bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of the overall paradigm context on intensity deviant responses in healthy subjects
Authors: Yukhnovich Ekaterina A , Alter Kai , Sedley William
Published: January 2024

Three experiments have been carried out to explore Mismatch Negativity responses to intensity deviants in a roving intensity deviant paradigm in control and tinnitus groups. The first experiment used interspersed blocks of two tinnitus-like frequencies set by each participant with tinnitus, which were usually around 1/3 of an octave apart. On the other hand, two later studies used interspersed blocks tones at tinnitus-like frequencies and at 1 kHz. This was the only difference in the paradigms used, however, there were differences in the patterns displayed by the control group in the first study compared to the other two. Three groups of healthy controls were recruited to measure responses to intensity deviants when different frequencies were used for the alternating blocks. For one group, the whole experiment was set at a single frequency; for the next, blocks were played at 6 kHz and at a frequency 1/3 octave below 6 kHz (small difference); the last group was presented with blocks that had tones at 6 kHz and 1 kHz frequencies (large difference). Overall, the Mismatch Negativity responses in the small difference group were opposite to the large difference and the single frequency group. It would be useful to see whether these results generalise to other experiment designs such as attended and ignored stimulus conditions, different stimulus durations, non-isochronous, or paradigms with frequency deviants.
Genomics 2 Proteins portal: A resource and discovery tool for linking genetic screening outputs to protein sequences and structures
Authors: Kwon Seulki , Safer Jordan , Nguyen Duyen T. , Hoksza David , May Patrick , Arbesfeld Jeremy A. , Rubin Alan F. , Campbell Arthur J , Burgin Alex , Iqbal Sumaiya
Published: January 2024

Recent advances in AI-based methods have revolutionized the field of structural biology. Concomitantly, high-throughput sequencing and functional genomics technologies have enabled the detection and generation of variants at an unprecedented scale. However, efficient tools and resources are needed to link these two disparate data types – to “map” variants onto protein structures, to better understand how the variation causes disease and thereby design therapeutics. Here we present the Genomics 2 Proteins Portal (G2P; g2p.broadinstitute.org/): a human proteome-wide resource that maps 19,996,443 genetic variants onto 42,413 protein sequences and 77,923 structures, with a comprehensive set of structural and functional features. Additionally, the G2P portal generalizes the capability of linking genomics to proteins beyond databases by allowing users to interactively upload protein residue-wise annotations (variants, scores, etc.) as well as the protein structure to establish the connection. The portal serves as an easy-to-use discovery tool for researchers and scientists to hypothesize the structure-function relationship between natural or synthetic variations and their molecular phenotype.
Tracing India’s Canine Heritage through SNP-Based Haplotype Identification
Authors: Singh Dapinder , Mahajan Shashi Kant , Kashyap Neeraj , Mukhopadhyay Chandra Sekhar
Published: January 2024

Dog breeds/germplasm in India is mostly unexplored and the population structure of owned dogs has not been studied at all. The current study was designed to determine the haplotypes and explore the population structure among divergent breeds of dogs using genome-wide distributed SNPs, followed by validation of selected haplotypes through PCR-sequencing. The research employed custom ddRAD-GBS sequencing carried out using Illumina 150bp paired-end sequencing of 50 dog samples generated 2,18,433 high-quality SNPs meeting the screening criteria. The data was further analyzed for population structure assessment and haplotype identification using bash, and R-environment. Subsequently, three haplotypes (on AFAP1, CELSR1, and GBGT1 genes) were selected (based on SNP density and haplotype length) for validation via PCR followed by paired-end Sanger sequencing in seven different dog breeds (n=21). The results revealed notable connections between dog breeds from Punjab and Haryana, while the affiliation with Karnataka was found to be less pronounced. The sequencing results indicated that CELSR1 and GBGT1 genes contained SNPs, with the AFAP1 gene lacking SNPs. The results also provided insights into the molecular-level population structure, SNPs, and haplotypes of diverse dog breeds reared in India. This SNP variation could be used for molecular characterization of indigenous dogs.
Novel metagenomics analysis of stony coral tissue loss disease
Authors: Heinz Jakob M. , Lu Jennifer , Huebner Lindsay K. , Salzberg Steven L. , Sommer Markus , Rosales Stephanie M.
Published: January 2024

Stony coral tissue loss disease (SCTLD) has devastated coral reefs off the coast of Florida and continues to spread throughout the Caribbean. Although a number of bacterial taxa have consistently been associated with SCTLD, no pathogen has been definitively implicated in the etiology of SCTLD. Previous studies have predominantly focused on the prokaryotic community through 16S rRNA sequencing of healthy and affected tissues. Here, we provide a different analytical approach by applying a bioinformatics pipeline to publicly available metagenomic sequencing samples of SCTLD lesions and healthy tissues from four stony coral species. To compensate for the lack of coral reference genomes, we used data from apparently healthy coral samples to approximate a host genome and healthy microbiome reference. These reads were then used as a reference to which we matched and removed reads from diseased lesion tissue samples, and the remaining reads associated only with disease lesions were taxonomically classified at the DNA and protein levels. For DNA classifications, we used a pathogen identification protocol originally designed to identify pathogens in human tissue samples, and for protein classifications, we used a fast protein sequence aligner. To assess the utility of our pipeline, a species-level analysis of a candidate genus,Vibrio, was used to demonstrate the pipeline’s effectiveness. Our approach revealed both complementary and unique coral microbiome members compared to a prior metagenome analysis of the same dataset.

