bluuebunny/crossref_metadata_2025 huggingface dataset:
Assessing Protein Surface-Based Scoring for Interpreting Genomic Variants
Authors: Dsouza Nikita R. , Haque Neshatul , Tripathi Swarnendu , Zimmermann Michael T.
Published: December 2023

Clinical genomics sequencing is rapidly expanding the number of variants that need to be functionally elucidated. Interpreting genetic variants (i.e., mutations) usually begins by identifying how they affect protein-coding sequences. Still, the 3D protein molecule is rarely considered for large-scale variant analysis, nor for their effects on how proteins interact with each other and their environment. We propose a standardized approach to scoring protein surface property changes as a new dimension for functionally and mechanistically interpreting genomic variants. Further, it directs hypothesis generation for functional genomics research to learn more about the encoded protein’s function. We developed a novel method leveraging 3D structures and time-dependent simulations to score and statistically evaluate protein surface property changes. We evaluated positive controls composed of 8 thermophilic versus mesophilic orthologs and variants that experimentally change protein solubility, which all showed large and statistically significant differences in charge distribution (p<0.01). We scored static 3D structures and dynamic ensembles for 43 independent variants (23 pathogenic and 20 uninterpreted) across four proteins. Focusing on the potassium ion channel, KCNK9, average local surface potential shifts were 0.41 kBT/ec with an average p-value of 1×10−2. In contrast, dynamic ensemble shifts averaged 1.15 kBT/ec and an average p-value of 1×10−5, enabling the identification of changes far from mutated sites. This study demonstrates that an objective assessment of how mutations affect electrostatic distributions of protein surfaces can aid in interpreting genomic variants discovered through clinical genomics sequencing.
A computational procedure for predicting excipient effects on protein-protein affinities
Authors: Dignon Gregory L. , Dill Ken A.
Published: December 2023

Protein-protein interactions lie at the center of much biology and are a challenge in formulating biological drugs such as antibodies. A key to mitigating protein association is to use small molecule additives, i.e. excipients that can weaken protein-protein interactions. Here, we develop a computationally efficient model for predicting the viscosity-reducing effect of different excipient molecules by combining atomic-resolution MD simulations, binding polynomials and a thermodynamic perturbation theory. In a proof of principle, this method successfully rank orders four types of excipients known to reduce the viscosity of solutions of a particular monoclonal antibody. This approach appears useful for predicting effects of excipients on protein association and phase separation, as well as the effects of buffers on protein solutions.

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Brood care in shell-dwelling cichlids is timed by independent maternal and larval clocks
Authors: Parker Ash V , Stemmer Manuel , Grätsch Swantje , Dorigo Alessandro , Ramirez Oriolson Rodriguez , Adel Abdelrahman , Jordan Alex , Baier Herwig
Published: December 2023

Successful brood care requires a finely tuned interplay between caregiving parent and offspring. Cichlids are a diverse group of teleost fish known for their extensive parental care of altricial young. FemaleLamprologus ocellatuslay their eggs in the protective confines of an abandoned snail shell, where they raise them to free-swimming larvae. Here we investigated whether the transition to larval independence is driven by the behavior of the mother, the offspring, or an interaction between the two. Using 3D printed shells with a window, we were able to observe the behavior of both mother and offspring inside the nest using automated object detection and tracking. We found that, once hatched, the larvae remain in the deep chambers of the shell, and at 9 dpf, actively emerge from the shell during the day. An inversion of larval phototactic behavior from dark- to light-seeking leads up to this emergence. Removal of the mother causes premature emergence of the larvae by a few days. The natural schedule (emergence at 9 dpf) is restored if the larvae are supplied with fresh water in the mother’s absence. In cross-fostering experiments, we found that swapping out the mother’s own clutch for older larvae forces a delay in emergence, as the foster mother prevents the larvae from leaving at 9 dpf. We conclude that larval and maternal behavior are each controlled by independent internal clocks, which, although normally synchronized, can be experimentally manipulated and brought into conflict. This study has thus revealed an innate sequence of behavioral adaptations that orchestrate brood care in shell-dwelling cichlids.
Visualizing PIEZO1 Localization and Activity in hiPSC-Derived Single Cells and Organoids with HaloTag Technology
Authors: Bertaccini Gabriella A. , Casanellas Ignasi , Evans Elizabeth L. , Nourse Jamison L. , Dickinson George D. , Liu Gaoxiang , Seal Sayan , Ly Alan T. , Holt Jesse R. , Wijerathne Tharaka D. , Yan Shijun , Hui Elliot E. , Lacroix Jerome J. , Panicker Mitradas M. , Upadhyayula Srigokul , Parker Ian , Pathak Medha M.
Published: December 2023

