bluuebunny/crossref_metadata_2025 huggingface dataset:
Structure and flexibility of the DNA polymerase holoenzyme of vaccinia virus
Authors: Burmeister Wim P. , Boutin Laetitia , Balestra Aurelia C. , Gröger Henri , Ballandras-Colas Allison , Hutin Stephanie , Kraft Christian , Grimm Clemens , Böttcher Bettina , Fischer Utz , Tarbouriech Nicolas , Iseni Frédéric
Published: September 2023

The year 2022 was marked by the mpox outbreak caused by human monkeypox virus (MPXV), which is about 98 % identical to vaccinia virus (VACV) at the sequence level regarding the proteins involved in DNA replication. We present the strategy for the production of the VACV DNA polymerase holoenzyme composed of the E9 polymerase associated with its co-factor, the A20-D4 heterodimer, which led to the 3.8 Å cryo-electron microscopy (cryo-EM) structure of the DNA-free form of the holoenzyme. Model building used high-resolution structures of components of the complex and the A20 structure predicted by AlphaFold 2. The structure of E9 does not change in context of the holoenzyme compared to the crystal structure. As for the MPXV holoenzyme, a contact between E9 and D4 is mediated by a cluster of hydrophobic residues. The holoenzyme structure is quite compact and surprisingly similar to the MPXV holoenzyme in presence of a DNA template, with the exception of a movement of the finger domain and the thumb domain, which becomes ordered in presence of DNA. Even in absence of DNA, the VACV holoenzyme structure is too compact for an agreement with SAXS data. This suggests the presence of more open conformations in solution, which are also predicted by Alphafold 2 indicating hinge regions located within A20. Using biolayer interferometry we showed that indeed, the E9-D4 interaction is weak and transient although very important as it has not been possible to obtain viable viruses carrying mutations of key residues in the E9-D4 interface.

Author Summary

The 2022 outbreak of mpox is caused by monkeypox virus closely related to the best studied model, vaccinia virus. Genome replication, which takes place largely autonomously in the cytosol of the infected cell, is still not really understood. Viral DNA synthesis involves a DNA repair enzyme, the uracil-DNA glycosylase D4 linked to the structural protein A20 forming the processivity factor, which in turn binds to E9 forming the complex required for processive DNA synthesis. Here we present the first structure of the vaccinia virus polymerase holoenzyme E9-A20-D4 at 3.8 Å obtained by cryo-electron microscopy. This structure, together with several recent structures from monkeypox virus, provide a static view of the complex with a previously undescribed contact between E9 and D4. Our small-angle scattering data show that other conformations, taking advantage of 2 hinge regions in A20, exist in solution. Using site-directed mutagenesis and binding studies we show that the contact between E9 and D4, which serves to encircle the template strand, is important, but transient. Thus the current model of the orientation of the holoenzyme on the replication fork may not be the only one possible.
Population-based Risk of Psychiatric Disorders Associated with Recurrent CNVs
Authors: Vaez Morteza , Montalbano Simone , Calle Sánchez Xabier , Georgii Hellberg Kajsa-Lotta , Rasekhi Dehkordi Saeid , Dybdahl Krebs Morten , Meijsen Joeri , Shorter John , Byberg-Grauholm Jonas , Mortensen Preben B , Børglum Anders D , Hougaard David M , Nordentoft Merete , Geschwind Daniel H , Buil Alfonso , Schork Andrew J ,   iPSYCH Investigators, Helenius Dorte , Raznahan Armin , Thompson Wesley K , Werge Thomas , Ingason Andrés
Published: September 2023

Recurrent copy number variants (rCNVs) are associated with increased risk of neuropsychiatric disorders but their pathogenic population-level impact is unknown. We provide population-based estimates of rCNV-associated risk of neuropsychiatric disorders for 34 rCNVs in the iPSYCH2015 case-cohort sample (n=120,247).

Most observed significant increases in rCNV-associated risk for ADHD, autism or schizophrenia were moderate (HR:1.42-5.00), and risk estimates were highly correlated across these disorders, the most notable exception being high autism-associated risk with Prader-Willi/ Angelman Syndrome duplications (HR=20.8). No rCNV was associated with significant increase in depression risk. Also, rCNV-associated risk was positively correlated with locus size and gene constraint. Comparison with published rCNV studies suggests that prevalence of some rCNVs is higher, and risk of psychiatric disorders lower, than previously estimated.

In an era where genetics is increasingly being clinically applied, our results highlight the importance of population-based risk estimates for genetics-based predictions.
Wastewater-based surveillance of respiratory syncytial virus epidemic at the national level in Finland
Authors: Länsivaara Annika , Lehto Kirsi-Maarit , Hyder Rafiqul , Luomala Oskari , Lipponen Anssi , Hokajärvi Anna-Maria , Heikinheimo Annamari , Pitkänen Tarja , Oikarinen Sami ,   WastPan Study Group
Published: September 2023