Article Summary

Studies of stony coral tissue loss disease (SCTLD), a devastating coral disease, have primarily used 16S rRNA sequencing approaches to identify putative pathogens. In contrast, this study applied human tissue pathogen identification protocols to SCTLD metagenomic DNA samples. Diseased samples were filtered of host sequences using a k-mer based method since host genomes were unavailable. DNA and protein-level classifications from this novel approach revealed both complementary and unique microbiome members compared to a prior metagenome analysis of the same dataset.
Cardiac Localized Polycystin-2 plays a Functional Role in Natriuretic Peptide Production and its Absence Contributes to Hypertension
Authors: Elliott Brandon , Márquez-Nogueras Karla M. , Thuo Paula , DiNello Elisabeth , Knutila Ryne M. , Fritzmann Geena E. , Willis Monte , Chapman Arlene B. , Cao Quan , Barefield David Y. , Kuo Ivana Y.
Published: January 2024

Cardiovascular complications are the most common cause of mortality in patients with autosomal dominant polycystic kidney disease (ADPKD). Hypertension is seen in 70% of patients by the age of 30 prior to decline in kidney function. The natriuretic peptides (NPs), atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), are released by cardiomyocytes in response to membrane stretch, increasing urinary excretion of sodium and water. Mice heterozygous forPkd2have attenuated NP responses and we hypothesized that cardiomyocyte-localized polycystin proteins contribute to production of NPs. Cardiomyocyte-specific knock-out models of polycystin-2 (PC2), one of the causative genes of ADPKD, demonstrate diurnal hypertension. These mice have decreased ANP and BNP expression in the left ventricle. Analysis of the pathways involved in production, maturation, and activity of NPs identified decreased transcription of CgB, PCSK6, and NFAT genes in cPC2-KOs. Engineered heart tissue with human iPSCs driven into cardiomyocytes with CRISPR/Cas9 KO ofPKD2failed to produce ANP. These results suggest that PC2 in cardiomyocytes are involved in NP production and lack of cardiac PC2 predisposes to a hypertensive volume expanded phenotype, which may contribute to the development of hypertension in ADPKD.
Biofilm formation and maize root-colonization of seed-endophytic Bacilli isolated from native maize landraces
Authors: Gastélum Gabriela , Ángeles Alejandra , Arellano-Wattenbarger Guillermo , Coronado Yaxk’in , Guevara Eduardo , Rocha Jorge
Published: January 2024

Agricultural microbiology seeks to replace the use of agrochemicals with microbe-based products. Plant growth-promoting bacteria (PGPB) are often selected based on their functionsin vitro, and then, their effect on plant development is tested. However, this approach neglects the study of their survival in soil, root-colonization, and the monitoring of beneficial functions in the rhizosphere. This could explain the recurrent lack of success in the transition from lab tests to field applications of natural isolates from novel habitats. In our recent studies, we found that native maize seeds from traditional agroecosystems carry endophytic bacterial communities that are dominated by members of the class Bacilli. As an approach to grasp their PGP potential, we developed protocols to test maize root-colonization of these natural isolates in 1) a short-term hydroponics assayin vitroand 2) a long-term assay in non-sterile soil pots. Our results show thatin vitrobiofilm formation was only partially associated to short-term colonizationin vitro; furthermore, long-term root-colonization in soil pots was not correlated to thein vitroassays. This work highlights the necessity to incorporate root-colonization assays as part of the research strategies in the search for PGPB natural isolates from unexplored habitats, towards the generation of inoculants with increased success in the field.
Non-genetic adaptation by collective migration
Authors: Vo Lam , Avgidis Fotios , Mattingly Henry H. , Edmonds Karah , Burger Isabel , Balasubramanian Ravi , Shimizu Thomas S. , Kazmierczak Barbara I. , Emonet Thierry
Published: January 2024

Cell populations must adjust their phenotypic composition to adapt to changing environments. One adaptation strategy is to maintain distinct phenotypic subsets within the population and to modulate their relative abundances via gene regulation. Another strategy involves genetic mutations, which can be augmented by stress-response pathways. Here, we studied how a migrating bacterial population regulates its phenotypic distribution to traverse diverse environments. We generated isogenicEscherichia colipopulations with varying distributions of swimming behaviors and observed their phenotype distributions during migration in liquid and porous environments. We found that the migrating populations became enriched with high-performing swimming phenotypes in each environment, allowing the populations to adapt without requiring mutations or gene regulation. This adaptation is dynamic and rapid, reversing in a few doubling times when migration ceases. By measuring the chemoreceptor abundance distributions during migration towards different attractants, we demonstrated that adaptation acts on multiple chemotaxis-related traits simultaneously. These measurements are consistent with a general mechanism in which adaptation results from a balance between cell growth generating diversity and collective migration eliminating under-performing phenotypes. Thus, collective migration enables cell populations with continuous, multi-dimensional phenotypes to flexibly and rapidly adapt their phenotypic composition to diverse environmental conditions.