PIEZO1 is critical to numerous physiological processes, transducing diverse mechanical stimuli into electrical and chemical signals. Recent studies underscore the importance of visualizing endogenous PIEZO1 activity and localization to understand its functional roles. To enable physiologically and clinically relevant studies on human PIEZO1, we genetically engineered human induced pluripotent stem cells (hiPSCs) to express a HaloTag fused to endogenous PIEZO1. Combined with advanced imaging, our chemogenetic platform allows precise visualization of PIEZO1 localization dynamics in various cell types. Furthermore, the PIEZO1-HaloTag hiPSC technology facilitates the non-invasive monitoring of channel activity across diverse cell types using Ca2+-sensitive HaloTag ligands, achieving temporal resolution approaching that of patch clamp electrophysiology. Finally, we used lightsheet imaging of hiPSC-derived neural organoids to achieve molecular scale imaging of PIEZO1 in three-dimensional tissue organoids. Our advances offer a novel platform for studying PIEZO1 mechanotransduction in human cells and tissues, with potential for elucidating disease mechanisms and targeted therapeutic development.
Combining transcutaneous spinal stimulation and functional electrical stimulation increases force generated by lower limbs: When more is more
Authors: Steele Alexander G. , Vette Albert H. , Martin Catherine , Masani Kei , Sayenko Dimitry G.
Published: December 2023

Background

Transcutaneous Spinal Stimulation (TSS) has been shown to promote activation of the lower limb and trunk muscles and is being actively explored for improving the motor outcomes of people with neurological conditions. However, individual responses to TSS vary, and often the muscle responses are insufficient to produce enough force for self-supported standing. Functional electrical stimulation (FES) can activate individual muscles and assist in closing this functional gap, but it introduces questions regarding timing between modalities.

Methods

To assess the effects of TSS and FES on force generation, ten neurologically intact participants underwent (1) TSS only, (2) FES only, and (3) TSS + FES. TSS was delivered using four electrodes placed at T10–T11 through the L1–L2 intervertebral spaces simultaneously, while FES was delivered to the skin over the right knee extensors and plantarflexors. For all conditions, TSS and FES were delivered using three 0.5 ms biphasic square-wave pulses at 15 Hz. During the TSS + FES condition, timing between the two modalities was adjusted in increments of ¼ time between pulses (16.5 ms).

Results

When TSS preceded FES, a larger force production was observed. We also determined several changes in muscle activation amplitude at different relative stimulus intervals, which help characterize our finding and indicate the facilitating and inhibitory effects of the modalities.

Conclusions

Utilizing a delay ranging from 15 to 30 ms between stimuli resulted in higher mean force generation in both the knee and ankle joints, regardless of the selected FES location (Average; knee: 112.0%, ankle: 103.1%).
LinkedOXTRVariants Are Associated with Social Behavior Differences in Bonobos (Pan paniscus)
Authors: Skiba Sara A. , Hansen Alek , McCall Ryan , Byers Azeeza , Waldron Sarah , Epping Amanda J. , Taglialatela Jared P. , Hudson Martin L.
Published: December 2023

Single-nucleotide polymorphisms (SNPs) in forkhead box protein P2 (FOXP2) and oxytocin receptor (OXTR) genes have been associated with linguistic and social development in humans, as well as to symptom severity in autism spectrum disorder (ASD). Studying biobehavioral mechanisms in the species most closely related to humans can provide insights into the origins of human communication, and the impact of genetic variation on complex behavioral phenotypes. Here, we aimed to determine if bonobos (Pan paniscus) exhibit individual variation inFOXP2andOXTRloci that have been associated with human social development and behavior. Although the ASD-related variants were reported in 13-41% of the human population, we did not find variation at these loci in our sample of 13 bonobos. However, we did identify a novel variant in bonoboFOXP2, as well as four novel variants in bonoboOXTRthat were 17-184 base pairs from the human ASD variants. We also found the same linked, homozygous allelic combination across the 4 novelOXTRSNPs (homozygous TGTC) in 6 of the 13 bonobos, indicating that this combination may be under positive selection. When comparing the combinedOXTRgenotypes, we found significant group differences in social behavior; bonobos with zero copies of the TGTC combination were less social than bonobos with one copy of the TGTC combination. Taken together, our findings suggest that theseOXTRvariants may influence individual-level social behavior in bonobos and support the notion that linked genetic variants are promising risk factors for social communication deficits in humans.
Human cerebrospinal fluid single exosomes in Parkinson’s and Alzheimer’s diseases
Authors: Yakabi Koya , Berson Eloise , Montine Kathleen S. , Bendall Sean C. , MacCoss Michael J. , Poston Kathleen L. , Montine Thomas J.
Published: December 2023

Exosomes are proposed to be important in the pathogenesis of prevalent neurodegenerative diseases. We report the first application of solid-state technology to perform multiplex analysis of single exosomes in human cerebrospinal fluid (CSF) obtained from the lumbar sac of people diagnosed with Alzheimer’s disease dementia (ADD, n=30) or Parkinson’s disease dementia (PDD, n=30), as well as age-matched health controls (HCN, n=30). Single events were captured with mouse monoclonal antibodies to one of three different tetraspanins (CD9, CD63, or CD81) or with mouse (M) IgG control, and then probed with fluorescently labeled antibodies to prion protein (PrP) or CD47 to mark neuronal or presynaptic origin, as well as ADD- and PDD-related proteins: amyloid beta (Aβ), tau, α-synuclein, and Apolipoprotein (Apo) E. Data were collected only from captured events that were within the size range of 50 to 200 nm. Exosomes were present at approximately 100 billion per mL human CSF and were similarly abundant for CD9+ and CD81+ events, but CD63+ were only 22% to 25% of CD9+ (P<0.0001) or CD81+ (P<0.0001) events. Approximately 24% of CSF exosomes were PrP+, while only 2% were CD47+. The vast majority of exosomes were surface ApoE+, and the number of PrP-ApoE+ (P<0.001) and PrP+ApoE+ (P<0.01) exosomes were significantly reduced in ADD vs. HCN for CD9+ events only. Aβ, tau, and α-synuclein were not detected on the exosome surface or in permeabilized cargo. These data provide new insights into single exosome molecular features and highlight reduction in the CSF concentration of ApoE+ exosomes in patients with ADD.
CompHEAR: A Customizable and Scalable Web-Enabled Auditory Performance Evaluation Platform for Cochlear Implant Sound Processing Research
Authors: Merrill Kris , Muller Leah , Beim Jordan A. , Hehrmann Phillipp , Swan Dean , Alfsmann Daniel , Spahr Tony , Litvak Leo , Oxenham Andrew J. , Tward Aaron D.
Published: December 2023

