bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantitative single-cell transcriptome-based ranking of engineered AAVs in human retinal explants
Authors: Xi Zhouhuan , Öztürk Bilge E. , Johnson Molly E. , Stauffer William R. , Byrne Leah C.
Published: September 2021

Gene therapy is a rapidly developing field, and adeno-associated virus (AAV) is a leading viral vector candidate for therapeutic gene delivery. Newly engineered AAVs with improved abilities are now entering the clinic. It has proven challenging, however, to predict the translational potential of gene therapies developed in animal models, due to cross-species differences. Human retinal explants are the only available model of fully developed human retinal tissue, and are thus important for the validation of candidate AAV vectors. In this study, we evaluated 18 wildtype and engineered AAV capsids in human retinal explants using a recently developed single-cell RNA-Seq AAV engineering pipeline (scAAVengr). Human retinal explants retained the same major cell types as fresh retina, with similar expression of cell-specific markers, except for a cone population with altered expression of cone-specific genes. The efficiency and tropism of AAVs in human explants were quantified, with single-cell resolution. The top performing serotypes, K91, K912, and 7m8, were further validated in non-human primate and human retinal explants. Together, this study provides detailed information about the transcriptome profiles of retinal explants, and quantifies the infectivity of leading AAV serotypes in human retina, accelerating the translation of retinal gene therapies to the clinic.

Graphic abstract
Regional and clonal T cell dynamics at single cell resolution in immune checkpoint blockade
Authors: Pai Joy A. , Chow Andrew , Sauter Jennifer L. , Mattar Marissa , Rizvi Hira , Woo Hyung Jun , Shah Nisargbhai , Uddin Fathema , Quintanal-Villalonga Alvaro , Chan Joseph M. , Manoj Parvathy , Allaj Viola , Baine Marina , Chaft Jamie , Plodkowski Andrew J. , Won Helen , Wells Daniel , Donoghue Mark T.A. , de Stanchina Elisa , Sen Triparna , Wolchok Jedd D. , Houck-Loomis Brian , Merghoub Taha , Rudin Charles M. , Satpathy Ansuman T. , Hellmann Matthew D.
Published: September 2021

Paired T cell receptor and RNA single cell sequencing (scTCR/RNA-seq) has allowed for enhanced resolution of clonal T cell dynamics in cancer. Here, we report a scTCR/RNA-seq dataset of 162,062 single T cells from 31 tissue regions, including tumor, adjacent normal tissues, and lymph nodes (LN), from three patients who underwent resections for progressing lung cancers after immune checkpoint blockade (ICB). We found marked regional heterogeneity in tumor persistence that was associated with heterogeneity in CD4 and CD8 T cell phenotypes; regions with persistent cancer cells were enriched for follicular helper CD4 T cells (TFH), regulatory T cells (Treg), and exhausted CD8 T cells. Clonal analysis demonstrated that highly-expanded T cell clones were predominantly of the CD8 subtype, were ubiquitously present across all sampled regions, found in the peripheral circulation, and expressed gene signatures of ‘large’ and ‘dual-expanded’ clones that have been predictive of response to ICB. Longitudinal tracking of CD8 T cell clones in the peripheral blood revealed that the persistence of ubiquitous CD8 T cell clones, as well as phenotypically distinct clones with tumor-reactive features, correlated with systemic tumor control. Finally, tracking CD8 T cell clones across tissues revealed the presence of TCF-1+precursor exhausted CD8 T cells in tumor draining LNs that were clonally linked to expanded exhausted CD8 T cells in tumors. Altogether, this comprehensive scTCR/RNA-seq dataset with regional, longitudinal, and clonal resolution provides fundamental insights into the tissue distribution, persistence, and differentiation trajectories of ICB-responsive T cells that underlie clinical responses to ICB.
Gene Expression Analysis of Glioma Neural Stem Cells Shows Disturbed Amino Acids Metabolism and Axonal Growth Cone Dynamics in Glioblastoma Multiforme
Authors: Vaja Rutvi
Published: September 2021