Significance statement

Conventional cell adaptation mechanisms, like gene regulation and stochastic phenotypic switching, act swiftly but are limited to a few traits, while mutation-driven adaptations unfold slowly. By quantifying phenotypic diversity during bacterial collective migration, we discovered an adaptation mechanism that rapidly and reversibly adjusts multiple traits simultaneously. By balancing the generation of diversity through growth with the loss of phenotypes unable to keep up, this process tunes the phenotypic composition of migrating populations to the environments they traverse, without gene regulation or mutations. Given the prevalence of collective migration in microbes, cancers, and embryonic development, non-genetic adaptation through collective migration may be a universal mechanism for populations to navigate diverse environments, offering insights into broader applications across various fields.
Background food influences rate of encounter and efficacy of rodenticides in wild house mice
Authors: Brown Peter R. , Henry Steve , Hinds Lyn A. , Robinson Freya , Duncan Richard P. , Ruscoe Wendy A.
Published: January 2024

Baiting is widely used in wildlife management for various purposes, including lethal control, fertility control, disease and parasite control, and conditioned aversion programs for many invasive vertebrate species. The efficacy of baiting programs relies on the likelihood that target animals will encounter the bait, consume it, and receive an appropriate dose of the active ingredient. However, there has been little focus on encounter rate of toxic baits combined with behavioural aversion, which are likely to be significant factors affecting efficacy.

The likelihood of an animal encountering and consuming a toxic grain bait should theoretically increase in proportion to its availability relative to background food quantity if it is neither more or less detectable or palatable. Furthermore, the probability of consuming toxic baits might also be influenced by bait aversion following ingestion of a non-lethal dose of toxin.

Using a model system of wild house mice (Mus musculusL.) in mouse-proof enclosures in Australia, we manipulated background food, applied zinc phosphide (ZnP) baits and measured the mortality of mice. When background food was scarce, mouse mortality was high, whereas an increasing abundance of background food led to reduced mortality. A scenario modelling random encounters and including bait aversion explained 78% of the variation in observed mortality outcomes and achieved a closer fit to the data. Mortality rates were predicted to be higher with a higher strength bait which overcomes behavioural aversion.

Ensuring that animals locate and consume a lethal dose of toxic bait is a critical factor for successful bait delivery and efficacy. This is particularly significant in toxic baiting programs, where sublethal doses can make animals feel sick, leading to a negative association with the bait, and the development of aversion.

Synthesis and applications:Our findings explain why some toxic baiting programs might fail. To achieve successful control, efforts should be directed at reducing the availability of background food to increase the probability of encounter and uptake of toxic baits. It is important to measure and understand the role of background food on toxic baiting programs to explain variable outcomes and inform strategies for successful bait delivery.

Graphical abstract

Our findings explain why some toxic baiting programs might fail. To achieve successful control, efforts should be directed at reducing the availability of background food to increase the probability of encounter and uptake of toxic baits. It is important to measure and understand the role of background food on toxic baiting programs to explain variable outcomes and inform strategies for successful bait delivery.
Metabolomics unveiled the impact of alterations in nucleotide glucose metabolism on palmitic acid-induced TLR4 activation in THP-1 macrophages
Authors: Wu Niting , Ran Xiaodong , Wang Jiawei , Wang Pengbo , Li Lin , She Yuanting , Luo Yadan , Li Xiaohui , Jia Yi , Huang Yan
Published: January 2024

OBJECTIVE

The association of palmitic acid with macrophage inflammation and its promotion of the expression of inflammatory factors through TLR4 have been demonstrated. It has been observed that immature TLR4 localizes to the endoplasmic reticulum and Golgi apparatus, necessitating glycosylation for migration to the cell membrane. The objective of this study was to identify potential biomarkers associated with N-glycosylation subsequent to the activation of TLR4 inflammatory signaling in human macrophages by palmitic acid.

APPROACH AND RESULTS

The co-cultivation of palmitic acid with THP-1 macrophages was conducted for a duration of 24 hours, followed by the collection of cell extracts for subsequent metabolomic and lipidomic analyses using high performance liquid chromatography-tandem mass spectrometry. Multivariate and univariate statistical analyses were conducted to identify potential biomarkers, in accordance with established scientific protocols. The impact of palmitic acid on the TLR4 signaling pathway and macrophage N-glycosylation was assessed at various time points using Western blot analysis, immunofluorescence staining, Elisa assays, and chemical labeling techniques. The TLR4 inflammatory signaling pathway was examined in macrophages at different time points, revealing that PA induced the upregulation of MyD88 and TRAF6 expression as well as NF-κB phosphorylation, indicating the activation of classical NF-κB signaling. After 24 hours, TLR4 translocated from the cell membrane to the cytoplasm and initiated internalization, accompanied by significant colocalization with GalAz in the cytosol. In addition, the metabolites in the cell extract were found to be altered in both the control and model groups. Significantly alterations in two N-glycosylation related metabolites were observed in the model group, including guanosine diphosphate-L-fucose and uridine diphosphate-N-acetylglucosamine/uridine diphosphate-N-acetylgalactosamine.