Objective

Cochlear implants (CIs) are auditory prostheses for individuals with severe to profound hearing loss, offering substantial but incomplete restoration of hearing function by stimulating the auditory nerve using electrodes. However, progress in CI performance and innovation has been constrained by the inability to rapidly test multiple sound processing strategies. Current research interfaces provided by major CI manufacturers have limitations in supporting a wide range of auditory experiments due to portability, programming difficulties, and the lack of direct comparison between sound processing algorithms. To address these limitations, we present the CompHEAR research platform, designed specifically for the Cochlear Implant Hackathon, enabling researchers to conduct diverse auditory experiments on a large scale.

Study Design

Quasi-experimental

Setting

Virtual

Methods

CompHEAR is an open-source, user-friendly platform which offers flexibility and ease of customization, allowing researchers to set up a broad set of auditory experiments. CompHEAR employs a vocoder to simulate novel sound coding strategies for CIs. It facilitates even distribution of listening tasks among participants and delivers real-time metrics for evaluation. The software architecture underlies the platform’s flexibility in experimental design and its wide range of applications in sound processing research.

Results

Performance testing of the CompHEAR platform ensured that it could support at least 10,000 concurrent users. The CompHEAR platform was successfully implemented during the COVID-19 pandemic and enabled global collaboration for the CI Hackathon (www.cihackathon.com).

Conclusion

The CompHEAR platform is a useful research tool that permits comparing diverse signal processing strategies across a variety of auditory tasks with crowdsourced judging. Its versatility, scalability, and ease of use can enable further research with the goal of promoting advancements in cochlear implant performance and improved patient outcomes.
Sunfish as zooplankton control agents to improve yields of wastewater-cultivated algae
Authors: Rakowski Chase J. , Leibold Mathew A. , Manning Schonna R.
Published: December 2023

Wastewater algal cultivation (WAC) in outdoor raceway ponds is a promising means of removing polluting nutrients from water while producing biomass feedstock for products like fertilizer or biofuel. However, algal cultivation in open ponds is often plagued by underproduction due to contamination by zooplankton. The use of biological controls, i.e., natural predators to control zooplankton, was evaluated as a potential low-cost solution in this study. Eighteen pilot-scale raceway ponds were inoculated with cultured algae and naturally-occurring zooplankton; bluegill sunfish (Lepomis macrochirus) were tested as biological control agents to suppress zooplankton. We installed cages in the ponds to protect the fish from the paddlewheels and control the area the fish could access. The cage either surrounded the paddlewheel – in which case the fish had access to the remaining area – or the cage was opposite the paddlewheel, in which case the fish had access to the smaller area inside the cage. We also implemented fishless controls. After 19 days, ponds with cages surrounding the paddlewheel had 93% more zooplankton, but the fish had reduced zooplankton by 89% on average across cage treatments. Cage placement did not affect algal biomass, but ponds with fish had 45% higher algal dry weight and 84% higher algal biovolume. However, no differences in nutrient levels were recorded among treatments after 19 days. We concluded hardy zooplanktivorous fish represent a promising means of boosting algal production in WAC, although further research is needed to determine whether the change in algal biomass could eventually translate to improved nutrient removal.
Decoding the Epigenetics and Chromatin Loop Dynamics of Androgen Receptor-Mediated Transcription
Authors: Altıntaş Umut Berkay , Seo Ji-Heui , Giambartolomei Claudia , Ozturan Dogancan , Fortunato Brad J. , Nelson Geoffrey M. , Goldman Seth Raphael , Adelman Karen , Hach Faraz , Freedman Matthew L. , Lack Nathan A.
Published: December 2023

Androgen receptor (AR)-mediated transcription plays a critical role in normal prostate development and prostate cancer growth. AR drives gene expression by binding to thousands of cis-regulatory elements (CRE) that loop to hundreds of target promoters. With multiple CREs interacting with a single promoter, it remains unclear how individual AR bound CREs contribute to gene expression. To characterize the involvement of these CREs, we investigated the AR-driven epigenetic and chromosomal chromatin looping changes. We collected a kinetic multiomic dataset comprised of steady-state mRNA, chromatin accessibility, transcription factor binding, histone modifications, chromatin looping, and nascent RNA. Using an integrated regulatory network, we found that AR binding induces sequential changes in the epigenetic features at CREs, independent of gene expression. Further, we showed that binding of AR does not result in a substantial rewiring of chromatin loops, but instead increases the contact frequency of pre-existing loops to target promoters. Our results show that gene expression strongly correlates to the changes in contact frequency. We then proposed and experimentally validated an unbalanced multi-enhancer model where the impact on gene expression of AR-bound enhancers is heterogeneous, and is proportional to their contact frequency with target gene promoters. Overall, these findings provide new insight into AR-mediated gene expression upon acute androgen simulation and develop a mechanistic framework to investigate nuclear receptor mediated perturbations.
Modulators of MAPK pathway activity during filamentous growth inSaccharomyces cerevisiae
Authors: Pujari Atindra N. , Cullen Paul J.
Published: December 2023