The purpose of this study was to evaluate the potential of wastewater-based surveillance in the monitoring of epidemics at the national level in Finland. To discover the correlation of wastewater data and register data, the 2021–2022 respiratory syncytial virus (RSV) epidemic in Finland was analyzed from wastewater and the Finnish National Infectious Diseases Register. The study was performed using samples that were collected monthly from May 2021 to July 2022 from ten wastewater treatment plants that cover 40% of the Finnish population. Respiratory syncytial virus detection in 24-h composite samples of influent wastewater was performed using RT-qPCR. Respiratory syncytial virus wastewater data were positively correlated with the National Infectious Diseases Register data for the sampling week (correlation coefficient, CC min = 0.412, max = 0.865). Furthermore, the cumulative incidence of respiratory syncytial virus from the sampling week to three weeks afterward was strongly correlated with the wastewater data (CC min = 0.482, max = 0.814), showing the potential of wastewater-based surveillance for use in estimating the course of the epidemic. When the register-based incidence of RSV was at least four cases/100,000 persons/week in the sampling week, it was detected in all wastewater samples. This study showed that wastewater surveillance is useful in the surveillance of respiratory syncytial virus epidemics, and its potential in the surveillance of other epidemics should be explored further.

Synopsis

Environmental surveillance has been previously used to monitor viruses such as poliovirus and SARS-CoV-2. This study shows that wastewater-based surveillance can be used to surveil the respiratory syncytial virus and its epidemics, at a national level.
Psychotherapies and Psychological Support for Individuals Facing Psychological Distress during the COVID-19 Pandemic: A Scoping Review
Authors: Yagihashi Mao , Sakuma Atsushi , Murakami Michio
Published: September 2023

In this scoping review, we investigated psychotherapies and psychological support provided the coronavirus disease 2019 (COVID-19) pandemic to clarify its recipients and the methods employed. We used Scopus and PubMed as the search engines on October 18, 2022, employing specific search terms ("COVID*" AND ("Psychotherap*" OR "psychological support*") AND "psychological distress*"). The initial search yielded 153 articles, of which 18 met the eligibility criteria after two rounds of screening. The distribution of participants ranged from the general population to patients with COVID-19 and those who had recovered. However, no studies of patients with post-COVID-19 sequelae were found. The distribution of the types of and psychological support varied and the use of new technology was suggested. Online interventions comprised the majority of the means of psychotherapies and psychological support. This study suggests that psychotherapies and psychological support during the COVID-19 pandemic were influenced by the social situation.
Statistical significance misuse in public health research: an investigation of the current situation and possible solutions
Authors: Rovetta Alessandro
Published: September 2023

Background

Despite the efforts of leading statistical authorities and experts worldwide and the inherent dangers of interpretative errors in clinical research, misuses of statistical significance remain a common practice in the field of public health. Currently, there is a need to attempt to quantify this phenomenon.

Methods

97 studies were randomly selected within the PubMed database. An evaluation scale for the interpretation and presentation of statistical results (SRPS) was adopted. The maximum achievable score was 4 points. The abstracts and the full texts of the manuscripts were evaluated separately to highlight any differences in presentation. In this regard, a paired Student t-test was implemented.

Results

All studies failed to adopt statistical significance as a continuous measure of compatibility between the model and the test result. The vast majority of them did not provide information on the validation of the model used. However, in most cases, all results were reported in full within the manuscripts. Substantial differences (a-f) between abstracts (a) and full texts (f) were highlighted when applying the SRPS (null hypothesis, P<.0001, a-f=1.2, Cohen’s D=1.7; a-f=0.5 hypothesis, P<.0001, a-f=0.7, Cohen’s D=1.0).

Conclusion

When contextualized in the current scenario, these findings provide evidence of widespread and severe shortcomings in the use and interpretation of statistical significance measures in clinical and public health research during 2023. Therefore, it is essential for academic journals to compulsorily demand higher scientific quality standards.
Views and experiences of young people on using mHealth platforms for sexual and reproductive health services in rural low- and middle-income countries: a qualitative systematic review
Authors: Laar Alexander S. , Harris Melissa L. , Khan Md N , Loxton Deborah
Published: September 2023

In low- and middle-income countries (LMICs), reproductive health programs use mobile health (mHealth) platforms to deliver a broad range of SRH information and services to young people in rural areas. However, young people’s experiences of using mobile phone platforms for SRH services in the rural contexts of LMICs remains unexplored. This review qualitatively explored the experiences and perceptions of young people’s use of mobile phone platforms for SRH information and services. This qualitative evidence synthesis was conducted through a systematic search of online databases: Medline, Embase, CINAHL, PsycInfo and Scopus. We included peer reviewed articles that were conducted between 2000 and 2020 and used qualitative methods. The methodological quality of papers was assessed by two authors using Grading of Recommendations, Assessment, Development and Evaluation (GRADE) and Confidence in Evidence from Reviews of Qualitative research (CERQual) approach with the identified papers synthesized using a narrative thematic analysis approach. The 26 studies included in the review were conducted in a wide range of LMIC rural settings. The studies used seven different types of mHealth platforms in providing access to SRH information and services on contraception, family planning, sexually transmitted infections (STIs) and human immunodeficiency virus (HIV) education. Participant preferences for use of SRH service platforms centred on convenience, privacy and confidentiality, as well as ease and affordability. High confidence was found in the studies preferencing text messaging, voice messaging, and interactive voice response services while moderate confidence was found in studies focused on phone calls. The overall constraint for platforms services included poor and limited network and electricity connectivity (high confidence in the study findings), limited access to mobile phones and mobile credit due to cost, influence from socio-cultural norms and beliefs and community members (moderate confidence in the study findings), language and literacy skills constraints (high confidence in the study findings). The findings provide valuable information on the preferences of mHealth platforms for accessing SRH services among young people in rural settings in LMICs and the quality of available evidence on the topic. As such, the findings have important implications for health policy makers and implementers and mHealth technology platform developers on improving services for sustainable adoption and integration in LMIC rural health systems.
Small protein mediates inhibition of ammonium transport in Methanosarcina mazei – an ancient mechanism?
Authors: Habenicht Tim , Weidenbach Katrin , Velazquez-Campoy Adrian , Buey Ruben M. , Balsera Monica , Schmitz Ruth A.
Published: September 2023