CONCLUSIONS

THP-1 macrophages incubated with palmitic acid exhibited a distinct metabolomic profile compared to the control group. Our findings suggest that metabolomics analysis holds promise in identifying disease-specific biomarkers for diagnosing fatty acid-induced inflammatory responses in macrophages.
Spatiotemporal Mapping and Molecular Basis of Whole-brain Circuit Maturation
Authors: Xue Jian , Brawner Andrew T. , Thompson Jacqueline R. , Yelhekar Tushar D. , Newmaster Kyra T. , Qiu Qiang , Cooper Yonatan A. , Yu C. Ron , Ahmed-Braima Yasir H. , Kim Yongsoo , Lin Yingxi
Published: January 2024

Brain development is highly dynamic and asynchronous, marked by the sequential maturation of functional circuits across the brain. The timing and mechanisms driving circuit maturation remain elusive due to an inability to identify and map maturing neuronal populations. Here we create DevATLAS (Developmental Activation Timing-based Longitudinal Acquisition System) to overcome this obstacle. We develop whole-brain mapping methods to construct the first longitudinal, spatiotemporal map of circuit maturation in early postnatal mouse brains. Moreover, we uncover dramatic impairments within the deep cortical layers in a neurodevelopmental disorders (NDDs) model, demonstrating the utility of this resource to pinpoint when and where circuit maturation is disrupted. Using DevATLAS, we reveal that early experiences accelerate the development of hippocampus-dependent learning by increasing the synaptically mature granule cell population in the dentate gyrus. Finally, DevATLAS enables the discovery of molecular mechanisms driving activity-dependent circuit maturation.
Dynamical constraints on neural population activity
Authors: Oby Emily R. , Degenhart Alan D. , Grigsby Erinn M. , Motiwala Asma , McClain Nicole T. , Marino Patrick J. , Yu Byron M. , Batista Aaron P.
Published: January 2024

The manner in which neural activity unfolds over time is thought to be central to sensory, motor, and cognitive functions in the brain. Network models have long posited that the brain’s computations involve time courses of activity that are shaped by the underlying network. A prediction from this view is that the activity time courses should be difficult to violate. We leveraged a brain-computer interface (BCI) to challenge monkeys to violate the naturally-occurring time courses of neural population activity that we observed in motor cortex. This included challenging animals to traverse the natural time course of neural activity in a time-reversed manner. Animals were unable to violate the natural time courses of neural activity when directly challenged to do so. These results provide empirical support for the view that activity time courses observed in the brain indeed reflect the underlying network-level computational mechanisms that they are believed to implement.
YY1 is a transcriptional activator of mouse LINE-1 Tf subfamily
Authors: Saha Karabi , Nielsen Grace I. , Nandani Raj , Kong Lingqi , Ye Ping , An Wenfeng
Published: January 2024

Long interspersed element type 1 (LINE-1, L1) is an active autonomous transposable element (TE) in the human genome. The first step of L1 replication is transcription, which is controlled by an internal RNA polymerase II promoter in the 5’ untranslated region (UTR) of a full-length L1. It has been shown that transcription factor YY1 binds to a conserved sequence motif at the 5’ end of the human L1 5’UTR and dictates where transcription initiates but not the level of transcription. Putative YY1-binding motifs have been predicted in the 5’UTRs of two distinct mouse L1 subfamilies, Tf and Gf. Using site-directed mutagenesis, in vitro binding, and gene knockdown assays, we experimentally tested the role of YY1 in mouse L1 transcription. Our results indicate that Tf, but not Gf subfamily, harbors functional YY1-binding sites in its 5’UTR monomers. In contrast to its role in human L1, YY1 functions as a transcriptional activator for the mouse Tf subfamily. Furthermore, YY1-binding motifs are solely responsible for the synergistic interaction between monomers, consistent with a model wherein distant monomers act as enhancers for mouse L1 transcription. The abundance of YY1-binding sites in Tf elements also raise important implications for gene regulation at the genomic level.
Suppression of epileptic seizures by transcranial activation of K+-selective channelrhodopsin
Authors: Duan Xiaodong , Zhang Chong , Wu Yujie , Ju Jun , Xu Zhe , Li Xuanyi , Liu Yao , Ohdah Schugofa , Constantin Oana M. , Lu Zhonghua , Wang Cheng , Chen Xiaojing , Gee Christine E. , Nagel Georg , Hou Sheng-Tao , Gao Shiqiang , Song Kun
Published: January 2024