Mitogen-activated protein kinase (MAPK) pathways control the response to intrinsic and extrinsic stimuli. In the budding yeastSaccharomyces cerevisiae, cells undergo filamentous growth, which is regulated by the fMAPK pathway. To better understand the regulation of the fMAPK pathway, a genetic screen was performed to identify spontaneous mutants with elevated activity of an fMAPK-pathway dependent growth reporter (ste4 FUS1-HIS3). In total, 159 mutants were isolated and analyzed by secondary screens for invasive growth by the plate-washing assay, and filament formation by microscopy. Thirty-two mutants were selected for whole-genome sequencing, which identified new alleles in genes encoding known regulators of the fMAPK pathway. These included gain-of-function alleles inSTE11,which encodes the MAPKKK, as well as loss-of-function alleles inKSS1,which encodes the MAP kinase, andRGA1,which encodes a GTPase activating protein (GAP) forCDC42. New alleles in previously identified pathway modulators were also uncovered inALY1, AIM44, RCK2, IRA2, REG1and in genes that regulate protein folding (KAR2), glycosylation (MNN4), and turnover (BLM10). C-terminal truncations in the transcription factor Ste12p were also uncovered that resulted in elevated reporter activity, presumably identifying an inhibitory domain in the C-terminus of the protein. We also show that a wide variety of filamentous growth phenotypes result from mutations in different regulators of the response. The alleles identified here expand the connections surrounding MAPK pathway regulation and reveal new features of proteins that function in the signaling cascade.

ARTICLE SUMMARY

Signaling pathways control the response to stimuli. In yeast, a signaling (MAPK) pathway controls a fungal behavioral response called filamentous growth. A genetic screen was performed to identify spontaneous mutants that show hyperactivity of a MAPK pathway-dependent reporter. Select mutants were analyzed by whole-genome sequencing. New alleles in known regulatory proteins were identified. A potential inhibitory domain in the C-terminus of the transcription factor Ste12p was also uncovered. Our results indicate that filamentous growth is determined by the combinatorial effects of multiple positive and negative regulatory inputs.
TGM6, a helminth secretory product, mimics TGF-β binding to TβRII to antagonize TGF-β signaling in fibroblasts
Authors: White Stephen E. , Schwartze Tristin A. , Mukundan Ananya , Schoenherr Christina , Singh Shashi P. , van Dinther Maarten , Cunningham Kyle T. , White Madeleine P. J. , Campion Tiffany , Pritchard John , Hinck Cynthia S. , ten Dijke Peter , Inman Gareth , Maizels Rick M. , Hinck Andrew P.
Published: December 2023

The murine helminth parasiteHeligmosomoides polygyrusexpresses a family of proteins structurally related to TGF-β Mimic 1 (TGM1), a secreted five domain protein that activates the TGF-β pathway and converts naïve T lymphocytes to immunosuppressive Tregs. TGM1 signals through the TGF-β type I and type II receptors, TβRI and TβRII, with domains 1-2 and 3 binding TβRI and TβRII, respectively, and domains 4-5 binding CD44, a co-receptor abundant on T cells. TGM6 is a homologue of TGM1 that is co-expressed with TGM1, but lacks domains 1 and 2. Herein, we show that TGM6 binds TβRII through domain 3, but does not bind TβRI, or other type I or type II receptors of the TGF-β family. In TGF-β reporter assays in fibroblasts, TGM6, but not truncated TGM6 lacking domains 4 and 5, potently inhibits TGF-β- and TGM1-induced signaling, consistent with its ability to bind TβRII but not TβRI or other receptors of the TGF-β family. However, TGM6 does not bind CD44 and is unable to inhibit TGF-β and TGM1 signaling in T cells. To understand how TGM6 binds TβRII, the X-ray crystal structure of the TGM6 domain 3 bound to TβRII was determined at 1.4 Å. This showed that TGM6 domain 3 binds TβRII through an interface remarkably similar to the TGF-β:TβRII interface. These results suggest that TGM6 has adapted its domain structure and sequence to mimic TGF-β binding to TβRII and function as a potent TGF-β and TGM1 antagonist in fibroblasts. The coexpression of TGM6, along with the immunosuppressive TGMs that activate the TGF-β pathway, may prevent tissue damage caused by the parasite as it progresses through its life cycle from the intestinal lumen to submucosal tissues and back again.
Acclimation Capacity to Global Warming of Amphibians and Freshwater Fishes: Drivers, Patterns, and Data Limitations
Authors: Ruthsatz Katharina , Dahlke Flemming , Alter Katharina , Wohlrab Sylke , Eterovick Paula C. , Lyra Mariana L. , Gippner Sven , Cooke Steven J. , Peck Myron A.
Published: December 2023