In the past decade, small open reading frames (sORFs) coding for proteins less than 70 amino acids (aa) in length have moved into the focus of Science. sORFs and corresponding small proteins have been recently identified in all three domains of life. However, the majority of small proteins remain functionally uncharacterized. While several bacterial small proteins have already been described, the number of identified and functionally characterized small proteins in archaea is still limited. In this study, we have discovered that the small protein 36 (sP36), which consists of only 61 aa, plays a critical role in regulating nitrogen metabolism inMethanosarcina mazei.The absence of sP36 significantly delays the growth ofM. mazeiwhen transitioning from nitrogen limitation to nitrogen sufficiency, as compared to the wild type. Through ourin vivoexperiments, we have observed that during nitrogen limitation, sP36 is dispersed throughout the cytoplasm; however, upon shifting the cells to nitrogen sufficiency, it relocates to the cytoplasmic membrane. Moreover, in vitro biochemical analysis clearly showed that sP36 interacts with high-affinity with the ammonium transporter AmtB1present in the cytoplasmic membrane during nitrogen limitation, as well as with the PII-like protein GlnK1. Based on our findings, we propose that in response to an ammonium up-shift, sP36 targets the ammonium transporter AmtB1and inhibits its activity by mediating the interaction with GlnK1.

Importance

Small proteins containing fewer than 70 aa, which were previously disregarded due to computational prediction and biochemical detection challenges, have gained increased attention in the scientific community in recent years. However, the number of functionally characterized small proteins, especially in archaea, is still limited. Here, by using biochemical and genetic approaches, we demonstrate a crucial role for the small protein sP36 in the nitrogen metabolism ofM. mazei, regulating the ammonium transporter AmtB1according to nitrogen availability. This regulation might represent an ancient archaeal mechanism of AmtB1inhibition by GlnK, in contrast to the well-studied regulation in bacteria, which depends on covalent modification of GlnK.
The association betweenDioscorea sansibarensisandOrrella dioscoreaeas a model for hereditary leaf symbiosis
Authors: Acar Tessa , Moreau Sandra , Jardinaud Marie-Françoise , Houdinet Gabriella , Maviane-Macia Felicia , De Meyer Frédéric , Hoste Bart , Leroux Olivier , Coen Olivier , le Ru Aurélie , Peeters Nemo , Carlier Aurelien
Published: September 2023

Hereditary, or vertically-transmitted, symbioses affect a large number of animal species and some plants. The precise mechanisms underlying transmission of functions of these associations are often difficult to describe, due to the difficulty in separating the symbiotic partners. This is especially the case for plant-bacteria hereditary symbioses, which lack experimentally tractable model systems. Here, we demonstrate the potential of the leaf symbiosis between the wild yamDioscorea sansibarensisand the bacteriumOrrella dioscoreae(O. dioscoreae) as a model system for hereditary symbiosis.O. dioscoreaeis easy to grow and genetically manipulate, which is unusual for hereditary symbionts. These properties allowed us to design an effective antimicrobial treatment to rid plants of bacteria and generate whole aposymbiotic plants, which can later be re-inoculated with bacterial cultures. Aposymbiotic plants did not differ morphologically from symbiotic plants and the leaf forerunner tip containing the symbiotic glands formed normally even in the absence of bacteria, but microscopic differences between symbiotic and aposymbiotic glands highlight the influence of bacteria on the development of trichomes and secretion of mucilage. This is to our knowledge the first leaf symbiosis where both host and symbiont can be grown separately and where the symbiont can be genetically altered and reintroduced to the host.
Xenomake: a pipeline for processing and sorting xenograft reads from spatial transcriptomic experiments
Authors: Strope Benjamin S , Pendleton Katherine E , Bowie William Z , Echeverria Gloria V , Zhu Qian
Published: September 2023

Xenograft models are attractive models that mimic human tumor biology and permit one to perturb the tumor microenvironment and study its drug response. Spatially resolved transcriptomics (SRT) provide a powerful way to study the organization of xenograft models, but currently there is a lack of specialized pipeline for processing xenograft reads originated from SRT experiments. Xenomake is a standalone pipeline for the automated handling of spatial xenograft reads. Xenomake handles read processing, alignment, xenograft read sorting, quantification, and connects well with downstream spatial analysis packages. We additionally show that Xenomake can correctly assign organism specific reads, reduce sparsity of data by increasing gene counts, while maintaining biological relevance for studies.
Deprivation-induced plasticity in the early central circuits of the rodent visual, auditory, and olfactory systems: a systematic review and meta-analysis of the literature
Authors: Huang Li , Hardyman Francesca , Edwards Megan , Galliano Elisa
Published: September 2023

