals to a resolution of 2.1 Å. The data collected were complicated by the presence of anisotropy and twinning, and while initial space group determination pointed to a higher apparent tetragonal crystal system, the data statistics suggested twinning, placing the crystal in an orthorhombic system. Twinning was confirmed by the Padilla and Yeates test, H-test, and Britton test based on local intensity differences with a twin fraction of 0.4. Molecular replacement produced the best solution in the orthorhombic space group P21212 with four molecules in the asymmetric unit and we were able to model over 96% of the residues in the electron density with a final Rworkand Rfreeof 0.1988 and 0.2289 upon refinement.

Synopsis

The crystal structure of a specific nanobody against NEIL1 was determined to 2.1 Å. The structure was ultimately solved in an orthorhombic space group after diffraction data analysis revealed mild anisotropy as well as pseudo-merohedral twinning
AAV-HBV mouse model replicates immune exhaustion patterns of chronic HBV patients at single-cell level
Authors: Conceição-Neto Nádia , Han Qinglin , Yao Zhiyuan , Pierson Wim , Wu Qun , Dockx Koen , Aerts Liese , De Maeyer Dries , Van den Berge Koen , Li Chris , Kukolj George , Zhu Ren , Podlaha Ondřej , Nájera Isabel , Van Gulck Ellen
Published: August 2023

Background and Aims

Unresolved hepatitis B virus (HBV) infection leads to a progressive state of immune exhaustion that impairs resolution of infection, leading to chronic infection (CHB). The immune-competent AAV-HBV mouse is a common HBV preclinical immune competent model, though a comprehensive characterization of the liver immune microenvironment and its translatability to human infection is still lacking. We investigated the intrahepatic immune profile of the AAV-HBV mouse model at a single-cell level and compared with data from CHB patients in immune tolerant (IT) and immune active (IA) clinical stages.

Methods

Immune exhaustion was profiled through an iterative subclustering approach for cell-typing analyses of single-cell RNA-sequencing data in CHB donors and compared to the AAV-HBV mouse model 24-weeks post-transduction to assess its translatability. This was validated using an exhaustion flow cytometry panel at 40 weeks post-transduction.

Results

Using single-cell RNA-sequencing, CD8 pre-exhausted T-cells with self-renewing capacity (TCF7+), and terminally exhausted CD8 T-cells (TCF7-) were detected in the AAV-HBV model. These terminally exhausted CD8 T-cells (expressingPdcd1,Tox,Lag3,Tigit) were significantly enriched versus control mice and independently identified through flow cytometry. Importantly, comparison to CHB human data showed a similar exhausted CD8 T-cell population in IT and IA donors, but not in healthy individuals.

Conclusions

Long term high titer AAV-HBV mouse liver transduction led to T-cell exhaustion, as evidenced by expression of classical immune checkpoint markers at mRNA and protein levels. In both IT and IA donors, a similar CD8 exhausted T-cell population was identified, with increased frequency observed in IA donors. These data support the use of the AAV-HBV mouse model to study T-cell exhaustion in HBV infection and the effect of immune-based therapeutic interventions.

Lay Summary

The AAV-HBV mouse model is used as a research tool to study hepatitis B infection. In this study we evaluated the translation value from mouse to human with regards to T-cell exhaustion.

Highlights

AAV-HBV mice transduced with a high titer vector showed presence of CD8 exhausted T-cells after 24 weeks.

High titer transduced mice, but not lower titer show increased expression of LAG-3, TOX, TIM-3 and TIGIT in CD8 T-cells. PD-1 was increased in CD8 T-cells, independent of HBV transduction titer.

A similar exhausted CD8 T-cell population could be found in chronic HBV donors, but not in healthy individuals.