Amphibians and fishes play a central role in shaping the structure and function of freshwater environments. These organisms have a limited capacity to disperse across different habitats and the thermal buffer offered by freshwater systems is small. Understanding determinants and patterns of their physiological sensitivity across life history is, therefore, imperative to predicting the impacts of climate change in freshwater systems. Based on a systematic literature review including 345 experiments with 998 estimates on 96 amphibian (Anura/Caudata) and 93 freshwater fish species (Teleostei), we conducted a quantitative synthesis to explore phylogenetic, ontogenetic, and biogeographic (thermal adaptation) patterns in upper thermal tolerance (CTmax) and thermal acclimation capacity (Acclimation Response Ratio, ARR) as well as the influence of the methodology used to assess these thermal traits using a conditional inference tree analysis. We found globally consistent patterns in CTmax and ARR, with phylogeny (taxa/order), experimental methodology, climatic origin, and life stage as significant determinants of thermal traits. The analysis demonstrated that CTmax does not primarily depend on the climatic origin but on experimental acclimation temperature and duration, and life stage. Higher acclimation temperatures and longer acclimation times led to higher CTmax values, whereby Anuran larvae revealed a higher CTmax than older life stages. The ARR of freshwater fishes was more than twice that of amphibians. Differences in ARR between life stages were not significant. In addition to phylogenetic differences, we found that ARR also depended on acclimation duration, ramping rate, and adaptation to local temperature variability. However, the amount of data on early life stages is too small, methodologically inconsistent, and phylogenetically unbalanced to identify potential life cycle bottlenecks in thermal traits. We therefore propose methods to improve the robustness and comparability of CTmax/ARR data across species and life stages, which is crucial for the conservation of freshwater biodiversity under climate change.
Electrostatic-driven Interactions Enhance Intratumoral Retention and Antitumor Efficacy of Immune Checkpoint Blockade Antibodies
Authors: Mohanty Rashmi P , Pan Yuting , Lewis Mae M , Soto Melissa , Maier Esther Y , Alzhrani Riyad F , Ghosh Debadyuti
Published: December 2023

Tumor extracellular matrix (ECM) forms a net negative charged network that interacts with and hinders the transport of molecules partly based on electrostatic interactions. The focus on drug delivery in solid tumors has traditionally been on developing neutral charge coatings to minimize interactions with the ECM for improved transport. In contrast to this prior work, we recently found a cationic peptide that interacted electrostatically with the negatively charged components of the ECM, resulting in enhanced uptake and retention of nanoparticles in tumor ECM and tumor tissue. Based on this previous study, here, we hypothesize that the electrostatically driven interactions of the cationic peptide will improve the binding and retention of immune checkpoint blockade antibodies (ICBs), ultimately enhancing their antitumor immunogenic responses. We prepared peptide antibody (Ab) conjugates by conjugating the cationic peptide to ICBs, anti-cytotoxic T lymphocyte antigen 4 (∝-CTLA4) and anti-programmed cell death ligand-1 (∝-PD-L1) Abs, using copper-free click chemistry. We confirmed an average of 1 – 2 peptides per Ab. The cationic peptide electrostatically interacted with the net negatively charged tumor ECM and improved the binding of the Abs to the tumor ECM without affecting their antigen recognition capacities. Modifying the Abs due to cationic peptide conjugation reduced the systemic exposure of the Abs and did not induce treatment-related toxicities. We quantified a significantly higher population of tumor-infiltrating CD8+T cells and a significant depletion of regulatory T cells in the tumor and tumor-draining lymph nodes upon peptide conjugation, which resulted in a better therapeutic outcome of the ICBs.

ONE SENTENCE SUMMARY

Electrostatic interaction-based intratumoral retention enhances antitumor responses of immune checkpoint blockade antibodies upon local administration.
Piezo-dependent surveillance of matrix stiffness generates transient cells that repair the basement membrane
Authors: Stricker Aubrie M. , Hutson M. Shane , Page-McCaw Andrea
Published: December 2023

Basement membranes are extracellular matrix sheets separating tissue layers and providing mechanical support, and Collagen IV (Col4) is their most abundant protein. Although basement membranes are repaired after damage, little is known about repair, including whether and how damage is detected, what cells repair the damage, and how repair is controlled to avoid fibrosis. Using the intestinal basement membrane of adultDrosophilaas a model, we show that after basement membrane damage, there is a sharp increase in enteroblasts transiently expressing Col4, or “matrix mender” cells. Enteroblast-derived Col4 is specifically required for matrix repair. The increase in matrix mender cells requires the mechanosensitive ion channel Piezo, expressed in intestinal stem cells. Matrix menders are induced by the loss of matrix stiffness, as specifically inhibiting Col4 crosslinking is sufficient for Piezo-dependent induction of matrix mender cells. Our data suggest that epithelial stem cells control basement membrane integrity by monitoring stiffness.
Sleep-Deep-Net learns sleep wake scoring from the end-user and completes each record in their style
Authors: Katsuki Fumi , Spratt Tristan J. , Brown Ritchie E. , Basheer Radhika , Uygun David S.
Published: December 2023