Activity-dependent neuronal plasticity is crucial for animals to adapt to dynamic sensory environments. Traditionally, research on activity dependent-plasticity has used sensory deprivation approaches in animal models, and it has focused on its effects in primary sensory cortices. However, emerging evidence emphasizes the importance of activity-dependent plasticity both in the sensory organs and in sub-cortical regions where cranial nerves relay information to the brain. Additionally, a critical question arises: do different sensory modalities share common cellular mechanisms for deprivation-induced plasticity at these central entry-points? Furthermore, does the duration of deprivation correlate with specific plasticity mechanisms? This study aims to systematically review and meta-analyse research papers that investigated visual, auditory, or olfactory deprivation in rodents. Specifically, it explores the consequences of sensory deprivation in homologous regions at the first central synapse after the cranial nerve: vision—lateral geniculate nucleus and superior colliculus; audition— ventral and dorsal cochlear nucleus; olfaction—olfactory bulb. The systematic search yielded 91 research papers (39 vision, 22 audition, 30 olfaction), revealing significant heterogeneity in publication trends, experimental methods of inducing deprivation, measures of deprivation-induced plasticity, and reporting, across the three sensory modalities. Nevertheless, despite these methodological differences, commonalities emerged when correlating the plasticity mechanisms with the duration of the sensory deprivation. Following short-term deprivations (up to 1 day) all three systems showed reduced activity levels and increased disinhibition. Medium-term deprivation (1 day to a week) induced greater glial involvement and synaptic remodelling. Long-term deprivation (over a week) predominantly led to macroscopic structural changes including tissue shrinkage and apoptosis. These findings underscore the importance of standardizing methodologies and reporting practices. Additionally, they highlight the value of cross-modals synthesis for understanding how the nervous system, including peripheral, pre-cortical, and cortical areas, respond to and compensate for sensory inputs loss.
Inferring cell trajectories of spatial transcriptomics via optimal transport analysis
Authors: Shen Xunan , Huang Ke , Zuo Lulu , Ye Zhongfei , Li Zeyu , Yu Qichao , Zou Xuanxuan , Wei Xiaoyu , Xu Ping , Jin Xin , Xu Xun , Wu Liang , Zhu Hongmei , Qin Pengfei
Published: September 2023

The integration of cell transcriptomics and spatial coordinates to organize differentiation trajectories remains a challenge. Here we introduce spaTrack, a trajectory inference method using optimal transport to incorporate both transcriptomics and distance of spatial transcriptomics sequencing data into transition costs. spaTrack could construct fine spatial trajectories reflecting the true differentiation topology, as well as trace cell dynamics across multiple samples with temporal intervals. To capture the dynamic drivers, spaTrack models the cell fate as a function of expression profile along temporal intervals driven by transcription factors. Applying spaTrack, we successfully disentangle spatiotemporal trajectories of axolotl telencephalon regeneration and mouse midbrain development. Furthermore, we uncover diverse malignant lineages expanding in a primary tumor. One of the lineages with upregulated extracellular matrix organization implants to the metastatic site and subsequently colonizes to a secondary tumor. Overall, spaTrack greatly facilitates trajectory inference from spatial transcriptomics, providing insights in cell differentiation of broad areas.
Astrocytic uptake of posttranslationally modified amyloid-β leads to endolysosomal system disruption and induction of pro-inflammatory signaling
Authors: Wirth S , Schlößer A , Beiersdorfer A , Schweizer M , Lohr C , Grochowska KM
Published: September 2023

The disruption of astrocytic catabolic processes contributes to the impairment of amyloid-β (Aβ) clearance, neuroinflammatory signaling, and the loss of synaptic contacts in late-onset Alzheimer’s disease (AD). While it is known that the posttranslational modifications of Aβ have significant implications on biophysical properties of the peptides, their consequences for clearance impairment are not well understood. It was previously shown that N-terminally pyroglutamylated Aβ3(pE)-42, a significant constituent of amyloid plaques, is efficiently taken up by astrocytes, leading to the release of pro-inflammatory cytokine tumor necrosis factor α (TNFα) and synapse loss. Here we report that Aβ3(pE)-42, but not Aβ1-42, gradually accumulates within the astrocytic endolysosomal system, disrupting this catabolic pathway and inducing formation of heteromorphous vacuoles. This accumulation alters lysosomal kinetics and lysosome-dependent calcium signaling, and upregulates lysosomal stress response. These changes correlate with the upregulation of glial fibrillary acidic protein (GFAP) and increased activity of nuclear factor kappa-light-chain-enhancer of activated B cells (NFᴋB). Treatment with a lysosomal protease inhibitor, E64, rescues GFAP upregulation, NFᴋB activation, and synapse loss, indicating that abnormal lysosomal protease activity is upstream of pro-inflammatory signaling and related synapse loss. Collectively, our data suggest that Aβ3(pE)-42-induced disruption of the astrocytic endolysosomal system leads to cytoplasmic leakage of lysosomal proteases, promoting pro-inflammatory signaling and synapse loss, hallmarks of AD-pathology.
Regulation potential of transcribed simple repeated sequences in developing neurons
Authors: Chung Tek Hong , Zhuravskaya Anna , Makeyev Eugene V.
Published: September 2023