Graphical abstract
Discovery of a Well-Folded Protein Interaction Hub Within the Human Long Non-Coding RNANORAD
Authors: Kumar Ananthanarayanan , Wan Han , Perry Zion , Patel Shivali , Tavares Rafael , Pyle Anna Marie
Published: August 2023

The long non-coding RNANORADfunctions in maintaining genomic stability in humans via sequestering Pumilio proteins from the cytoplasm, and thereby modulating the gene expression of mRNA targets of Pumilio proteins. Despite its role in fundamental cellular pathways including chromosome segregation and DNA damage response, there have been limited structural and biophysical descriptions ofNORAD. Here, using an integrative approach combining chemical probing coupled to high throughput sequencing, and RNA-pull downs coupled with mass spectrometry, we discovered a well-folded and structured protein interaction hub within the functional core ofNORAD. Ourin vitrobiochemical reconstitutions using purified recombinant proteins and aNORADrepeat unit region within this hub reveal the assembly of a higher-order multimeric RNA-protein complex.
Unravelling the complex biogeographic and anthropogenic history of Alaska’s mountain goats
Authors: Young Kiana B. , White Kevin S. , Shafer Aaron B.A.
Published: August 2023

Aim

We used genetic tools to examine the population structure of mountain goats in Alaska, USA and assessed the demographic history of this species in relation to the natural and anthropogenic forces.

Location

Alaska, USA

Taxon

North American mountain goat (Oreamnos americanus)

Methods

Samples were collected between 2006 - 2020 from harvested animals and live captures. We genotyped 816 mountain goats at 18 microsatellite loci and identified the number of genetically distinct subpopulations across the state and assessed their genetic diversity. We used Bayesian computation software to investigate the demographic history relative to the known biogeographic history of the state. We also simulated island translocation events and compared simulations to empirical data to address the hypothesis that Baranof Island was a cryptic refugia.

Results

We showed that Alaska has four genetically distinct subpopulations of mountain goats with some additional genetic structure within those subpopulations. The main split of mountain goats between Southcentral and Southeast Alaska occurred ∼14,000 years ago. Simulations of translocation events largely aligned with the current populations observed today except for Baranof Island which showed greater diversity than the translocation simulation.

Main Conclusions

The distribution and genetic structure of mountain goats in Alaska reflects a combination of natural and anthropogenic forces. A rapid northerly expansion through an ice-free corridor in combination with the isolated nature of the landscape led to low diversity and isolation 14,000 years ago in Southcentral Alaska and higher diversity in Southeast Alaska. Two of the three islands where mountain goat translocations have occurred match genetically with their source population, while Baranof Island appears to have a divergent population, consistent with the hypothesis of an endemic or cryptic population prior to the translocation event. This study highlights the value of considering both the natural and anthropogenic forces when assessing the biogeographic history of a species.
Genome-scale quantification and prediction of pathogenic stop codon readthrough by small molecules
Authors: Toledano Ignasi , Supek Fran , Lehner Ben
Published: August 2023

Premature termination codons (PTCs) cause ∼10-20% of Mendelian diseases and are the major mechanism of tumor suppressor gene inactivation in cancer. A general strategy to alleviate the effects of PTCs would be to promote translational readthrough. Nonsense suppression by small molecules has proven effective in diverse disease models, but translation into the clinic is hampered by ineffective readthrough of many PTCs. Here we directly tackle the challenge of defining drug efficacy by quantifying readthrough of ∼5,800 human pathogenic stop codons by 8 drugs. We find that different drugs promote readthrough of complementary subsets of PTCs defined by local sequence context. This allows us to build interpretable models that accurately predict drug-induced readthrough genome-wide. Accurate readthrough quantification and prediction will empower clinical trial design and the development of personalized nonsense suppression therapies.
Deep learning reduces data requirements and allows real-time measurements in Imaging Fluorescence Correlation Spectroscopy
Authors: Tang Wai Hoh , Sim Shao Ren , Aik Daniel Ying Kia , Nelanuthala Ashwin Venkata Subba , Athilingam Thamarailingam , Röllin Adrian , Wohland Thorsten
Published: August 2023