Sleep-wake scoring is a time-consuming, tedious but essential component of clinical and pre-clinical sleep research. Sleep scoring is even more laborious and challenging in rodents due to the smaller EEG amplitude differences between states and the rapid state transitions which necessitate scoring in shorter epochs. Although many automated rodent sleep scoring methods exist, they do not perform as well when scoring new data sets, especially those which involve changes in the EEG/EMG profile. Thus, manual scoring by expert scorers remains the gold-standard. Here we take a different approach to this problem by using a neural network to accelerate the scoring of expert scorers. Sleep-Deep-Net (SDN) creates a bespoke deep convolution neural network model for individual electroencephalographic or local-field-potential records via transfer learning of GoogleNet, by learning from a small subset of manual scores of each EEG/LFP record as provided by the end-user. SDN then automates scoring of the remainder of the EEG/LFP record. A novel REM scoring correction procedure further enhanced accuracy. SDN reliably scores EEG and LFP data and retains sleep-wake architecture in wild-type mice, in sleep induced by the hypnotic zolpidem, in a mouse model of Alzheimer’s disease and in a genetic knock-down study, when compared to manual scoring. SDN reduced manual scoring time to 1/12. Since SDN uses transfer learning on each independent recording, it is not biased by previously scored existing data sets. Thus, we find SDN performs well when used on signals altered by a drug, disease model or genetic modification.

STATEMENT OF SIGNIFICANCE

Sleep medicine is often critically advanced by translational research based onin vivoelectrophysiologic mouse data. A necessary but time-consuming step in this field is scoring epochs of recordings into wakefulness, non-rapid-eye-movement sleep and non-rapid-eye-movement sleep. Despite efforts to automate this, manual scoring remains the gold-standard since automatic methods poorly handle data that is not similar enough to data used during development. Here, we describe a novel automated sleep scoring method that involves retraining a deep-convolution-neural-net capable of computer vision to score sleep-wake patterns after learning from a small set of manual scores within a record. This avoids biasing the model to expect data to be the same as its training set from previous records.
Distinct transcriptional programs define a heterogeneous neuronal ensemble for social interaction
Authors: Walker Hailee , Frost Nicholas A.
Published: December 2023

Reliable representations of information regarding complex behaviors including social interactions require the coordinated activity of heterogeneous cell types within distributed brain regions. Activity in the medial prefrontal cortex is critical in regulating social behavior, but our understanding of the specific cell types which comprise the social ensemble has been limited by available mouse lines and molecular tagging strategies which rely on the expression of a single marker gene. Here we sought to quantify the heterogeneous neuronal populations which are recruited during social interaction in parallel in a non-biased manner and determine how distinct cell types are differentially active during social interactions. We identify distinct populations of prefrontal neurons activated by social interaction by quantification of immediate early gene (IEG) expression in transcriptomically clustered neurons. This approach revealed variability in the recruitment of different excitatory and inhibitory populations within the medial prefrontal cortex. Furthermore, evaluation of the populations of IEGs recruited following social interaction revealed both cell-type and region-specific transcriptional programs, suggesting that reliance on a single molecular marker is insufficient to quantify activation across all cell types. Our findings provide a comprehensive description of cell-type specific transcriptional programs invoked by social interactions and reveal new insights into the heterogeneous neuronal populations which compose the social ensemble.
Multicellularity and increasing Reynolds number impact on the evolutionary shift in photo-induced ciliary response in Volvocales
Authors: Ueki Noriko , Wakabayashi Ken-ichi
Published: December 2023

Volvocales include species with different cell numbers and sizes, reflecting a history of gradual size increase evolution. Unicellular species live in low Reynolds-number (Re) environments where viscous forces dominate, whereas multicellular species live in higherReenvironments with nonnegligible inertial forces. Despite significant changes in the physical environment, during the evolution of multicellularity they maintained photobehaviors (i.e., photoshock and phototactic responses), which allows them to survive under changing light conditions. In this study, we classified photo-induced ciliary responses in Volvocales into four patterns: temporal waveform conversion, no obvious response, pause in ciliary beating, temporal changes in ciliary beating directions. We found that which species exhibit which pattern depends onReassociated with the individual size of each species rather than phylogenetic relationships. These results suggest that species with increased cell numbers acquired their responses adapted to higherRefluid environments.

Significance Statement

Volvocales green algae include species with various cell numbers and are excellent organisms for studying the evolution of multicellularity. They exhibit photobehaviors by changing the pattern of ciliary beating, which could be categorized into four patterns. We found that the difference in patterns among the organisms is due to the Reynolds number, the ratio of viscous and inertia forces, rather than their phylogenetic relationships. This study indicates that the fluid environment was an important factor in natural selection for behavioral changes in microalgae during evolution. The results link evolution and physics while contributing to the design of micromachines.
Detection of genetically modified organisms using highly multiplexed amplicon sequencing
Authors: Reyes-Avila C. Sarai , Waldvogel Dominique , Pradervand Nicolas , Aubry Sylvain , Croll Daniel
Published: December 2023