Simple repeated sequences (SRSs), defined as tandem iterations of microsatellite- to satellite-sized DNA units, occupy a substantial part of the human genome. Some of these elements are known to be transcribed in the context of repeat expansion disorders. Mounting evidence suggests that the transcription of SRSs may also contribute to normal cellular functions. Here, we used genome-wide bioinformatics approaches to systematically examine SRS transcriptional activity in cells undergoing neuronal differentiation. We identified thousands of long noncoding RNAs containing >200-nucleotide-long SRSs (SRS-lncRNAs), with hundreds of these transcripts significantly upregulated in the neural lineage. We show that SRS-lncRNAs often originate from telomere-proximal regions and that they have a strong potential to form multivalent contacts with a wide range of RNA-binding proteins. Our analyses also uncovered a cluster of neurally upregulated SRS-lncRNAs encoded in a centromere-proximal part of chromosome 9, which underwent an evolutionarily recent segmental duplication. Using a newly established in vitro system for rapid neuronal differentiation of induced pluripotent stem cells, we demonstrate that at least some of the bioinformatically predicted SRS-lncRNAs, including those encoded in the segmentally duplicated part of chromosome 9, indeed increase their expression in developing neurons to readily detectable levels. These data suggest that many SRSs may be expressed in a cell type and developmental stage-specific manner, providing a valuable resource for further studies focused on the functional consequences of SRS-lncRNAs in the normal development of the human brain.
Distractor-specific control adaptation in multidimensional environments
Authors: Gheza Davide , Kool Wouter
Published: September 2023

Goal-directed behavior requires humans to constantly manage and switch between multiple, independent, and conflicting, sources of information. Conventional cognitive control tasks, however, only feature one task and one source of distraction. Therefore, it is unclear how control is allocated in multidimensional environments. To address this question, we developed a multi-dimensional task-set interference paradigm, in which people need to manage distraction from three independent dimensions. We use this task to test whether people adapt to prior conflict by enhancing task-relevant information or suppressing task-irrelevant information. Three experiments provided strong evidence for the latter hypothesis. Moreover, control adaptation was highly dimension-specific. Conflict from a given dimension only affected processing of that same dimension on subsequent trials, with no evidence for generalization. A new neural network model shows that our results can only be simulated when including multiple independent conflict-detector units. Our results call for an update to classic models of cognitive control, and their neurocomputational underpinnings.
FYN regulates aqueous humor outflow and IOP through the phosphorylation of VE-cadherin
Authors: Kizhatil Krishnakumar , Clark Graham , Sunderland Daniel , Bhandari Aakriti , Horbal Logan , Balasubramanian Revathi , John Simon
Published: September 2023

The exact sites and molecules that determine resistance to aqueous humor drainage and control intraocular pressure (IOP) need further elaboration. Proposed sites include the inner wall of Schlemms’s canal and the juxtacanalicular trabecular meshwork ocular drainage tissues. The adherens junctions (AJs) of Schlemm’s canal endothelial cells (SECs) must both preserve the blood-aqueous humor (AQH) barrier and be conducive to AQH drainage. How homeostatic control of AJ permeability in SC occurs and how such control impacts IOP is unclear. We hypothesized that mechano-responsive phosphorylation of the junctional molecule VE-CADHERIN (VEC) by SRC family kinases (SFKs) regulates the permeability of SEC AJs. We tested this by clamping IOP at either 16 mmHg, 25 mmHg, or 45 mmHg in mice and then measuring AJ permeability and VEC phosphorylation. We found that with increasing IOP: 1) SEC AJ permeability increased, 2) VEC phosphorylation was increased at tyrosine-658, and 3) SFKs were activated at the AJ. Among the two SFKs known to phosphorylate VEC, FYN, but not SRC, localizes to the SC. Furthermore, FYN mutant mice had decreased phosphorylation of VEC at SEC AJs, dysregulated IOP, and reduced AQH outflow. Together, our data demonstrate that increased IOP activates FYN in the inner wall of SC, leading to increased phosphorylation of AJ VEC and, thus, decreased resistance to AQH outflow. These findings support a crucial role of mechanotransduction signaling in IOP homeostasis within SC in response to IOP. These data strongly suggest that the inner wall of SC partially contributes to outflow resistance.
A short course of antibiotics selects for persistent resistance in the human gut
Authors: Yaffe Eitan , Dethlefsen Les , Patankar Arati V. , Gui Chen , Holmes Susan , Relman David A.
Published: September 2023

Understanding the relationship between antibiotic use and the evolution of antimicrobial resistance is vital for effective antibiotic stewardship, yet animal models andin vitroexperiments poorly replicate real-world conditions. To elucidate how resistance evolvesin vivo, we exposed 60 human subjects to ciprofloxacin and used longitudinal stool samples and a new computational method to assemble genomes for 5665 populations of commensal bacterial species within subjects. Analysis of 2.27M polymorphic sequence variants revealed 513 populations that underwent selective sweeps. We found convergent evolution focused on DNA gyrase and evidence of dispersed selective pressure at other genomic loci. Nearly 10% of susceptible bacterial populations evolved towards resistance through sweeps that involved mutations in a specific amino acid in gyrase. Evolution towards resistance was predicted by population abundances before and during the exposure. 89% of gyrase sweeps and the majority of all sweeps persisted more than 10 weeks. This work quantifies the direct relationship between antibiotic usage and the evolution of resistance within the gut communities of individual human hosts.
Multi-method quantification of acetyl-CoA and further acyl-CoA species in normal and ischemic rat liver
Authors: Tokarska-Schlattner Malgorzata , Zeaiter Nour , Cunin Valérie , Attia Stéphane , Meunier Cécile , Kay Laurence , Achouri Amel , Hiriart-Bryant Edwige , Couturier Karine , Tellier Cindy , Harras Abderrafek El , Elena-Herrmann Bénédicte , Khochbin Saadi , Gouellec Audrey Le , Schlattner Uwe
Published: September 2023

