bluuebunny/crossref_metadata_2025 huggingface dataset:
Activator-blocker model of transcriptional regulation by pioneer-like factors
Authors: Riesle Aileen Julia , Gao Meijiang , Rosenblatt Marcus , Hermes Jacques , Hass Helge , Gebhard Anna , Veil Marina , Grüning Björn , Timmer Jens , Onichtchouk Daria
Published: January 2023

Zygotic genome activation (ZGA) in the development of flies, fish, frogs and mammals depends on pioneer-like transcription factors (TFs). Those TFs create open chromatin regions, promote histone acetylation on enhancers, and activate transcription. Here, we use the panel of single, double and triple mutants for zebrafish genome activators Pou5f3, Sox19b and Nanog, multi-omics and mathematical modeling to investigate the combinatorial mechanisms of genome activation. We show that Pou5f3 and Nanog act differently on synergistic and antagonistic enhancer types. Pou5f3 and Nanog both bind as pioneer-like TFs on synergistic enhancers, promote histone acetylation and activate transcription. Antagonistic enhancers are activated by binding of one of these factors. The other TF binds as non-pioneer-like TF, competes with the activator and blocks all its effects, partially or completely. This activator-blocker mechanism mutually restricts widespread transcriptional activation by Pou5f3 and Nanog and prevents premature expression of late developmental regulators in the early embryo.
Pressure and ultrasound activate mechanosensitive TRAAK K+channels through increased membrane tension
Authors: Sorum Ben , Docter Trevor , Panico Vincent , Rietmeijer Robert A. , Brohawn Stephen G.
Published: January 2023

TRAAK is a mechanosensitive two-pore domain K+(K2P) channel found in nodes of Ranvier within myelinated axons. It displays low leak activity at rest and is activated up to one hundred-fold by increased membrane tension. Structural and functional studies have led to physical models for channel gating and mechanosensitivity, but no quantitative analysis of channel activation by tension has been reported. Here, we use simultaneous patch-clamp recording and fluorescent imaging to determine the tension response characteristics of TRAAK. TRAAK shows high sensitivity and a broad response to tension spanning nearly the entire physiologically relevant tension range. This graded response profile distinguishes TRAAK from similarly low-threshold mechanosensitive channels Piezo1 and MscS, which activate in a step-like fashion over a narrow tension range. We further use patch imaging to show that ultrasonic activation of TRAAK and MscS is due to increased membrane tension. Together, these results provide mechanistic insight into TRAAK tension gating, a framework for exploring the role of mechanosensitive K+channels at nodes of Ranvier, and biophysical context for developing ultrasound as a mechanical stimulation technique for neuromodulation.
Uptake-independent killing of macrophages by extracellular aggregates ofMycobacterium tuberculosisis ESX-1 and PDIM-dependent
Authors: Toniolo Chiara , Dhar Neeraj , McKinney John D.
Published: January 2023

Mycobacterium tuberculosis(Mtb) infection is initiated by inhalation of small numbers of bacteria into lung alveoli, where they are phagocytosed by resident macrophages. Intracellular replication ofMtbleads to death of the infected macrophages, release of bacterial aggregates, and rapid growth of the extracellular aggregates on host-cell debris. Here, we show that extracellularMtbaggregates can evade phagocytosis by killing macrophages in a contact-dependent but uptake-independent manner. We use single-cell time-lapse fluorescence microscopy to show that contact with extracellularMtbaggregates triggers macrophage plasma membrane perturbation, cytoplasmic calcium accumulation, and pyroptotic cell death. These effects depend on theMtbtype VII secretion system ESX-1, however, this system alone cannot induce calcium accumulation and macrophage death in the absence of theMtbsurface-exposed lipid phthiocerol dimycocerosate. Unexpectedly, we found that ESX-1-mediated secretion of the EsxA/EsxB virulence factors is not required for uptake-independent killing of macrophages after contact with extracellularMtbaggregates. In the absence of EsxA/EsxB secretion, killing is mediated by the 50-kDa isoform of the ESX-1-secreted protein EspB, while blocking secretion of both EsxA/EsxB and processed EspB reduces killing to background levels. Treatment with a small-molecule ESX-1 inhibitor reduces uptake-independent killing of macrophages byMtbaggregates, suggesting that novel therapies targeting this anti-phagocytic mechanism could prevent the propagation of extracellular bacteria within the lung.

Significance statement

Mycobacterium tuberculosis(Mtb) can survive inside the lung macrophages that normally provide the first line of defense against bacterial infections. Intracellular replication ofMtbultimately results in the death and lysis of infected macrophages, allowing the bacteria to spread to other cells and propagate the infection. Our study shows that extracellularMtbaggregates that form on the debris of dead host cells can induce macrophage death in a contact-dependent but uptake-independent manner, allowing the bacteria to evade the host defenses associated with uptake by macrophages. Killing of macrophages by extracellularMtbaggregates is driven by theMtbESX-1 secretion system and the surface-exposed lipid phthiocerol dimycocerosate. Our results suggest that novel drugs targetingMtbfactors required for host-cell killing by extracellularMtbaggregates may reduce bacterial spreading and expansion of necrotic tuberculosis lesions, which are known to be poorly penetrated by conventional antibiotics.
Modulation and Neural Correlates of Postmating Sleep Plasticity inDrosophilaFemales
Authors: Duhart José M. , Buchler Joseph R. , Inami Sho , Kennedy Kyle J. , Jenny B. Peter , Afonso Dinis J.S. , Koh Kyunghee
Published: January 2023