The circulation of products based on genetically modified (GM) organisms is highly regulated by some governments having implemented strict rules on the breeding, planting, marketing, labelling, and trading of such products. To ensure compliance, accurate detection methods for GM events are necessary, along with assurance that GM material falls within relevant threshold levels. The increasing complexity and potential of undocumented GM are a growing challenge for genetic screening. In this study, we developed and assessed a highly multiplexed amplicon sequencing assay for the detection of GM events based on a microfluidics platform and next-generation sequencing (NGS). To probe GM events comprehensively, we designed a total of 230 new amplicons to cover flanking, promoter, junction and coding sequences of GM sequences. In addition, we designed and implemented parallel amplification of ribosomal and chloroplast markers to define crop species identity from potentially mixed samples. Using reference GM material of 11 crop species and multiple amplicons, we successfully detected the presence of 10 known modifications per GM event. We also find that reported flanking sequences of GM events may not be all useful for diagnostic. We assessed the assay’s potential to detect GM events in mixed samples as well as in highly diluted DNA. Finally, we performed a prospective search of potentially undocumented GM events in plant material. Our microfluidics-based amplicon GM detection approach fills important gaps in detecting potentially undocumented and complex GM events by recovering a wide range of specific amplicon sequences for evaluation. Integrating highly parallel amplicon assays in GM screening efforts should be an effective complement to aid post-market monitoring and regulatory compliance efforts.
News without the buzz: reading out weak theta rhythms in the hippocampus
Authors: Agarwal Gautam , Lustig Brian , Akera Seiji , Pastalkova Eva , Lee Albert K. , Sommer Friedrich T.
Published: December 2023

Local field potentials (LFPs) reflect the collective dynamics of neural populations, yet their exact relationship to neural codes remains unknown1. One notable exception is the theta rhythm of the rodent hippocampus, which seems to provide a reference clock to decode the animal’s position from spatiotemporal patterns of neuronal spiking2or LFPs3. But when the animal stops, theta becomes irregular4, potentially indicating the breakdown of temporal coding by neural populations. Here we show that no such breakdown occurs, introducing an artificial neural network that can recover position-tuned rhythmic patterns (pThetas) without relying on the more prominent theta rhythm as a reference clock. pTheta and theta preferentially correlate with place cell and interneuron spiking, respectively. When rats forage in an open field, pTheta is jointly tuned to position and head orientation, a property not seen in individual place cells but expected to emerge from place cell sequences5. Our work demonstrates that weak and intermittent oscillations, as seen in many brain regions and species, can carry behavioral information commensurate with population spike codes.
CCDC78: unveiling the function of a novel gene associated to hereditary myopathy
Authors: Lopergolo Diego , Gallus Gian Nicola , Pieraccini Giuseppe , Boscaro Francesca , Berti Gianna , Serni Giovanni , Volpi Nila , Formichi Patrizia , Bianchi Silvia , Cassandrini Denise , Sorrentino Vincenzo , Rossi Daniela , Santorelli Filippo Maria , De Stefano Nicola , Malandrini Alessandro
Published: December 2023

CCDC78was indicated about ten years ago as novel candidate gene for the autosomal dominant centronuclear myopathy-4 (CNM4). However, to date, only one family has been described and CCDC78 function remains unclear. Here we deeply analyze for the first time a family harbouring aCCDC78nonsense mutation. Histopathological features included, as novel histological hallmark, peculiar sarcoplasmic reticulum (SR) abnormalities. We provided evidence of nonsense mediated mRNA decay, defined novelCCDC78transcripts and, through transcriptome profiling, detected 1035 muscular differentially expressed genes including a series of genes involved in SR. Through coimmunoprecipitation assay and mass spectrometry studies we demonstrated that CCDC78 interacts with two pivotal SR proteins: SERCA1 and CASQ1. We also found an interaction with MYH1, ACTN2 and ACTA1. Our findings shed light on interactors and possible role of CCDC78 in skeletal muscle, thus allowing us to locate the protein in SR and to considerCCDC78as CNM4 causative gene.
Bacteroidia and Clostridia genomes collectively encode for a progressive cascade of marine polysaccharide degradation along the hindgut of the herbivorous fishKyphosus sydneyanus
Authors: Facimoto Cesar T. , Clements Kendall D. , Lindsey White W. , Handley Kim M.
Published: December 2023

The gut microbiota of the marine herbivorous fishKyphosus sydneyanusare thought to play an important role in host nutrition by supplying short-chain fatty acids (SCFA) through fermentation of dietary macroalgae. Here, we assembled 645 metagenome-assembled genomes (MAGs) from wild fish to determine the capacity of different bacterial taxa to degrade seaweed carbohydrates along the gut. Most bacteria (99%) were unclassified at the species level, highlighting taxonomic novelty dominated by Bacteroidia and Clostridia within the gut community. The presence of genes encoding endo-acting CAZymes in both phyla suggest they have a role in initiating glycan depolymerization. Bacteroidia also contributed the most to CAZyme-related gene expression in the distal hindgut, and encoded the highest densities of CAZymes within the community. In particular, the enrichment of CAZyme gene clusters (CGCs) within the Bacteroidia genusAlistipes(n = 73 versus just 59 distributed across all other taxa) points to an enhanced capacity for macroalgal polysaccharide utilization (e.g., alginate, laminarin and sulfated polysaccharides). Pairwise correlations of MAG relative abundances and encoded CAZyme compositions provide evidence of potential inter-species collaborations, whereby co-abundant MAGs exhibited complementary degradative capacities for specific substrates. Results indicated flexibility across these co-abundant groups in their capacity to source carbon (e.g., glucose or galactose-rich glycans), which possibly facilitates coexistence via niche partitioning. Our results indicate the potential for collaborative microbial carbohydrate metabolism in the gut ofK. sydneyanusby Bacteroidia and Clostridia, and suggest that members of the genusAlistipesare a metabolically and taxonomically diverse group of specialized macroalgae biomass degraders.
Combinatory differentiation of human induced pluripotent stem cells generates thymic epithelium that supports thymic crosstalk and directs dendritic- and CD4/CD8 T-cell full development
Authors: Provin Nathan , d’Arco Manon , Kervagoret Erwan , Bruneau Alexandre , Brusselle Lucas , Fourgeux Cynthia , Poschmann Jérémie , Saulquin Xavier , Guillonneau Carole , David Laurent , Giraud Matthieu
Published: December 2023