Imaging Fluorescence Correlation Spectroscopy (Imaging FCS) is a powerful tool to extract information on molecular mobilities, actions and interactions in live cells, tissues and organisms. Nevertheless, several limitations restrict its applicability. First, FCS is data hungry, requiring 50,000 frames at 1 ms time resolution to obtain accurate parameter estimates. Second, the data size makes evaluation slow. Thirdly, as FCS evaluation is model-dependent, data evaluation is significantly slowed unless analytic models are available. Here we introduce two convolutional neural networks (CNNs) –FCSNetandIm-FCSNet– for correlation and intensity trace analysis, respectively.FCSNetrobustly predicts parameters in 2D and 3D live samples.ImFCSNetreduces the amount of data required for accurate parameter retrieval by at least one order of magnitude and makes correct estimates even in moderately defocused samples. Both CNNs are trained on simulated data, are model-agnostic, and allow autonomous, real-time evaluation of Imaging FCS measurements.
Single-molecule analysis of transcription activation: dynamics of SAGA co-activator recruitment
Authors: Jeon Jongcheol , Friedman Larry J. , Seo Hogyu David , Adeleke Augustus , Graham Bria , Patteson Emily , Gelles Jeff , Buratowski Stephen
Published: August 2023

Transcription activators are said to stimulate gene expression by “recruiting” coactivators to promoters, yet this term fits several different kinetic models. To directly analyze dynamics of activator-coactivator interactions, single-molecule microscopy was used to image promoter DNA, a transcription activator, and the Spt-Ada-Gcn5 Acetyltransferase (SAGA) complex within nuclear extract. SAGA readily, but transiently, binds nucleosome-free DNA without activator, while chromatin template association occurs nearly exclusively when activator is present. On both templates, activator increases SAGA association rates by up to an order of magnitude, and dramatically extends its dwell times. These effects reflect direct interactions with the transactivation domain, as VP16 or Rap1 activation domains produce different SAGA dynamics. Despite multiple bromodomains, acetyl-CoA or histone H3/H4 tail acetylation only modestly improves SAGA binding. Unexpectedly, histone acetylation more strongly affects activator residence. Our studies thus reveal two modes of SAGA interaction with the genome: a short-lived activator-independent interaction with nucleosome-free DNA, and a state tethered to promoter-bound transcription activators that can last up to several minutes.
A novel lncRNA PROCA11 regulates cell plasticity in response to androgen deprivation of prostate cancer cells
Authors: Cipolla Rocco , Gabriel Marc , Regin Giorgia Ianese , Piemontese Micaela , Szachnowski Ugo , Firlej Virginie , Pinskaya Marina , Morillon Antonin
Published: August 2023

Long non-coding RNAs (lncRNAs) represent vast and yet poorly characterized family of genes that can fine tune cellular plasticity, thereby allowing the emergence of aggressive therapy-resistant and metastatic cancers. Androgen deprivation therapies (ADT) are commonly used to treat prostate cancer by inactivating the Androgen Receptor (AR). However, castration-resistant prostate cancer (CRPC) with neuroendocrine subtypes (NEPC) often emerge. In this study, we explore the role of lncRNAs in response to androgen deprivation. Using a dynamic prostate cancer cell system mimicking the CRPC and NEPC onset, we identified 15 novel lncRNAs, with PROCA11 standing out as a first-choice candidate, being also highly abundant in high-risk prostate cancer tumors. This majorly nuclear lncRNA is expressed at low levels in androgen-dependent conditions of growth and strongly activated upon hormone withdrawal, preceding neuroendocrine genes and persisting at high levels in neuroendocrine cells. Extensive computational analysis of clinical data and functional studies in cells revealed PROCA11 association with basal-to-luminal transformation of the transcriptomic landscape and activation of metabolic and signaling pathways reminiscent of neurogenesis and of maintenance of AR signaling. We propose that PROCA11 is involved in the intricate circuits regulating cellular plasticity enabling cell survival and proliferation and emergence of the NE phenotype in response to ADT.
Season of transmission of Ross River/Barmah Forest Virus andMycobacterium ulceransclosely align in southeastern Australia, supporting mosquitoes as the vector of Buruli ulcer
Authors: Buultjens Andrew H. , Tay Ee Laine , Yuen Aidan , Friedman N. Deborah , Stinear Timothy P. , Johnson Paul D.R.
Published: August 2023