Sleep is essential, but animals may forgo sleep to engage in other critical behaviors, such as feeding and reproduction. Previous studies have shown that female flies show decreased sleep after mating, but our understanding of the process is limited. Here, we report that postmating nighttime sleep loss is modulated by diet and sleep deprivation, demonstrating a complex interaction among sleep, reproduction, and diet. We also report that female-specific pC1 neurons and sleep-promoting dorsal fan-shaped body (dFB) neurons are required for postmating sleep plasticity. Activating pC1 neurons leads to sleep suppression on standard fly culture media but has little sleep effect on sucrose-only food. Published connectome data suggest indirect, inhibitory connections among pC1 subtypes. Using calcium imaging, we show that activating the pC1e subtype inhibits dFB neurons. We propose that pC1 and dFB neurons integrate the mating status, food context, and sleep drive to modulate postmating sleep plasticity.

Highlights

Diet and sleep drive modulate female nighttime postmating sleep loss

Female-specific pC1 neurons are required for postmating sleep loss

Sleep-promoting dFB-projecting neurons are required for postmating sleep loss

Activating pC1 subtypes promotes wakefulness and inhibits dFB-projecting neurons

eTOC blurb

Animals evaluate environmental conditions and internal states to make behavioral choices. Duhart et al. show that the decision to stay awake after mating inDrosophilafemales is modulated by food composition and sleep history and mediated by female-specific pC1 neurons acting upstream of the dFB sleep center.
Population demographic history and evolutionary rescue: influence of a bottleneck event
Authors: Olazcuaga Laure , Lincke Beatrice , Delacey Sarah , Durkee Lily F. , Melbourne Brett A. , Hufbauer Ruth A.
Published: January 2023

Rapid environmental change presents a significant challenge to the persistence of natural populations. Rapid adaptation that increases population growth, enabling populations that declined following severe environmental change to grow and avoid extinction, is called evolutionary rescue. Numerous studies have shown that evolutionary rescue can indeed prevent extinction. Here, we extend those results by considering the demographic history of populations. To evaluate how demographic history influences evolutionary rescue, we created 80 populations of red flour beetle,Tribolium castaneum, with three classes of demographic history: diverse populations that did not experience a bottleneck, and populations that experienced either an intermediate or a strong bottleneck. We subjected these populations to a new and challenging environment for six discrete generations and tracked extinction and population size. Populations that did not experience a bottleneck in their demographic history avoided extinction entirely, while more than 20% of populations that experienced an intermediate or strong bottleneck went extinct. Similarly, among the extant populations at the end of the experiment, adaptation increased the growth rate in the novel environment the most for populations that had not experienced a bottleneck in their history. Taken together, these results highlight the importance of considering the demographic history of populations to make useful and effective conservation decisions and management strategies for populations experiencing environmental change that pushes them toward extinction.
Urinary Glycosaminoglycans are Associated with Recurrent UTI and Urobiome Ecology in Postmenopausal Women
Authors: Neugent Michael L. , Hulyalkar Neha V. , Kumar Ashwani , Xing Chao , Zimmern Philippe E. , Shulaev Vladimir , De Nisco Nicole J.
Published: January 2023

Glycosaminoglycans (GAGs) are linear, negatively charged polysaccharides composed of repeating disaccharide units of uronic acid and amino sugars. The luminal surface of the bladder epithelium is coated with a GAG layer. These urothelial GAGs are thought to provide a protective barrier and serve as a potential interaction site with the urinary microbiome (urobiome). Previous studies have profiled urinary GAG composition in mixed cohorts, but the urinary GAG composition in postmenopausal women remains undefined. To investigate the relationship between GAGs and recurrent UTI (rUTI), we profiled urinary GAGs in a controlled cohort of postmenopausal women. We found that chondroitin sulfate (CS) is the major urinary GAG in postmenopausal women and that urinary CS was elevated in women with active rUTI. We also associated urinary GAGs with urobiome composition and identified bacterial species that significantly associated with urinary GAG concentration.Corynebacterium amycolatum, Porphyromonas somerae, andStaphylococcus pasteuriwere positively associated with heparin sulfate or hyaluronic acid and bacterial species associated with vaginal dysbiosis were negatively correlated to urinary CS. Altogether, this work defines changes in urinary GAG composition associated with rUTI and identifies new associations between urinary GAGs and the urobiome that may play a role in rUTI pathobiology.
DNA metabarcoding using newrbcLandITS2metabarcodes collectively enhance detection efficiency of medicinal plants in single and polyherbal formulations
Authors: Travadi Tasnim , Shah Abhi P. , Pandit Ramesh , Sharma Sonal , Joshi Chaitanya , Joshi Madhvi
Published: January 2023