Optogenetics is a valuable tool for studying the mechanisms of neurological diseases and is now being developed for therapeutic applications. In rodents and macaques, improved channelrhodopsins have been applied to achieve transcranial optogenetic stimulation. While transcranial photoexcitation of neurons has been achieved, noninvasive optogenetic inhibition for treating hyperexcitability-induced neurological disorders has remained elusive. There is a critical need for effective inhibitory optogenetic tools that are highly light-sensitive and capable of suppressing neuronal activity in deep brain tissue. In this study, we developed a highly sensitive K+-conductive channelrhodopsin (hsKCR) by molecular engineering of the recently discoveredHyphochytrium catenoideskalium (potassium) channelrhodopsin 1. Transcranial activation of hsKCR significantly prolongs the time to the first seizure, increases survival, and decreases seizure activity in several mouse epileptic models. Our approach for transcranial optogenetic inhibition of neural hyperactivity may be adapted for cell type-specific neuromodulation in both basic and preclinical settings.
Medial prefrontal cortex suppresses reward-seeking behavior with risk of positive punishment by reducing sensitivity to reward
Authors: Nishio Monami , Kondo Masashi , Yoshida Eriko , Matsuzaki Masanori
Published: January 2024

Reward-seeking behavior is frequently associated with risk of punishment. There are two types of punishment: positive, resulting in an unpleasant outcome, and negative, resulting in omission of a reinforcing outcome. Although the medial prefrontal cortex (mPFC) is important in avoiding punishment, whether it is important for avoiding both positive and negative punishment and how it contributes to such avoidance are not clear. In this study, we trained male mice to perform decision-making tasks under the risks of positive (air-puff stimulus) and negative (reward omission) punishment. We found that pharmacological inactivation of mPFC enhanced the reward-seeking choice under the risk of positive, but not negative, punishment. In reinforcement learning models, this behavioral change was well-explained by hypersensitivity to the reward, rather than a decrease in the strength of aversion to punishment. Our results suggest that mPFC suppresses reward-seeking behavior by reducing sensitivity to reward under the risk of positive punishment.
A combinatory genetic strategy for targeting neurogliaform neurons in the mouse basolateral amygdala
Authors: Ozsvár Attila , Sieburg Meike Claudia , Sietam Monica Dahlstrup , Hou Wen-Hsien , Capogna Marco
Published: January 2024

The mouse basolateral amygdala (BLA) contains various GABAergic interneuron subpopulations, which have distinctive roles in the neuronal microcircuit controlling numerous behavioral functions.

In mice, roughly 15% of the BLA GABAergic interneurons express neuropeptide Y (NPY), a reasonably characteristic marker for neurogliaform cells (NGFCs) in cortical-like brain structures. However, genetically labeled putative NPY-expressing interneurons in the BLA yield a mixture of interneuron subtypes besides NGFCs. Thus, selective molecular markers are lacking for genetically accessing NGFCs in the BLA.

Here, we validated the NGFC-specific labeling with a molecular marker, neuron-derived neurotrophic factor (NDNF), in the mouse BLA, as such specificity has been demonstrated in the neocortex and hippocampus. We characterized genetically defined NDNF-expressing (NDNF+) GABAergic interneurons in the mouse BLA by combining the Ndnf-IRES2-dgCre-D transgenic mouse line with viral labeling, immunohistochemical staining, and in vitro electrophysiology.

We found that BLA NDNF+ GABAergic cells mainly expressed NGFC neurochemical markers NPY and reelin (Reln) and exhibited small round soma and dense axonal arborization. Whole-cell patch clamp recordings indicated that most NDNF+ interneurons showed late spiking and moderate firing adaptation. Moreover, ∼81% of BLA NDNF+ cells generated retroaxonal action potential after current injections or optogenetic stimulations, frequently developing into persistent barrage firing. Optogenetic activation of the BLA NDNF+ cell population yielded both GABAA- and GABABreceptor-mediated currents onto BLA pyramidal neurons (PNs). We demonstrate a combinatory strategy combining the NDNF-cre mouse line with viral transfection to specifically target adult mouse BLA NGFCs and further explore their functional and behavioral roles.
Multimodal profiling reveals tissue-directed signatures of human immune cells altered with age
Authors: Wells Steven B. , Rainbow Daniel B. , Mark Michal , Szabo Peter A. , Ergen Can , Maceiras Ana Raquel , Caron Daniel P. , Rahmani Elior , Benuck Eli , Amiri Valeh Valiollah Pour , Chen David , Wagner Allon , Howlett Sarah K. , Jarvis Lorna B. , Ellis Karen L. , Kubota Masaru , Matsumoto Rei , Mahbubani Krishnaa , Saeb-Parsy Kouresh , Dominguez-Conde Cecilia , Richardson Laura , Xu Chuan , Li Shuang , Mamanova Lira , Bolt Liam , Wilk Alicja , Teichmann Sarah A. , Farber Donna L. , Sims Peter A. , Jones Joanne L. , Yosef Nir
Published: January 2024