Thioesters of coenzyme A (CoA) carrying different acyl chains (acyl-CoAs) are central intermediates of many metabolic pathways and donor molecules for protein lysine acylation. Acyl-CoA species largely differ in cellular concentrations and physico-chemical properties, rendering their analysis challenging. Here we compare several approaches to quantify cellular acyl-CoAs concentrations in normal and ischemic rat liver, using HPLC and LC-MS/MS for multi-acyl-CoA analysis, as well as NMR, fluorimetric and spectrophotometric techniques for quantification of acetyl-CoA. In particular, we describe a simple LC-MS/MS protocol that is suitable for the relative quantification of short and medium chain acyl-CoAs species. We show that ischemia induces specific changes in the short chain acyl-CoA relative concentrations, while mild ischemia (1-2 min), although reducing succinyl-CoA, has little effects on acetyl-CoA, and even increases some acyl-CoAs species upstream of the tricarboxylic acid cycle. In contrast, advanced ischemia (5-6 min) also reduces acetyl-CoA levels. Our approach provides the keys to accessing the acyl-CoA metabolome for a more in-depth analysis of metabolism, protein acylation and epigenetics.
UBC13-mediated template switching promotes replication stress resistance in FBH1-deficient cells
Authors: Hawks Alexandra L. , Bergmann Amy , McCraw Tyler J. , Mason Jennifer M.
Published: September 2023

The proper resolution of DNA damage during replication is essential for genome stability. FBH1, a UvrD, helicase plays crucial roles in the DNA damage response. FBH1 promotes double strand break formation and signaling in response to prolonged replication stress to initiate apoptosis. Human FBH1 regulates RAD51 to inhibit homologous recombination. A previous study suggested that mis-regulation of RAD51 may contribute to replication stress resistance in FBH1-deficient cells, but the underlying mechanism remains unknown. Here, we provide direct evidence that RAD51 promotes replication stress resistance in FBH1-deficient cells. We demonstrate inhibition of RAD51 using the small molecule, B02, partially rescues double strand break signaling in FBH1-deficient cells. We show that inhibition of only the strand exchange activity of RAD51 rescues double strand break signaling in FBH1 knockout cells. Finally, we show that depletion of UBC13, a E2 protein that promotes RAD51-dependent template switching, rescues double strand break formation and signaling sensitizing FBH1- deficient cells to replication stress. Our results suggest FBH1 regulates template switching to promote replication stress sensitivity.
FRET-based sensor for measuring adenine nucleotide binding to AMPK
Authors: Nahed Roland Abi , Pelosse Martin , Aulicino Francesco , Cottaz Florine , Berger Imre , Schlattner Uwe
Published: September 2023

AMP-activated protein kinase (AMPK) has evolved to detect a critical increase in cellular AMP/ATP and ADP/ATP concentration ratios as a signal for limiting energy supply. Such energy stress then leads to AMPK activation and downstream events that maintain cellular energy homeostasis. AMPK activation by AMP, ADP or pharmacological activators involves a conformational switch within the AMPK heterotrimeric complex. We have engineered an AMPK-based sensor, AMPfret, which translates the activating conformational switch into a fluorescence signal, based on increased fluorescence resonance energy transfer (FRET) between donor and acceptor fluorophores. Here we describe how this sensor can be used to analyze direct AMPK activation by small moleculesin vitrousing a fluorimeter, or to estimate changes in the energy state of cells using standard fluorescence or confocal microscopy.
SPAG7 deletion causes intrauterine growth restriction, resulting in adulthood obesity and metabolic dysfunction
Authors: Flaherty Stephen E , Bezy Olivier , Paulhus Brianna LaCarubba , Song LouJin , Piper Mary , Pang Jincheng , Park Yoson , Asano Shoh , Lien Yu-Chin , Griffin John D , Robertson Andrew , Opsahl Alan , Hirenallur-Shanthappa Dinesh , Ahn Youngwook , Pashos Evanthia , Simmons Rebecca A , Birnbaum Morris J , Wu Zhidan
Published: September 2023

From a forward mutagenetic screen to discover mutations associated with obesity, we identified mutations in thespag7gene linked to metabolic dysfunction in mice. Here we show that SPAG7 KO mice are born smaller and develop obesity and glucose intolerance in adulthood. This obesity does not stem from hyperphagia, but a decrease in energy expenditure. The KO animals also display reduced exercise tolerance and muscle function due to impaired mitochondrial function. Furthermore, SPAG7-deficiency in developing embryos leads to intrauterine growth restriction, brought on by placental insufficiency, likely due to abnormal development of the placental junctional zone. This insufficiency leads to loss of SPAG7-deficient fetuses in utero and reduced birth weights of those that survive. We hypothesize that a “thrifty phenotype” is ingrained in SPAG7 KO animals during development that leads to adult obesity. Collectively, these results indicate that SPAG7 is essential for embryonic development and energy homeostasis later in life.
Arthropod communities associated with gall-inducingAciurina bigeloviaeandAciurina trixa(Diptera: Tephritidae) in New Mexico
Authors: Baine Quinlyn , Casares Emily E , Hughes Daniel W.W. , Martinson Vincent G , Martinson Ellen O
Published: September 2023