With the widespread adoption of barcoding and next-generation sequencing, metabarcoding is emerging as a potential tool for detecting labelled and unlabelled plant species in herbal products. In this study, newly designedrbcLandITS2metabarcode primers were validated for metabarcoding using in-house mock controls of medicinal plant gDNA pools and biomass pools. The applicability of the multi-barcode sequencing approach was evaluated on 17 single drugs and 15 polyherbal formulations procured from the Indian market. TherbcLmetabarcode demonstrated detection efficiencies of 86.7% and 71.7% in gDNA plant pools and biomass mock controls, respectively, while theITS2metabarcode demonstrated 82.2% and 69.4%. In the gDNA plant pool and biomass pool mock controls, the cumulative detection efficiency increased by 100% and 90%, respectively. A total of 79% cumulative detection efficiency of both metabarcodes was observed in single drugs, while 76.3% was observed in polyherbal formulations. An average fidelity of 83.6% was observed for targeted plant species present within mock controls as well as in herbal formulations. Our results demonstrated the applicability of multilocus strategies in metabarcoding as a potential tool for detecting labelled and unlabelled plant species in herbal formulations.
Candida albicansoropharyngeal infection is an exception to iron-based nutritional immunity
Authors: Solis Norma V. , Wakade Rohan S. , Filler Scott G. , Krysan Damian J.
Published: January 2023

Candida albicansis a commensal of the human gastrointestinal tract and one of the most causes of human fungal disease, including mucosal infections such as oropharyngeal candidiasis and disseminated infections of the bloodstream and deep organs. We directly compared the in vivo transcriptional profile ofC. albicansduring oral infection and disseminated infection of the kidney to identify niche specific features. Although the expression of a set of environmentally responsive genes were correlated in the two infection sites (Pearson R2, 0.6), XXX genes were differentially expressed. Virulence associated genes such as hyphae-specific transcripts were expressed similarly in the two sites. Genes expressed during growth in a poor carbon source (ACS1andPCK1) were upregulated in oral tissue relative to kidney. Most strikingly,C. albicansin oral tissue shows the transcriptional hallmarks of an iron-replete state while in the kidney it is in the expected iron starved state. Interestingly,C. albicansexpresses genes associated with a low zinc environment in both niches. Consistent with these expression data, deletion of two transcription factors that activate iron uptake genes (SEF1,HAP5) have no effect on virulence in a mouse model of oral candidiasis. During microbial infection, the host sequesters iron and other metal nutrients to suppress growth of the pathogen in a process called nutritional immunity. Our results indicate thatC. albicansis subject to iron and zinc nutritional immunity during disseminated infection but is exempted from iron nutritional immunity during oral infection.
Family Vulnerability Scale: evidence of content and internal structure validity
Authors: de Souza Evelyn Lima , Eshriqui Ilana , Rebustini Flávio , Masuda Eliana Tiemi , de Paiva Neto Francisco Timbó , Lima Ricardo Macedo , Bonfim Daiana
Published: January 2023

Introduction

territory view based on families’ vulnerability strata allows identifying different health needs that, in their turn, can guide healthcare at primary care scope. Although there are instruments aimed at measuring family vulnerability, they still need robust validity evidences; therefore, they represent a limitation for usability in a country showing multiple socioeconomic and cultural realities, such as Brazil. The present study introduces the development and search for evidences about the validity of the Family Vulnerability Scale for Brazil, known as EVFAM-BR.

Methods

items were generated through exploratory qualitative study carried out with 123 professionals. Collected data subsidized the generation of 92 initial items that were subjected to a panel of multi-regional and multi-disciplinary judges (n = 73) to calculate the Content Validity Ratio (CVR) – this process resulted in a version of the scale comprising 38 items. Subsequently, it was applied to 1,255 individuals to find evidences about the internal-structure validity by using the Exploratory Factor Analysis (EFA). Dimensionality was assessed through Robust Parallel Analysis and the model was tested through cross-validation to find EVFAM-BR’s final version.

Results

the final version comprised 14 items distributed into four domains, with explained variance of 79.02%. All indicators were within adequate and satisfactory limits, without any cross-loading and Heywood Case issues. Reliability indices also reached adequate levels (α= 0.71;ω= 0.70; glb = 0.83 and ORION ranging from 0.80 to 0.93, between domains). Instrument’s score was subjected to normalization, and it pointed towards three vulnerability strata (0 to 4 – Low; 5 to 6 – Moderate; 7 to 14 - High).

Conclusion

the scale showed satisfactory validity evidences, which were consistent, reliable ad robust; it led to a synthesized instrument capable of accurately measuring and differentiating family vulnerability in the primary care territory, in Brazil.
Flat clathrin lattices are linked to metastatic potential in colorectal cancer
Authors: Cresens Charlotte , Solis-Fernandez Guillermo , Tiwari Astha , Nuyts Rik , Hofkens Johan , Barderas Rodrigo , Rocha Susana
Published: January 2023

Clathrin assembles at the cells’ plasma membrane in a multitude of clathrin-coated structures (CCSs). Among these are flat clathrin lattices (FCLs), alternative clathrin structures that have been found in specific cell types, including cancer cells. Here we show that these structures are also present in different colorectal cancer (CRC) cell lines, and that they are extremely stable with lifetimes longer than 8 hours. By combining cell models representative of CRC metastasis with advanced fluorescence imaging and analysis, we discovered that the metastatic potential of CRC is associated with an aberrant membranous clathrin distribution, resulting in a higher prevalence of FCLs in cells with a higher metastatic potential. These findings suggest that clathrin organization might play an important yet unexplored role in cancer metastasis.
The autism-associated loss of δ-catenin functions disrupts social behaviors
Authors: Mendez-Vazquez Hadassah , Roach Regan L. , Nip Kaila , Sathler Matheus F. , Garver Tyler , Danzman Rosaline A. , Moseley Madeleine C. , Roberts Jessica P. , Koch Olivia N. , Steger Ava A. , Lee Rahmi , Arikkath Jyothi , Kim Seonil
Published: January 2023