The immune system comprises multiple cell lineages and heterogeneous subsets found in blood and tissues throughout the body. While human immune responses differ between sites and over age, the underlying sources of variation remain unclear as most studies are limited to peripheral blood. Here, we took a systems approach to comprehensively profile RNA and surface protein expression of over 1.25 million immune cells isolated from blood, lymphoid organs, and mucosal tissues of 24 organ donors aged 20-75 years. We applied a multimodal classifier to annotate the major immune cell lineages (T cells, B cells, innate lymphoid cells, and myeloid cells) and their corresponding subsets across the body, leveraging probabilistic modeling to define bases for immune variations across donors, tissue, and age. We identified dominant tissue-specific effects on immune cell composition and function across lineages for lymphoid sites, intestines, and blood-rich tissues. Age-associated effects were intrinsic to both lineage and site as manifested by macrophages in mucosal sites, B cells in lymphoid organs, and T and NK cells in blood-rich sites. Our results reveal tissue-specific signatures of immune homeostasis throughout the body and across different ages. This information provides a basis for defining the transcriptional underpinnings of immune variation and potential associations with disease-associated immune pathologies across the human lifespan.
TIR domains produce histidine-ADPR conjugates as immune signaling molecules in bacteria
Authors: Sabonis Dziugas , Avraham Carmel , Lu Allen , Herbst Ehud , Silanskas Arunas , Leavitt Azita , Yirmiya Erez , Zaremba Mindaugas , Amitai Gil , Kranzusch Philip J. , Sorek Rotem , Tamulaitiene Giedre
Published: January 2024

TIR domains are central components of pattern recognition immune proteins across all domains of life. In both bacteria and plants, TIR-domain proteins were shown to recognize pathogen invasion and then produce immune signaling molecules exclusively comprising nucleotide moieties. Here we show that the TIR domain protein of the type II Thoeris defense system in bacteria produces a unique signaling molecule comprising the amino acid histidine conjugated to ADP-ribose (His-ADPR). His-ADPR is generated in response to phage infection and activates the cognate Thoeris effector by binding a Macro domain located at the C-terminus of the effector protein. By determining the crystal structure of a ligand-bound Macro domain, we describe the structural basis for His-ADPR recognition. Our analyses furthermore reveal a family of phage proteins that bind and sequester His-ADPR signaling molecules, allowing phages to evade TIR- mediated immunity. These data demonstrate diversity in bacterial TIR signaling and reveal a new class of TIR-derived immune signaling molecules combining nucleotide and amino acid moieties.
Spatially resolved quantification of oxygen consumption rate in ex vivo lymph node slices
Authors: Anbaei Parastoo , Stevens Marissa G. , Ball Alexander G. , Bullock Timothy N.J. , Pompano Rebecca R.
Published: January 2024

Cellular metabolism has been closely linked to activation state in cells of the immune system, and the oxygen consumption rate (OCR) in particular serves as a valuable metric for assessing metabolic activity. Several oxygen sensing assays have been reported for cells in standard culture conditions. However, none have provided a spatially resolved, optical measurement of local oxygen consumption in intact tissue samples, making it challenging to understand regional dynamics of consumption. Therefore, here we established a system to monitor the rates of oxygen consumption in ex vivo tissue slices, using murine lymphoid tissue as a case study. By integrating an optical oxygen sensor into a sealed perfusion chamber and incorporating appropriate correction for photobleaching of the sensor and of tissue autofluorescence, we were able to visualize and quantify rates of oxygen consumption in tissue. This method revealed for the first time that the rate of oxygen consumption in naïve lymphoid tissue was higher in the T cell region compared to the B cell and cortical regions. To validate the method, we measured OCR in the T cell regions of naïve lymph node slices using the optical assay and estimated the consumption rate per cell. The predictions from the optical assay were similar to reported values and were not significantly different from those of the Seahorse metabolic assay, a gold standard method for measuring OCR in cell suspensions. Finally, we used this method to quantify the rate of onset of tissue hypoxia for lymph node slices cultured in a sealed chamber and showed that continuous perfusion was sufficient to maintain oxygenation. In summary, this work establishes a method to monitor oxygen consumption with regional resolution in intact tissue explants, suitable for future use to compare tissue culture conditions and responses to stimulation.

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Interspecies surfactants serve as public goods enabling surface motility inPseudomonas aeruginosa
Authors: Warrell Delayna L. , Zarrella Tiffany M. , Machalek Christopher , Khare Anupama
Published: January 2024

In most natural environments, bacteria live in polymicrobial communities where secreted molecules from neighboring species alter bacterial behaviors including motility, but such interactions are understudied.Pseudomonas aeruginosais a motile opportunistic pathogen that exists in diverse multispecies environments such as the soil and is frequently found in human wound and respiratory tract co-infections with other bacteria includingStaphylococcus aureus. Here we show thatP. aeruginosacan co-opt secreted surfactants from other species for flagellar-based surface motility. We found that exogenous surfactants fromS. aureus, other bacteria, and interkingdom species enabledP. aeruginosato switch from swarming to an alternative surface spreading motility on semi-solid surfaces and allowed for the emergence of surface motility on hard agar whereP. aeruginosawas otherwise unable to move. This motility was distinct from the response of other motile bacteria in the presence of exogenous surfactants. Mutant analysis indicated that thisP. aeruginosamotility was similar to a previously described mucin-based motility, ‘surfing’, albeit with divergent regulation. Thus, our study demonstrates that secreted surfactants from the host as well as neighboring bacterial and interkingdom species act as public goods facilitatingP. aeruginosaflagella-mediated surfing-like surface motility, thereby allowing it to access different environmental niches.
SAXS/MC studies of the mixed-folded protein Cdt1 reveal monomeric, folded over conformations
Authors: Smith Kyle P. , Chakravarthy Srinivas , Rahi Amit , Chakraborty Manas , Vosberg Kristen M. , Tonelli Marco , Plach Maximilian G. , Grigorescu Arabela A. , Curtis Joseph E. , Varma Dileep
Published: January 2024