The thymus is a primary lymphoid organ playing a crucial role in immune tolerance, by educating thymocytes through a selection mediated by thymic epithelial cells (TEC). Recent advances in gene editing and immunotherapies have made in vitro generation of iPSC-derived T cells a crucial issue with promising therapeutic applications. Current state-of-the-art approaches often fail to accurately replicate the thymic niche, resulting in impaired T cell generation, particularly for CD4+ T cells. Here we address the production of functional mature iPSc-derived TECs that are able to support in vitro T cell generation. We designed a protocol allowing iPSc differentiation into thymic epithelial progenitors (TEP) through an unbiased multifactorial method based on optimal experimental design. Modulation of signaling pathways known to regulate embryonic thymus organogenesis resulted in the obtention of TEPs expressing typical thymic markers. We achieved TEP maturation into medullary and cortical TECs by setting up a hydrogel-based 3D culture supplemented in RANK ligand, resulting in the formation of thymic organoids. To assess the functionality of the generated TECs, primary hematopoietic progenitors were co-cultured in the organoids and showed significantly improved maturation in single-positive CD4 or CD8 T cells. Remarkably, our thymic organoid model shows multilineage differentiation potential, with generation of distinct dendritic cells populations in addition to the T lineages. Thus, generation of functional TECs and thymic organoids offers a practical platform for the study of thymic cellular interactions and paves the road to future cellular therapies by producing mature thymic immune cells of clinical interest.
Nitric oxide modulates contrast suppression in a subset of mouse retinal ganglion cells
Authors: Gonschorek Dominic , Goldin Matías A. , Oesterle Jonathan , Schwerd-Kleine Tom , Arlinghaus Ryan , Zhao Zhijian , Schubert Timm , Marre Olivier , Euler Thomas
Published: December 2023

Neuromodulators have major influences on the regulation of neural circuit activity across the nervous system. Nitric oxide (NO) has been shown to be a prominent neuromodulator in many circuits and has been extensively studied in the retina. Here, it has been associated with the regulation of light adaptation, gain control, and gap junctional coupling, but its effect on the retinal output, specifically on the different types of retinal ganglion cells (RGCs), is still poorly understood. In this study, we used two-photon Ca2+imaging and multi-electrode array (MEA) recordings to measure light-evoked activity of RGCs in the ganglion cell layer in theex vivomouse retina. This approach allowed us to investigate the neuromodulatory effects of NO on a cell type-level. Our findings reveal that NO selectively modulates the suppression of temporal responses in a distinct subset of contrast-suppressed RGC types, increasing their activity without altering the spatial properties of their receptive fields. Given that under photopic conditions, NO release is triggered by quick changes in light levels, we propose that these RGC types signal fast contrast changes to higher visual regions. Remarkably, we found that about one-third of the RGC types, recorded using two-photon Ca2+imaging, exhibited consistent, cell type-specific adaptational response changes throughout an experiment, independent of NO. By employing a sequential-recording paradigm, we could disentangle those additional adaptational response changes from drug-induced modulations. Taken together, our research highlights the selective neuromodulatory effects of NO on RGCs and emphasizes the need of considering non-pharmacological activity changes, like adaptation, in such study designs.
Active shape programming drivesDrosophilawing disc eversion
Authors: Fuhrmann Jana F. , Krishna Abhijeet , Paijmans Joris , Duclut Charlie , Eaton Suzanne , Popović Marko , Jülicher Frank , Modes Carl D. , Dye Natalie A.
Published: December 2023

How complex 3D tissue shape emerges during animal development remains an important open question in biology and biophysics. In this work, we study eversion of theDrosophilawing disc pouch, a 3D morphogenesis step when the epithelium transforms from a radially symmetric dome into a curved fold shape via an unknown mechanism. To explain this morphogenesis, we take inspiration from inanimate “shape-programmable” materials, which are capable of undergoing blueprinted 3D shape transformations arising from in-plane gradients of spontaneous strains. Here, we show that active, in-plane cellular behaviors can similarly create spontaneous strains that drive 3D tissue shape change and that the wing disc pouch is shaped in this way. We map cellular behaviors in the wing disc pouch by developing a method for quantifying spatial patterns of cell behaviors on arbitrary 3D tissue surfaces using cellular topology. We use a physical shape-programmability model to show that spontaneous strains arising from measured active cell behaviors create the tissue shape changes observed during eversion. We validate our findings using a knockdown of the mechanosensitive molecular motor MyoVI, which we find to reduce active cell rearrangements and disrupt wing pouch eversion. This work shows that shape programming is a mechanism for animal tissue morphogenesis and suggests that there exist intricate patterns in nature that could present novel designs for shape-programmable materials.