δ-catenin is expressed in excitatory synapses and functions as an anchor for the glutamatergic AMPA receptor (AMPAR) GluA2 subunit in the postsynaptic density. The glycine 34 to serine (G34S) mutation in theδ-cateningene is found in autism spectrum disorder (ASD) patients and induces loss of δ-catenin functions at excitatory synapses, which is presumed to underlie ASD pathogenesis in humans. However, how the G34S mutation causes loss of δ-catenin functions to induce ASD remains unclear. Here, using neuroblastoma cells, we discover that the G34S mutation generates an additional phosphorylation site for glycogen synthase kinase 3β (GSK3β). This promotes δ-catenin degradation and causes the reduction of δ-catenin levels, which likely contributes to the loss of δ-catenin functions. Synaptic δ-catenin and GluA2 levels in the cortex are significantly decreased in mice harboring the δ-catenin G34S mutation. The G34S mutation increases glutamatergic activity in cortical excitatory neurons while it is decreased in inhibitory interneurons, indicating changes in cellular excitation and inhibition. δ-catenin G34S mutant mice also exhibit social dysfunction, a common feature of ASD. Most importantly, inhibition of GSK3β activity reverses the G34S-induced loss of δ-catenin function effects in cells and mice. Finally, using δ-catenin knockout mice, we confirm that δ-catenin is required for GSK3β inhibition-induced restoration of normal social behaviors in δ-catenin G34S mutant animals. Taken together, we reveal that the loss of δ-catenin functions arising from the ASD-associated G34S mutation induces social dysfunction via alterations in glutamatergic activity and that GSK3β inhibition can reverse δ-catenin G34S-induced synaptic and behavioral deficits.

Significance Statement

δ-catenin is important for the localization and function of glutamatergic AMPA receptors at synapses in many brain regions. The glycine 34 to serine (G34S) mutation in theδ-cateningene is found in autism patients and results in the loss of δ-catenin functions. δ-catenin expression is also closely linked to other autism-risk genes involved in synaptic structure and function, further implying that it is important for the autism pathophysiology. Importantly, social dysfunction is a key characteristic of autism. Nonetheless, the links between δ-catenin functions and social behaviors are largely unknown. The significance of the current research is thus predicated on filling this gap by discovering the molecular, cellular, and synaptic underpinnings of the role of δ-catenin in social behaviors.
FOXM1 acts sexually dimorphically to regulate functional β-cell mass
Authors: Peng Guihong , Mosleh Elham , Yuhas Andrew , Katada Kay , Cherry Christopher , Golson Maria L.
Published: January 2023

The transcription factor FOXM1 regulates β-cell proliferation and insulin secretion. Our previous work demonstrates that expressing an activated form of FOXM1 (FOXM1*) in β cells increases β-cell proliferation and mass in aged male mice. Additionally, FOXM1* enhances β-cell function even in young mice, in which no β-cell mass elevation occurs. Here, we demonstrate that FOXM1 acts in a sexually dimorphic manner in the β cell. Expression of FOXM1* in female mouse β cells does not affect β-cell proliferation or glucose tolerance. Transduction of male but not female human islets with FOXM1* enhances insulin secretion in response to elevated glucose. Estrogen contributes to diabetes susceptibility differences between males and females, and the estrogen receptor (ER)α is the primary mediator of β-cell estrogen signaling. We show that FOXM1* can rescue impaired glucose tolerance in female mice with a pancreas-wide ERα deletion. Further, FOXM1 and ERα binding sites overlap with each other and with other β-cell-enriched transcription factors, including ISL1, PAX6, MAF, and GATA. These data indicate that FOMX1 and ERα cooperate to regulate β-cell function and suggest a general mechanism contributing to the lower incidence of diabetes observed in women.
Structure-based discovery of inhibitors of the SARS-CoV-2 Nsp14N7-methyltransferase
Authors: Singh Isha , Li Fengling , Fink Elissa , Chau Irene , Li Alice , Rodriguez-Hernández Annía , Glenn Isabella , Zapatero-Belinchón Francisco J. , Rodriguez Mario , Devkota Kanchan , Deng Zhijie , White Kris , Wan Xiaobo , Tolmachova Nataliya A. , Moroz Yurii S. , Kaniskan H. Ümit , Ott Melanie , Gastía-Sastre Adolfo , Jin Jian , Fujimori Danica Galonić , Irwin John J. , Vedadi Masoud , Shoichet Brian K.
Published: January 2023