Cdt1 is a protein critical for DNA replication licensing and is well-established to be a binding partner of the minichromosome maintenance (MCM) complex. Cdt1 has also been demonstrated to have an emerging, “moonlighting” role at the kinetochore via direct binding to microtubules and to the Ndc80 complex. However, it is not known how the structure and conformations of Cdt1 could allow for these multiple, completely unique sets of protein complexes. And while there exist multiple robust methods to study entirely folded or entirely unfolded proteins, structure-function studies of combined, mixed folded/disordered proteins remain challenging. It this work, we employ multiple orthogonal biophysical and computational techniques to provide a detailed structural characterization of human Cdt1 92-546. DSF and DSCD show both folded winged helix (WH) domains of Cdt1 are relatively unstable. CD and NMR show the N-terminal and the linker regions are intrinsically disordered. Using DLS and SEC-MALS, we show that Cdt1 is polydisperse, monomeric at high concentrations, and without any apparent inter-molecular self-association. SEC-SAXS of the monomer in solution enabled computational modeling of the proteinin silico. Using the program SASSIE, we performed rigid body Monte Carlo simulations to generate a conformational ensemble. Using experimental SAXS data, we filtered for conformations which did and did not fit our data. We observe that neither fully extended nor extremely compact Cdt1 conformations are consistent with our SAXS data. The best fit models have the N-terminal and linker regions extended into solution and the two folded domains close to each other in apparent “folded over” conformations. The best fit Cdt1 conformations are consistent with a function as a scaffold protein which may be sterically blocked without the presence of binding partners. Our studies also provide a template for combining experimental and computational biophysical techniques to study mixed-folded proteins.
Retention of ES cell-derived 129S genome drives NLRP1 hypersensitivity and transcriptional deregulation inNlrp3−/−mice
Authors: Weiss Felix D. , Alvarez Yubell , Sahu Anshupa , Shakeri Farhad , Lee Hye Eun , Gellner Anne-Kathrin , Buness Andreas , Latz Eicke , Meissner Felix
Published: January 2024

Immune response genes are highly polymorphic in humans and mice, with heterogeneity amongst loci driving strain-specific host defense responses. The inadvertent retention of polymorphic loci can introduce confounding phenotypes, leading to erroneous conclusions, and impeding scientific advancement. In this study, we employ a combination of RNAseq and variant calling analyses and identify a substantial region of 129S genome, including the highly polymorphicNlrp1locus proximal toNlrp3, in one of the most commonly used mouse models of NLRP3 deficiency. We show that increased expression of 129S NLRP1b sensitizesNlrp3−/−macrophages to NLRP1 inflammasome activation. Furthermore, the presence of 129S genome leads to altered gene and protein regulation across multiple cell-types, including of the key tissue-resident macrophage marker, TIM4. To address the challenge of resolving NLRP3-dependent phenotypes, we introduce and validate a conditionalNlrp3allele, enabling precise temporal and cell-type-specific control overNlrp3deletion. Our study establishes a generic framework to identify functionally relevant SNPs and assess genomic contamination in transgenic mice. This allows for unambiguous attribution of phenotypes to the target gene and advances the precision and reliability of research in the field of host defense responses.
Synergistic Modulation of Macrophages by Methotrexate and RELA siRNA Folate-Liposome: A Precision Therapy to Prevent Joint Degradation in Collagen-Induced Arthritic Rats
Authors: Nasra Simran , Bhatia Dhiraj , Kumar Ashutosh
Published: January 2024

Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by inflammation and joint destruction. Current treatments, such as Methotrexate (MTX), while effective, often have therapeutic limitations like high plasma Cmaxand lack of sustained release. This study explores a synergistic approach to RA therapy using folate-liposomal co-delivery of MTX and RELA siRNA, aimed at RAW264.7 macrophage repolarization through inhibition of the NF-κB pathway. Extensive invitro characterizations demonstrate the stability and biocompatibility of this combinatorial therapy in folate-liposomes. In collagen-induced arthritis (CIA) rat model, we observed a reduction in synovial inflammation and improved mobility following treatment. The combined MTX and RELA siRNA approach indirectly inhibits inflammatory cytokines and other biochemical parameters such as Rheumatoid factor (RF) and C-reactive protein (CRP). The targeted macrophage delivery yields a marked therapeutic effect in RAW264.7 murine macrophages, potentially modulating the M1 to M2 macrophage polarization. Overall, this research presents a promising avenue for innovative therapies in RA management by inhibiting the inflammatory cascade and preventing joint damage.
Sequence-based modelling of bacterial genomes enables accurate antibiotic resistance prediction
Authors: Wiatrak Maciej , Weimann Aaron , Dinan Adam , Brbić Maria , Floto R. Andres
Published: January 2024