Glioblastoma multiforme(GBM) is a group of fatal and aggressive tumors of the central nervous system. Despite advancements in the treatment of GBM, patients diagnosed with these tumors typically have a poor prognosis and poor quality of life as the disease develops. The single-cell RNA high-throughput sequencing processed data for Glioma cancer stem cells were taken from GEO and analyzed to find out the underlying expression differences at the gene level between glioma neural stem cells(GSCs) and Normal neural stem cells(NSCs). In the current study, we have performed an RNA-sequencing analysis between GSCs and NSCs to better understand the origin of GBM. We have performed bioinformatics analysis on the transcriptional profile of 134 samples which consisted of 75 GSCs and 59 NSCs obtained from the NCBI bio project(PRJNA546254). First, an exploratory analysis was performed which showed significant variation patterns between GSCs and NSCs. Subsequently, Deseq2 differential gene expression analysis identified 1436 differentially expressed genes between GSCs and NSCs[(padj. value <0.05, log2 fold change (>=+/-1.5)]. This study reveals genes like MAOA, MAOB, GATM, GLDC, AMT, and SHMT1 as the key features contributing to the disturbed processes of Glycine, threonine, and serine amino acid metabolism, axonal cone growth curve, and cell migration in Glioma. Conclusively, our study also depicts gene expression changes in amyloid beta-binding protein in between GSCs and NSCs which plays an important role in tumor microenvironment formation. Besides, the results presented here reveal new insight into the progression of GBM and the identification of novel genes involved in gliomagenesis.
Expert-integrated automated machine learning uncovers hemodynamic predictors in spinal cord injury
Authors: Chou Austin , Torres-Espin Abel , Kyritsis Nikos , Huie J. Russell , Khatry Sarah , Funk Jeremy , Hay Jennifer , Lofgreen Andrew , Shah Rajiv , McCann Chandler , Pascual Lisa U , Amorim Edilberto , Weinstein Philip R , Manley Geoffrey T , Dhall Sanjay S , Pan Jonathan Z , Bresnahan Jacqueline C , Beattie Michael S , Whetstone William D , Ferguson Adam R ,   the TRACK-SCI Investigators
Published: September 2021

Automated machine learning (AutoML) is positioned to democratize artificial intelligence (AI) by reducing the amount of human input and ML expertise needed to create prediction models. However, successful translation of ML in biomedicine requires moving beyond optimizing only for prediction accuracy and towards discovering reproducible clinical and biological inferences. Here, we present a model-agnostic framework to reinforce AutoML using strategies and tools of explainable and reproducible AI, including novel metrics for performance precision and feature instability. The framework enables clinicians to interpret AutoML-generated models for clinical and biological verifiability and consequently integrate domain expertise during model development. We applied the framework towards spinal cord injury prognostication and identified a detrimental relationship between intraoperative hypertension and patient outcome. Furthermore, our analysis captured evolving clinical practices such as faster time-to-surgery and blood pressure management that affected clinical model validation. Altogether, we illustrate how augmenting AutoML for inferential reproducibility empowers biomedical discovery and builds trust in AI processes towards effective clinical integration.
Plant-based biosensors for detecting CRISPR-mediated genome engineering
Authors: Yuan Guoliang , Hassan Md. Mahmudul , Yao Tao , Lu Haiwei , Vergara Michael Melesse , Labbé Jesse L. , Muchero Wellington , Pan Changtian , Chen Jin-Gui , Tuskan Gerald A. , Qi Yiping , Abraham Paul E. , Yang Xiaohan
Published: September 2021

CRISPR/Cas has recently emerged as the most reliable system for genome engineering in various species. However, concerns about risks associated with CRISPR/Cas9 technology are increasing on potential unintended DNA changes that might accidentally arise from CRISPR gene editing. Developing a system that can detect and report the presence of active CRIPSR/Cas tools in biological systems is therefore very necessary. Here, we developed the real-time detection systems that can spontaneously indicate CRISPR-Cas tools for genome editing and gene regulation including CRISPR/Cas9 nuclease, base editing, prime editing and CRISPRa in plants. Using the fluorescence-based molecular biosensors, we demonstrated that the activities of CRISPR/Cas9 nuclease, base editing, prime editing and CRIPSRa can be effectively detected in transient expression via protoplast transformation and leaf infiltration (in Arabidopsis, poplar, and tobacco) and stable transformation in Arabidopsis.
A serotonergic axon-cilium synapse drives nuclear signaling to maintain chromatin accessibility
Authors: Sheu Shu-Hsien , Upadhyayula Srigokul , Dupuy Vincent , Pang Song , Lemire Andrew L. , Walpita Deepika , Pasolli H. Amalia , Deng Fei , Wan Jinxia , Wang Lihua , Houser Justin , Sanchez-Martinez Silvia , Brauchi Sebastian E. , Banala Sambashiva , Freeman Melanie , Xu C. Shan , Kirchhausen Tom , Hess Harald F. , Lavis Luke , Li Yu-Long , Chaumont-Dubel Séverine , Clapham David E.
Published: September 2021