An under-explored target for SARS-CoV-2 is non-structural protein 14 (Nsp14), a crucial enzyme for viral replication that catalyzes the methylation ofN7-guanosine of the viral RNA at 5′-end; this enables the virus to evade the host immune response by mimicking the eukaryotic post-transcriptional modification mechanism. We sought new inhibitors of the S-adenosyl methionine (SAM)-dependent methyltransferase (MTase) activity of Nsp14 with three large library docking strategies. First, up to 1.1 billion make-on-demand (“tangible”) lead-like molecules were docked against the enzyme’s SAM site, seeking reversible inhibitors. On de novo synthesis and testing, three inhibitors emerged with IC50values ranging from 6 to 43 μM, each with novel chemotypes. Structure-guided optimization andin vitrocharacterization supported their non-covalent mechanism. In a second strategy, docking a library of 16 million tangible fragments revealed nine new inhibitors with IC50values ranging from 12 to 341 μM and ligand efficiencies from 0.29 to 0.42. In a third strategy, a newly created library of 25 million tangible, virtual electrophiles were docked to covalently modify Cys387 in the SAM binding site. Seven inhibitors emerged with IC50values ranging from 3.2 to 39 μM, the most potent being a reversible aldehyde. Initial optimization of a second series yielded a 7 μM acrylamide inhibitor. Three inhibitors characteristic of the new series were tested for selectivity against 30 human protein and RNA MTases, with one showing partial selectivity and one showing high selectivity. Overall, 32 inhibitors encompassing eleven chemotypes had IC50values <50 μM and 5 inhibitors in four chemotypes had IC50values <10 μM. These molecules are among the first non-SAM-like inhibitors of Nsp14, providing multiple starting points for optimizing towards antiviral activity.
A simple image processing pipeline to sharpen topology maps in multi-wavelength interference microscopy
Authors: Tinning Peter W. , Schniete Jana K. , Scrimgeour Ross , Kölln Lisa S. , Rooney Liam M. , Bushell Trevor J. , McConnell Gail
Published: January 2023

Multi-wavelength standing wave (SW) microscopy and interference reflection microscopy (IRM) are powerful techniques that use optical interference to study topographical structure. However, the use of more than two wavelengths to image the complex cell surface results in complicated topographical maps and it can be difficult to resolve the three-dimensional contours. We present a simple image processing method to reduce the thickness and spacing of antinodal fringes in multi-wavelength interference microscopy by up to a factor of two to produce clearer and more precise topographical maps of cellular structures. We first demonstrate this improvement using model non-biological specimens, and we subsequently demonstrate the benefit of our method for reducing the ambiguity of surface topography and revealing obscured features in live and fixed cell specimens.
Incorporating discovery and replication GWAS into summary data Mendelian randomization studies: A review of current methods and a simple, general and powerful alternative
Authors: Mounier Ninon , Robertson David S. , Kutalik Zoltán , Dudbridge Frank , Bowden Jack
Published: January 2023

Mendelian Randomization (MR) is a popular method for using genetics to estimate the causal effect of a modifiable exposure on a health outcome. Single Nucleotide Polymorphisms (SNPs) are typically selected for inclusion if they pass a genome-wide significance threshold in order to guarantee that they are strong genetic instruments, but this also induces Winner’s curse, as SNP-exposure associations tend to be overestimated. In this paper, we consider how to combine SNP-exposure data from discovery and replication samples using two-sample and three-sample approaches to best account for Winner’s curse, weak instrument bias, and pleiotropy within a summary data MR framework, using only GWAS summary statistics. After reviewing several existing methods, that often correct for Winner’s curse at the individual SNP level, we propose a simple alternative based on the technique of regression calibration that enacts a global correction to the causal effect estimate directly. This approach does not only correct for Winner’s curse, but also simultaneously accounts for weak instruments bias. Regression calibration can be used with a wide range of existing MR methods, including pleiotropy-robust methods such as median-based and mode-based estimators. Extensive simulations and real data examples are used to illustrate the utility of the new approach. Software is provided for users to implement the method in practice.

Author Summary

Mendelian randomization is a method to explore causation in health research which exploits the random inheritance of genes from parents to offspring as a ‘natural experiment’. It attempts to quantify the effect of intervening and modifying a health exposure, such as a person’s body mass, on a downstream outcome such as blood pressure. Causal estimates obtained using this method can be strongly influenced by the set of genes used, or more specifically, the rationale used to select them. For example, selecting only genes that are strongly associated with the health exposure can induce bias due to the ‘Winner’s curse’. Unfortunately, using genes with a small association can lead to so-called ‘weak instrument’ bias leading to a no-win paradox. In this paper, we present a novel approach based on the technique of regression calibration to de-bias causal estimates in an MR study. Our approach relies on the use of two independent samples for the exposure (discovery and replication) to estimate the amount of bias that is expected for a specific set of genes, so that causal estimates can be re-calibrated accordingly. We use extensive simulations and applied examples to compare our approach to current methods and provide software for researchers to implement our approach in future studies.
iTie controls intestinal tolerance through suppressing the NLRP6 inflammasome
Authors: Qin Xiwen , Zhu Fangrui , Li Weitao , Wang Shuo , Xia Pengyan
Published: January 2023

Intestines are full of commensal bacteria that possess numerous pathogen-associated molecular patterns. How the intestinal epithelial cells are tolerant to these stimuli under normal conditions is still elusive. Here we show that iTie is expressed in small intestinal enterocytes and its deficiency leads to body weight loss in mice, accompanied by length reduce of small intestines and intestinal villi. The activation of NLRP6 inflammasome is exacerbated uponiTiedeletion. iTie has a higher binding affinity for NLRP6 than NLRP6’s physiological ligand LTA does.iTiedeficiency gives rise to uncontrolled GSDMD activation and pyroptosis of small intestinal enterocytes. Inhibition of GSDMD permeabilization on cell membrane ameliorates the damage caused byiTiedeficiency. iTe’s expression is diminished in small intestines of patients with Crohn’s disease. Our results uncover a self-control system for mouse intestine to tolerate commensal bacteria which might shed new light on the treatment of bowel diseases.
Cryo-EM structure of the endothelin-1-ETB-Gicomplex
Authors: Sano Fumiya K. , Akasaka Hiroaki , Shihoya Wataru , Nureki Osamu
Published: January 2023