Ross River Virus and Barmah Forest Virus infections (alphaviruses) have short incubation periods and are transmitted to humans by mosquitoes.Mycobacterium ulceransinfection (Buruli ulcer) has a much longer incubation period and its mode of transmission is contested. We studied the relationship between month of notification of alphavirus infections and Buruli ulcer in the temperate Australian state of Victoria over the six-year period, 2017-2022. Usingcross-correlation, a signal processing technique, we found that a five-month temporal shift in month of Buruli ulcer notification provided optimal alignment with month of alphavirus notification. This closely matches the previously determined 5-month Buruli ulcer incubation period. Inferred transmission of both conditions showed coordinated maxima in summer and autumn and coordinated minima in winter and spring. The close alignment in season of transmission of alphavirus infection and Buruli ulcer in Victoria supports mosquitoes as the primary local vector ofM. ulcerans.
Target-directed microRNA degradation regulates developmental microRNA expression and embryonic growth in mammals
Authors: Jones Benjamin T. , Han Jaeil , Zhang He , Hammer Robert E. , Evers Bret M. , Rakheja Dinesh , Acharya Asha , Mendell Joshua T.
Published: August 2023

MicroRNAs (miRNAs) are post-transcriptional regulators of gene expression that play critical roles in development and disease. Target-directed miRNA degradation (TDMD), a pathway in which miRNAs that bind to specialized targets with extensive complementarity are rapidly decayed, has emerged as a potent mechanism of controlling miRNA levels. Nevertheless, the biological role and scope of miRNA regulation by TDMD in mammals remains poorly understood. To address these questions, we generated mice with constitutive or conditional deletion ofZswim8, which encodes an essential TDMD factor. Loss ofZswim8resulted in developmental defects in the heart and lungs, growth restriction, and perinatal lethality. Small RNA sequencing of embryonic tissues revealed widespread miRNA regulation by TDMD and greatly expanded the known catalog of miRNAs regulated by this pathway. These experiments also uncovered novel features of TDMD-regulated miRNAs, including their enrichment in cotranscribed clusters and examples in which TDMD underlies “arm switching,” a phenomenon wherein the dominant strand of a miRNA precursor changes in different tissues or conditions. Importantly, deletion of two miRNAs, miR-322 and miR-503, rescued growth ofZswim8-null embryos, directly implicating the TDMD pathway as a regulator of mammalian body size. These data illuminate the broad landscape and developmental role of TDMD in mammals.
Does cooperative membership stabilize farm income volatility? Evidence from the pig sector in China
Authors: Ji Chen , Zhuo Ni , Jin Songqing
Published: August 2023

Purpose

Farm income in the agricultural sector is susceptible to natural and market risks. A large body of literature has studied the effects of cooperative membership on household welfare, technical efficiency, productivity and production behavior, yet little has been known about the impact of cooperative membership on farm income volatility. This paper aims to fill this research gap by investigating the relationship between cooperative membership and farm income volatility of Chinese pig farmers and drawing policy implications.

Design/methodology/approach

This paper examines the effect of cooperative membership on farm income volatility, using data from a two-round survey of pig farmers in China. The authors employ an endogenous switching regression model to address the selection bias issues associated with unobserved factors simultaneously affecting farmers' participation in agricultural cooperatives and income earning activities.

Findings

Using household panel from a two-round survey of 193 pig farmers in China, this analysis highlights two key findings: (1) agricultural cooperative membership has significant and positive effect on farm income stability and (2) the impact of cooperative membership on farm income stability varies with production scale.

Originality/value

This research makes two contributions to the literature. First, this study contributes to the scant literature exploring the relationship between agricultural cooperatives and farm income stability. Second, to the best of the authors' knowledge, this is the first study that explores such relationship in a livestock sector. The pig sector in China and around the developing world has been increasingly challenged by multifaceted risks (e.g. price fluctuations, epidemic diseases, environmental regulations), and understanding the role of agricultural cooperatives in farm income stability of pig farmers is of great practical and policy significance.