Chemical synapses between axons and dendrites mediate much of the brain’s intercellular communication. Here we describe a new kind of synapse – the axo-ciliary synapse - between axons and primary cilia. By employing enhanced focused ion beam – scanning electron microscopy on samples with optimally preserved ultrastructure, we discovered synapses between the serotonergic axons arising from the brainstem, and the primary cilia of hippocampal CA1 pyramidal neurons. Functionally, these cilia are enriched in a ciliary-restricted serotonin receptor, 5-hydroxytryptamine receptor 6 (HTR6), whose mutation is associated with learning and memory defects. Using a newly developed cilia-targeted serotonin sensor, we show that optogenetic stimulation of serotonergic axons results in serotonin release onto cilia. Ciliary HTR6 stimulation activates a non-canonical Gαq/11-RhoA pathway. Ablation of this pathway results in nuclear actin and chromatin accessibility changes in CA1 pyramidal neurons. Axo-ciliary synapses serve as a distinct mechanism for neuromodulators to program neuron transcription through privileged access to the nuclear compartment.
The exocyst complex and intracellular vesicles mediate soluble protein trafficking to the primary cilium
Authors: Niedziółka SM , Datta S , Uśpieński T , Baran B , Skarżyńska W , Humke EW , Rohatgi R , Niewiadomski P
Published: September 2021

The efficient transport of proteins into the primary cilium is a crucial step for many signaling pathways. Dysfunction of this process can lead to the disruption of signaling cascades or cilium assembly, resulting in developmental disorders and cancer. Previous studies on the protein delivery to the cilium were mostly focused on the membrane-embedded receptors. In contrast, how soluble proteins are delivered into the cilium is poorly understood. In our work, we identify the exocyst complex as a key player in the ciliary trafficking of soluble Gli transcription factors. In line with the known function of the exocyst in intracellular vesicle transport, we demonstrate that soluble proteins, including Gli2/3 and Lkb1, can use the endosome recycling machinery for their delivery to the primary cilium. Finally, we identify GTPases: Rab14, Rab18, Rab23, and Arf4 that are involved in vesicle-mediated Gli protein ciliary trafficking. Our data pave the way for a better understanding of ciliary transport and uncover novel transport mechanisms inside the cell.
Towards optimization of oscillatory stimulation during sleep
Authors: Ladenbauer Julia , Khakimova Liliia , Malinowski Robert , Obst Daniela , Tönnies Eric , Antonenko Daria , Obermayer Klaus , Hanna Jeff , Flöel Agnes
Published: September 2021

Background

Oscillatory rhythms during sleep such as slow oscillations (SO) and spindles, and most importantly their coupling, are thought to underlie processes of memory consolidation. External slow oscillatory transcranial direct current stimulation (so-tDCS) with a frequency of 0.75 Hz has been shown to improve this coupling and memory consolidation, however, effects varied quite markedly between individuals, studies, and species. Here, we aimed to determine how precisely the frequency of stimulation has to match the naturally occurring SO frequency in individuals to optimally improve SO-spindle coupling. Moreover, we systematically tested stimulation durations necessary to induce changes.

Methods

We addressed these questions by comparing so-tDCS with individualized frequency to standardized frequency of 0.75 Hz in a within-subject design with 28 older participants during napping while systematically varying stimulation train durations between 30 s, 2 min, and 5 min.

Results

Stimulation trains as short as 30 s were sufficient to modulate the coupling between SOs and spindle activity. Contrary to our expectations, so-tDCS with standardized frequency indicated stronger aftereffects with regard to SO-spindle coupling compared to individualized frequency. Angle and variance of spindle maxima occurrence during the SO cycle were similarly modulated.

Conclusion

In sum, short stimulation trains were sufficient to induce significant changes in sleep physiology allowing for more trains of stimulation, which provides methodological advantages and possibly even larger behavioral effects in future studies. With regard to individualized stimulation frequency, further options of optimization need to be investigated, such as closed-loop stimulation to calibrate stimulation frequency to the SO frequency at time of stimulation onset.