The endothelin ETBreceptor is a promiscuous G-protein coupled receptor, activated by vasoactive peptide endothelins. ETBsignaling induces reactive astrocytes in the brain and vasorelaxation in vascular smooth muscle, and thus ETBagonists are expected to be utilized for neuroprotection and improved anti-tumor drug delivery. Here, we report a cryo-electron microscopy structure of the endothelin-1-ETB-Gicomplex at 2.8-Å resolution, with complex assembly stabilized by a newly established method. Comparisons with the inactive ETBreceptor structures revealed how endothelin-1 activates the ETBreceptor. The NPxxY motif, which is essential for G-protein activation, is not conserved in ETB, resulting in a unique structural change upon G-protein activation. As Compared with other GPCR-G-protein complexes, ETBbinds Giat the shallowest position, thus expanding the diversity of G-protein binding. This structural information will facilitate the elucidation of G-protein activation and the rational design of ETB-agonists.
The dietary polyphenol metabolite protocatechuic acid produces acute and sustained effects on hippocampal electrical activity
Authors: Montero-Atalaya Marta , Muñoz-Arnaiz Ricardo , Makarova Julia , Bartolomé Begoña , Moreno-Arribas María-Victoria , Herreras Oscar
Published: January 2023

Dietary polyphenols and in particular bioavailable metabolites resulting from gut microbiota transformations appear to have beneficial effects in situations of impaired cognition, combatting memory deficits in acute pathological models of neurodegeneration. Modifications to blood flow may underlie the effects of these molecules and although some such metabolites cross the blood-brain barrier, their targets and electrophysiological effects remain unknown. Hence, we explored the systemic and direct effects of protochatechuic acid (PCA) on electrical activity in the hippocampus and cortex of anesthetized female rats, recording evoked and spontaneous high-density field potentials (FPs) to mathematically derive pathway-specific FP generators. We found transient and sustained effects of PCA on evoked activity in the CA1 field, including paradoxical actions on excitatory transmission that depend on the route of administration. Systemic delivery of PCA altered the ongoing activity of some FP generators, albeit with marked inter-animal variation. Interestingly, PCA induced the detachment of infraslow cortico-hippocampal activities over a scale of minutes. These results point to direct actions of polyphenols on cell and network electrical activity, some of which reflect non-specific actions. Thus, dietary-derived polyphenols appear to fulfill neuromodulatory roles, encouraging the search for additional targets to better guide their use in preventing brain pathologies.
Multiparametric senescent cell phenotyping reveals CD24 osteolineage cells as targets of senolytic therapy in the aged murine skeleton
Authors: Doolittle Madison L. , Saul Dominik , Kaur Japneet , Rowsey Jennifer L. , Vos Stephanie J. , Pavelko Kevin D. , Farr Joshua N. , Monroe David G. , Khosla Sundeep
Published: January 2023

Senescence drives organismal aging, yet the deep characterization of senescent cellsin vivoremains incomplete. Here, we applied mass cytometry by time-of-flight (CyTOF) using carefully validated antibodies to analyze senescent cells at single-cell resolution. We used multiple criteria to identify senescent mesenchymal cells that were growth arrested and resistant to apoptosis (p16+/Ki67-/BCL-2+; “p16KB” cells). These cells were highly enriched for senescence-associated secretory phenotype (SASP) and DNA damage markers and were strongly associated with age. p16KB cell percentages were also increased in CD24+ osteolineage cells, which exhibited an inflammatory SASP in aged mice and were robustly cleared by both genetic and pharmacologic senolytic therapies. Following isolation, CD24+ skeletal cells exhibited growth arrest, SA-βgal positivity, and impaired osteogenesisin vitro. These studies thus provide a new approach using multiplexed protein profiling by CyTOF to define senescent mesenchymal cellsin vivoand identify a highly inflammatory, senescent CD24+ osteolineage population cleared by senolytics.
Goal-seeking compresses neural codes for space in the human hippocampus and orbitofrontal cortex
Authors: Muhle-Karbe PS , Sheahan H , Pezzulo G , Spiers H , Chien S , Schuck NW , Summerfield C
Published: January 2023

Humans can navigate flexibly to meet their goals. Here, we asked how the neural representation of allocentric space is distorted by goal-directed behaviour. Participants navigated an agent to two successive goal locations in a grid world environment comprising four interlinked rooms, with a contextual cue indicating the conditional dependence of one goal location on another. Examining the neural geometry by which room and context were encoded in fMRI signals, we found that map-like representations of the environment emerged in both hippocampus and neocortex. Cognitive maps in hippocampus and orbitofrontal cortices were compressed so that locations cued as goals were coded together in neural state space, and these distortions predicted successful learning. This effect was captured by a computational model in which current and prospective locations are jointly encoded in a place code, providing a theory of how goals warp the neural representation of space in macroscopic neural signals.
End Binding protein 1 promotes specific motor-cargo association in the cell body prior to axonal delivery of Dense Core Vesicles
Authors: Park Junhyun , Miller Kenneth G. , De Camilli Pietro , Yogev Shaul
Published: January 2023