Insect-induced galls are novel structures that serve as habitat to whole communities of associate arthropods that include predators, parasitoids and inquilines. Galling insects are generally under-described, but their associate communities, which can include many specialist organisms, are virtually unknown, particularly in the southwest United States.Aciurina bigeloviae(Cockerell 1890) andAciurina trixaCurran 1932 are unusually common and abundant galling flies in New Mexico. The 2 species are sister and occur in sympatric areas but have distinct gall morphologies. We reared all arthropods from 3800 galls from 14 sites in the northern and central regions of the state and as a result characterized the complete communities of both species, including barcode sequences and eclosion phenology. We also investigate interactions ofA. trixagalls with the abundant inquiline weevilAnthonomus cycliferusFall 1913 and find no measurable effect of inquiline abundance on the size of the emerged adult fly or gall. The total species count is 24 and includes 6 guilds; bothA. bigeloviaeandA. trixacommunities are richer and more complex than other documented Tephritidae-Asteraceae galling systems. This study highlights the potential of galling insects as ecosystem engineers to maintain large, rich and multi-trophic communities.
Enriched Single-Nucleus RNA-Sequencing reveals unique attributes of distal convoluted tubule cells
Authors: Su Xiao-Tong , Reyes Jeremiah V. , Lackey Anne E. , Maeoka Yujiro , Cornelius Ryan J. , McCormick James A. , Yang Chao-Ling , Jung Hyun Jun , Welling Paul A. , Nelson Jonathan W. , Ellison David H.
Published: September 2023

Background

The distal convoluted tubule (DCT) comprises two subsegments, DCT1 and DCT2, with different functional and molecular characteristics. The functional and molecular distinction between these segments, however, has been controversial.

Methods

To understand the heterogeneity within the DCT population with better clarity, we enriched for DCT nuclei by using a mouse line combining “Isolation of Nuclei TAgged in specific Cell Types” and NCC (sodium chloride cotransporter)-driven inducible Cre recombinase. We sorted the fluorescently labeled DCT nuclei using Fluorescence-Activated Nucleus Sorting, and performed single nucleus transcriptomics.

Results

Among 25,183 DCT cells, 70% were from DCT1 and 30% from DCT2. Additionally, there was a small population (<1%) enriched in proliferation-related genes, such asTop2a, Cenpp, and Mki67.Both DCT1 and DCT2 express NCC, magnesium transport genes are more abundant along DCT1; whereas calcium, electrogenic sodium and potassium transport genes are more abundant along DCT2. The transition between these two segments are gradual with a transitional zone where DCT1 and DCT2 cells are interspersed. The expression of the homeobox genes is not consistent between all DCT cells, suggesting that they develop along different trajectories.

Conclusion

Transcriptomics analysis of an enriched rare cell population using genetically targeted approach offers better clarification of the function and classification. The DCT segment is short, yet, can be separated into two sub-types that serve distinct functions, and are speculated to derive from different origins during development.

Significance Statement

High-resolution snRNAseq data indicate a clear separation between primary sites of calcium and magnesium handling within DCT. Both DCT1 and DCT2 expressSlc12a3, but these subsegments serve distinctive functions, with more abundant magnesium handling genes along DCT1 and more calcium handling genes along DCT2. The data also provides insight into the plasticity of the distal nephron-collecting duct junction, formed from cells of separate embryonic origins. By focusing/changing gradients of gene expression, the DCT can morph into different physiological cell states on demand.
Transcranial magnetic stimulation of primary motor cortex elicits a site-specific immediate transcranial evoked potential
Authors: Beck Mikkel Malling , Christiansen Lasse , Madsen Mads Alexander Just , Jadidi Armita Faghani , Vinding Mikkel Christoffer , Thielscher Axel , Bergmann Til Ole , Siebner Hartwig Roman , Tomasevic Leo
Published: September 2023

Background

Transcranial evoked potentials (TEPs) measured via electroencephalography (EEG) are widely used to study the cortical responses to transcranial magnetic stimulation (TMS). Immediate transcranial evoked potentials (i-TEPs) have been obscured by pulse and muscular artifacts. Thus, the TEP peaks that are commonly reported have latencies that are too long to be caused by direct excitation of cortical neurons.

Methods

In 14 healthy individuals, we recorded i-TEPs evoked by a single biphasic TMS pulse targeting the primary motor hand area (M1HAND) or parietal or midline control sites. Sampling EEG at 50 kHz enabled us to reduce the duration of the TMS pulse artifact to a few milliseconds, while minor adjustments of the TMS coil tilt or position enabled us to avoid cranial muscular twitches during the experiment.

Results

We observed an early positive EEG deflection starting after approx. 2 ms combined with a series of peaks with an inter-peak interval of ∼1.1-1.4 ms in electrodes overlying the stimulated sensorimotor region. This multi-peak i-TEP response was only evoked by TMS of the M1HANDregion and was modified by changes in stimulation intensity and current direction.