Abstract

Rapid detection of antibiotic-resistant bacteria and understanding the mecha- nisms underlying antimicrobial resistance (AMR) are major unsolved problems that pose significant threats to global public health. However, existing methods for predicting antibiotic resistance from genomic sequence data have had lim- ited success due to their inability to model epistatic effects and generalize to novel variants. Here, we present GeneBac, a deep learning method for predicting antibiotic resistance from DNA sequence through the integration of interactions between genes. We apply GeneBac to two distinct bacterial species and show that it can successfully predict the minimum inhibitory concentration (MIC) of multiple antibiotics. We use the WHO Mycobacterium tuberculosis mutation cat- alogue to demonstrate that GeneBac accurately predicts the effects of different variants, including novel variants that have not been observed during training. GeneBac is a modular framework which can be applied to a number of tasks including gene expression prediction, resistant gene identification and strain clus- tering. We leverage this modularity to transfer learn from the transcriptomic data to improve performance on the MIC prediction task.
Thermal energetics of bats of the family Vespertilionidae: an evolutionary approach
Authors: Ayala-Berdon Jorge , Medina-Bello Kevin I. , Carballo-Morales Jorge D. , Saldaña-Vázquez Romeo A. , Villalobos Federico
Published: January 2024

Thermal energetics define the way animals spend energy for thermoregulation. In this regard, numerous studies have determined that body mass (Mb) is the most influential morphological trait affecting the thermal traits in different species of birds and mammals. However, most of the studies have been focused on the basal metabolic rate (BMR), while other thermal traits have been less studied.

We addressed this gap by examining thermal variables on bats of the family Vespertilionidae. Using open-flow respirometry, we measuredBMR, absolute thermal conductance (C’), lower and upper critical temperatures (TLCandTUC), and the breadth of the thermoneutral zone (TNZb) of 15 bat species varying inMbfrom ∼ 4.0 to 21.0 g from central Mexico. We: 1) combined our empirical data with information gathered from the bibliography and conducted phylogenetic analyses to investigate the relationship betweenMband thermal traits, and 2) mapped the thermal energetic values along the phylogeny to explore how they may have evolved.

We found a positive relationship betweenMbandBMRand absoluteC’, and a negative relationship betweenMbandTLCandTUC. However, we did not find a relationship betweenMbandTNZbin bats. The phylogenetic approach suggested that over the evolutionary history of bats,BMRandC’ have decreased whileTLCandTUChave increased.

Our results suggest that adaptive changes inMband thermal traits may have influenced the geographical distribution and the use of energy-saving strategies of the different species of bats of the family Vespertilionidae.
Microbiome-induced Increases and Decreases in Bone Tissue Strength can be Initiated After Skeletal Maturity
Authors: Liu C , Cyphert E.L. , Stephen S.J. , Wang B. , Morales A.L. , Nixon J.C. , Natsoulas N.R. , Garcia M. , Blazquez Carmona P. , Vill A.C. , Donnelly E.L. , Brito I.L. , Vashishth D. , Hernandez C.J.
Published: January 2024

Recent studies in mice have indicated that the gut microbiome can regulate bone tissue strength. However, prior work involved modifications to the gut microbiome in growing animals and it is unclear if the same changes in the microbiome, applied later in life, would change matrix strength. Here we changed the composition of the gut microbiome before and/or after skeletal maturity (16 weeks of age) using oral antibiotics (ampicillin + neomycin). Male and female mice (n=143 total, n=12-17/group/sex) were allocated into five study groups:1) Unaltered, 2) Continuous (dosing 4-24 weeks of age), 3) Delayed (dosing only 16-24 weeks of age), 4) Initial (dosing 4-16 weeks of age, suspended at 16 weeks), and 5) Reconstituted (dosing from 4-16 weeks following by fecal microbiota transplant from Unaltered donors). Animals were euthanized at 24 weeks of age. In males, bone matrix strength in the femur was 25-35% less than expected from geometry in mice from the Continuous (p= 0.001), Delayed (p= 0.005), and Initial (p=0.040) groups as compared to Unaltered. Reconstitution of the gut microbiota, however, led to a bone matrix strength similar to Unaltered animals (p=0.929). In females, microbiome-induced changes in bone matrix strength followed the same trend as males but were not significantly different, demonstrating sex-related differences in the response of bone matrix to the gut microbiota. Minor differences in chemical composition of bone matrix were observed (Raman spectroscopy). Our findings indicate that microbiome-induced impairment of bone matrix in males can be initiated and/or reversed after skeletal maturity. The portion of the femoral cortical bone formed after skeletal maturity (16 weeks) is small; however, this suggests that microbiome-induced changes in bone matrix occur without osteoblast/osteoclast turnover using an, as of yet unidentified mechanism. These findings add to evidence that the mechanical properties of bone matrix can be altered in the adult skeleton.