Axonal transport is key to neuronal function. Efficient transport requires specific motor-cargo association in the soma, yet the mechanisms regulating this early step remain poorly understood. We found that EBP-1, theC. elegansortholog of the canonical microtubule end binding protein EB1, promotes the specific association between kinesin-3/KIF1A/UNC-104 and Dense Core Vesicles (DCVs) prior to their axonal delivery. Using single-neuron,in vivolabelling of endogenous cargo and EBs, we observed reduced axonal abundance and reduced secretion of DCV cargo, but not other KIF1A/UNC-104 cargo, inebp-1mutants. This reduction could be traced back to fewer exit events from the cell body, where EBP-1 colocalized with the DCV sorting machinery at the trans Golgi, suggesting that this is the site of EBP-1 function. In addition to its microtubule binding CH domain, mammalian EB1 interacted with mammalian KIF1A in an EBH domain dependent manner, and expression of mammalian EB1 or the EBH domain was sufficient to rescue DCV transport inebp-1mutants. Our results suggest a model in which kinesin-3 binding and microtubule binding by EBP-1 cooperate to transiently enrich the motor near sites of DCV biogenesis to promote motor-cargo association. In support of this model, tethering either EBP-1 or a kinesin-3 KIF1A/UNC-104 interacting domain from an unrelated protein to the Golgi restored the axonal abundance of DCV proteins inebp-1mutants. These results uncover an unexpected role for a microtubule associated protein and provide insight into how specific kinesin-3 cargo are delivered to the axon.
A highly conserved neuronal microexon in DAAM1 controls actin dynamics, RHOA/ROCK signaling, and memory formation
Authors: Poliński Patryk , Miret Cuesta Marta , Zamora-Moratalla Alfonsa , Mantica Federica , Cantero-Recasens Gerard , Normanno Davide , Iñiguez Luis P. , Morenilla-Palao Cruz , Ordoño Patricia , Bonnal Sophie , Ellis Jonathan D , Gómez Riera Raúl , De Lagrán Cabredo María Martínez , Fernández-Blanco Álvaro , Rodríguez-Marin Cristina , Permanyer Jon , Fölsz Orsolya , Dominguez-Sala Eduardo , Sierra Cesar , Legutko Diana , Wojnacki José , Musoles Lleo Juan Luis , Muñoz López Francisco José , Blencowe Benjamin J , Herrera Eloisa , Dierssen Mara , Irimia Manuel
Published: January 2023

Actin cytoskeleton dynamics is critical for nervous system development and function, yet the role of alternative splicing in controlling these processes is poorly understood. A highly conserved subset of neuronal-specific microexons coordinates fundamental aspects of nervous system biology. A subset of these exons is enriched in genes involved in actin cytoskeleton, yet their functions are unknown. Here, we focus on a microexon in DAAM1, a member of the formin-homology-2 (FH2) domain class of proteins, which have diverse functions associated with the reorganization of the actin cytoskeleton. Remarkably, splicing of the microexon extends the linker region of the DAAM1 FH2 domain and leads to qualitative and quantitative changes in actin polymerization. Deletion of the microexon results in neuritogenesis defects and increased calcium influx in differentiated neurons. Moreover, mice harboring the deletion exhibit postsynaptic defects, reduced number of immature dendritic spines, impaired long-term potentiation, and deficits in memory formation. These deficits are associated with increased RHOA/ROCK signaling, pivotal in controlling actin-cytoskeleton dynamics, and were rescued by treatment with a ROCK inhibitor. We thus demonstrate that a conserved neuronal microexon in DAAM1 is critical for controlling actin dynamics through the RHOA/ROCK signaling pathway and is necessary for normal cognitive functioning.
Imputation of label-free quantitative mass spectrometry-based proteomics data using self-supervised deep learning
Authors: Webel Henry , Niu Lili , Nielsen Annelaura Bach , Locard-Paulet Marie , Mann Matthias , Jensen Lars Juhl , Rasmussen Simon
Published: January 2023

Imputation techniques provide means to replace missing measurements with a value and are used in almost all downstream analysis of mass spectrometry (MS) based proteomics data using label-free quantification (LFQ). Here we demonstrate how collaborative filtering, denoising autoencoders, and variational autoencoders can impute missing values in the context of LFQ at different levels. We applied our method, proteomics imputation modeling mass spectrometry (PIMMS), to an alcohol-related liver disease (ALD) cohort with blood plasma proteomics data available for 358 individuals. Removing 20 percent of the intensities we were able to recover 15 out of 17 significant abundant protein groups using PIMMS-VAE imputations. When analyzing the full dataset we identified 30 additional proteins (+13.2%) that were significantly differentially abundant across disease stages compared to no imputation and found that some of these were predictive of ALD progression in machine learning models. We, therefore, suggest the use of deep learning approaches for imputing missing values in MS-based proteomics on larger datasets and provide workflows for these.
CoVar: A generalizable machine learning approach to identify the coordinated regulators driving variational gene expression
Authors: Roy Satyaki , Sheikh Shehzad Z. , Furey Terrence S.
Published: January 2023