Discussion

Single-pulse TMS of the M1HANDevokes an immediate local multi-peak response at the cortical site of stimulation. Our results suggest that the observed i-TEP patterns are genuine cortical responses evoked by TMS caused by synchronized excitation of pyramidal neurons in the targeted precentral cortex. This notion needs to be corroborated in future studies, including further investigations into the potential contribution of instrumental or physiological artifacts.
Structural mechanisms for VMAT2 inhibition by tetrabenazine
Authors: Dalton Michael P. , Cheng Mary Hongying , Bahar Ivet , Coleman Jonathan A.
Published: September 2023

The vesicular monoamine transporter 2 (VMAT2) is a proton-dependent antiporter responsible for loading monoamine neurotransmitters into synaptic vesicles. Dysregulation of VMAT2 can lead to several neuropsychiatric disorders including Parkinson’s disease and schizophrenia. Furthermore, drugs such as amphetamine and MDMA are known to act on VMAT2, exemplifying its role in the mechanisms of actions for drugs of abuse. Despite VMAT2’s importance, there remains a critical lack of mechanistic understanding, largely driven by a lack of structural information. Here we report a 3.1 Å resolution cryo-EM structure of VMAT2 complexed with tetrabenazine (TBZ), a non-competitive inhibitor used in the treatment of Huntington’s chorea. We find TBZ interacts with residues in a central binding site, locking VMAT2 in an occluded conformation and providing a mechanistic basis for non-competitive inhibition. We further identify residues critical for cytosolic and lumenal gating, including a cluster of hydrophobic residues which are involved in a lumenal gating strategy. Our structure also highlights three distinct polar networks that may determine VMAT2 conformational dynamics and play a role in proton transduction. The structure elucidates mechanisms of VMAT2 inhibition and transport, providing insights into VMAT2 architecture, function, and the design of small-molecule therapeutics.
Placebo treatment entails resource-dependent downregulation of negative inputs
Authors: Rauh Jonas , Mostauli Arasch , Gamer Matthias , Büchel Christian , Rief Winfried , Brassen Stefanie
Published: September 2023

Background

Clinical trials of antidepressants show improvements in placebo groups of up to 80% compared to the real treatment arm. The mechanism underlying this clinically important effect has been linked to expectation induced goal-directed control. Here, we investigated how cognitive resources influence the effects of positive expectations on emotional processing.

Methods

Forty-nine healthy volunteers participated in a cross-over fMRI study, in which expectancy of positive emotional changes was induced by an alleged oxytocin nasal spray combined with verbal instruction. Participants performed a spatial cueing paradigm that manipulated the attention to emotional face distractors in the scanner and were characterized regarding their general ability to control attention.

Results

Behavioral findings showed placebo treatment to improve mood and to reduce distractibility by fearful over happy faces, specifically when more attentional resources were available to process faces. This aligned with neural changes in activation and functional coupling in lateral prefrontal-limbic networks indicating an expectation induced top-down regulation of aversive inputs. In addition, behavioral effects and prefrontal-parietal engagement directly correlated with trait ability to control attention.

Conclusions

Our data show that placebo treatment combined with verbal instruction alone can improve mood and recruit top-down attentional selection. Changes in emotional processing critically depended on attentional context and individual control ability (i.e., contextual and general resources). These findings may be particularly relevant in patients with major depressive disorder, who often demonstrate a negativity bias and in whom placebo effects by verbal instructions alone may be limited due to reduced cognitive control capacity.
CREBBP lysine acetyltransferase domain mutations create zombie enzymes that alter chromatin loading dynamics and prevent EP300 redundancy
Authors: Yang Haopeng , Zhang Wenchao , Ravanmehr Vida , Mechaly Ariel , Regad Leslie , Haouz Ahmed , Chen Ruidong , Henderson Jared M. , Rojas Estela , Wilson Ashley , Parsons Sydney , Nastoupil Loretta , Davis R. Eric , Deng Qing , Samaniego Felipe , Rodrigues-Lima Fernando , Green Michael R.
Published: September 2023

Epigenetic regulation of gene expression is fundamental to cell state transitions. A prominent example of this is the epigenetically-regulated generation of transitory cells states within the germinal center (GC) reaction that are required for humoral immunity. The deregulation of these processes through somatic mutations can give rise to B-cell lymphoma, thus studying these mutations can provide insight into fundamental mechanisms underlying epigenetic regulation of gene expression as well as GC B-cell and lymphoma biology. Here we show that classes of mutations in theCREBBPlysine acetyltransferase gene result in different structure, catalytic activity and function. We discover that CREBBP and its paralog EP300 are dynamically and reciprocally loaded onto chromatin during a signal-responsive cell state transitions within the GC. Mutation ofCREBBPin GC-derived lymphoma interrupt signal-responsive CREBBP loading onto chromatin, while also inhibiting EP300 redundancy. These observations lead to a model in which CREBBP or EP300 loss of function through acetyltransferase domain mutations, which occur frequently in both hematopoietic and solid tumors, create a zombie enzyme that retards the function of the active paralog by binding limiting concentrations of transcription factor substrate and preventing signal-responsive enhancer activation.