Network inference is used to model transcriptional, signaling, and metabolic interactions among genes, proteins, and metabolites that identify biological pathways influencing disease pathogenesis. Advances in machine learning (ML)-based inference models exhibit the predictive capabilities of capturing latent patterns in genomic data. Such models are emerging as an alternative to the statistical models identifying causative factors driving complex diseases. We present CoVar, an inference framework that builds upon the properties of existing inference models, to find the central genes driving perturbed gene expression across biological states. We leverage ML-based network inference to find networks that capture the strength of regulatory interactions. Our model first pinpoints a subset of genes, termed variational, whose expression variabilities typify the differences in network connectivity between the control and perturbed data. Variational genes, by being differentially expressed themselves or possessing differentially expressed neighbor genes, capture gene expression variability. CoVar then creates subnetworks comprising variational genes and their strongly connected neighbor genes and identifies core genes central to these subnetworks that influence the bulk of the variational activity. Through the analysis of yeast expression data perturbed by the deletion of the mitochondrial genome, we show that CoVar identifies key genes not found through independent differential expression analysis.
The phenotype and genotype of fermentative microbes
Authors: Hackmann Timothy J.
Published: January 2023

Fermentation is a major type of metabolism carried out by many organisms. The study of this metabolism cuts across many fields, including cell biology, animal and human health, and biofuel production. Despite this broad importance, there has been no systematic study of fermentation across many organisms. Here we explore the phenotype and genotype of fermentative prokaryotes in order to gain insight into this metabolism. We assembled a dataset containing phenotypic records of 8,350 organisms (type strains) plus 4,355 genomes and 13.6 million genes. Fermentation was widespread, being found in 30% of all organisms and across the tree of life. Fermentative organisms were more likely than non-fermentative ones to have certain phenotypic traits. Some traits (such as oxygen insensitivity) were expected, but others (such as long cells) were surprising. Fermentative organisms also had a distinct genotype, with 9,450 gene functions and 337 metabolic pathways being more common in them. In a related analysis, we identified end products (metabolites) for 1,455 organisms fermenting 100 substrates. We found 55 products were formed in nearly 300 combinations, showing fermentation is more complex than previously realized. Additionally, we built metabolic models for 406 organisms and predicted which products they form. These models did not predict all products accurately, revealing gaps in our knowledge of metabolic pathways. Our study paints a full picture of fermentation while showing there is still much to learn about this type of metabolism. The microbiology community can explore resources in this work with an interactive tool (https://github.com/thackmann/FermentationExplorer).
Distinct subpopulations of D1 medium spiny neurons exhibit unique transcriptional responsiveness to cocaine
Authors: Phillips Robert A. , Tuscher Jennifer J. , Fitzgerald N. Dalton , Wan Ethan , Zipperly Morgan E. , Duke Corey G. , Ianov Lara , Day Jeremy J.
Published: January 2023

Drugs of abuse increase extracellular concentrations of dopamine in the nucleus accumbens (NAc), resulting in transcriptional alterations that drive long-lasting cellular and behavioral adaptations. While decades of research have focused on the transcriptional mechanisms by which drugs of abuse influence neuronal physiology and function, few studies have comprehensively defined NAc cell type heterogeneity in transcriptional responses to drugs of abuse. Here, we used single nucleus RNA-seq (snRNA-seq) to characterize the transcriptome of over 39,000 NAc cells from male and female adult Sprague-Dawley rats following acute or repeated cocaine experience. This dataset identified 16 transcriptionally distinct cell populations, including two populations of medium spiny neurons (MSNs) that express the Drd1 dopamine receptor (D1-MSNs). Critically, while both populations expressed classic marker genes of D1-MSNs, only one population exhibited a robust transcriptional response to cocaine. Validation of population-selective transcripts using RNA in situ hybridization revealed distinct spatial compartmentalization of these D1-MSN populations within the NAc. Finally, analysis of published NAc snRNA-seq datasets from non-human primates and humans demonstrated conservation of MSN subtypes across rat and higher order mammals, and further highlighted cell type-specific transcriptional differences across the NAc and broader striatum. These results highlight the utility in using snRNA-seq to characterize both cell type heterogeneity and cell type-specific responses to cocaine and provides a useful resource for cross-species comparisons of NAc cell composition.
Mathematical model of nuclear speckle morphology
Authors: Wakao Shingo , Saitoh Noriko , Awazu Akinori
Published: January 2023

Nuclear speckles are nuclear bodies consisting of populations of small and irregularly shaped droplet-like molecular condensates that contain various splicing factors. Recent experiments have shown the following morphological features of nuclear speckles: (I) Each molecular condensate contains SON and SRRM2 proteins, andMALAT1 non-coding RNA surrounds these condensates; (II) In the normal interphase of the cell cycle, these condensates are broadly distributed throughout the nucleus in multicellular organisms. In contrast, the fusion of condensates leads to the formation of strongly condensed spherical droplets when cell transcription is suppressed; (III) SON is dispersed spatially inMALAT1knocked-down cells, whereasMALAT1is dispersed in SON knocked-down cells by the collapse of nuclear speckles. However, the detailed interactions among molecules that reveal the mechanisms of this rich variety of morphologies remain unknown. In this study, a coarse-grained molecular dynamics model of the nuclear speckle was developed considering the dynamics of SON, SRRM2 or SRSF2,MALAT1, and pre-mRNA as representative components of condensates. The simulations reproduced the abovementioned morphological changes, by which the interaction strength among the representative components of the condensates was predicted.