Significance statement

Application of slow oscillatory transcranial direct current stimulation during sleep has been shown to enhance specific memory-relevant sleep parameters and memory performance after sleep, albeit with a high degree of variability. Here, we systematically explored two major stimulation parameters possibly accounting for this variability in humans: frequency and duration of stimulation. We found, contrary to our expectations, standardized frequency stimulation with 0.75 Hz being superior to individualized frequency stimulation in enhancing specific sleep parameters. Moreover short stimulation trains of 30 seconds were as effective as 5 min in modulating aftereffects. These are encouraging findings, implying methodological advantages as larger quantity of aftereffects data can be obtained within the same time window, which may also lead to enhanced behavioral stimulation effects.
HIV-1 hijacks the cell extracellular matrix to spread collectively and efficiently between T lymphocytes
Authors: Inizan C. , Caillet M. , Desrames A. , David A. , Bomme P. , Mallet A. , Bourdic K. , Lambotte O. , Boufassa F. , Mesel-Lemoine M. , Tangy F. , Pancino G. , Mouquet H. , Lagache T. , Dufour A. , Olivo-Marin J.- C. , Rey F.A. , Saez-Cirion A. , Thoulouze M.-I.
Published: September 2021

Collective transmissionviastructures containing several virions has recently emerged as a highly efficient mode of viral spread. Here, we demonstrate that HIV-1 spreads between T lymphocytes in the form of viral particles colonies that are concentrated and sheltered in an extracellular matrix (ECM) lattice enabling their collective transmission upon cell contacts. Intrinsically, ECM-clustered viruses infect T lymphocytes more efficiently than individual viral particles. They preserve HIV-1 transmission from antiretroviral treatment (ArT) and potent broadly neutralizing antibodies. We also show that collagen induced by HIV-1 infection controls the clustering of virions and their collective spread, thereby enhancing infectivity. CD4+ T cells from HIV-1-infected patients produce and transmit ECM-virus clusters, supporting that they could be involvedin vivo. This study provides new insights into modes of HIV-1 transmission and identifies a novel fundamental role for collagen in this process. HIV-1 spreadviaECM-virus clusters may have important implications for viral dissemination and persistence, including during therapy.
Quantitative phosphoproteomics reveals regulatory state dependent efficacy of chemical Cdk1 inhibition and distinct Cdk5 complexes as novel Wee1 substrates
Authors: Grassetti Andrew V. , Hards Rufus , Gerber Scott A.
Published: September 2021

Wee1 kinase plays a central role in the eukaryotic cell cycle via its well-known negative regulation of Cdk1 activity at the G2/M transition, preventing progression into mitosis until DNA replication and/or DNA damage repair is complete. Recent genetic evidence in yeast, flies and human cells have suggested additional functions of Wee1 in mitosis and during mitotic exit, respectively. To discover new candidate substrates of Wee1 kinase, we used SILAC-based phosphoproteomics and selective chemical inhibition to quantitatively compare phosphorylation site abundances in the presence and absence of Wee1 activity. Unexpectedly, we uncovered a role for the Wee1-dependent phosphorylation of Cdk1-cyclin B at tyrosine 15 (Y15) in facilitating chemical inhibition of Cdk1-cyclin B by the inhibitor RO3306. Thermal shift stability assays demonstrated greater binding affinity of RO3306 for Y15-phosphorylated Cdk1-cyclin B versus unphosphorylated complex, providing an additional molecular basis for the observed Wee1 inhibitor-based toxicity in human cells. In addition, our experiments identified Cdk5-CABLES and Cdk5-cyclin B as novel substrates of Wee1 during chemically induced exit from mitosis. Collectively, these experiments facilitate a greater understanding of the Wee1-Cdk1 signaling axis and uncover new candidate substrates for Wee1.
The Clock:cycle complex is a major transcriptional regulator ofDrosophilaphotoreceptors that protects the eye from retinal degeneration and oxidative stress
Authors: Jauregui-Lozano Juan , Hall Hana , Stanhope Sarah C. , Bakhle Kimaya , Marlin Makayla M. , Weake Vikki M.
Published: September 2021

The aging eye experiences physiological changes that include decreased visual function and increased risk of retinal degeneration. Although there are transcriptomic signatures in the aging retina that correlate with these physiological changes, the gene regulatory mechanisms that contribute to cellular homeostasis during aging remain to be determined. Here, we integrated ATAC-seq and RNA-seq data to identify 61 transcription factors that showed differential activity in agingDrosophilaphotoreceptors. These 61 age-regulated transcription factors include two circadian regulators, Clock and cycle, that showed sustained increases in activity during aging. When we disrupted Clock activity in adult photoreceptors, we observed changes in expression of 15 – 20% of genes including key components of the phototransduction machinery and many eye-specific transcription factors. Using ATAC-seq, we showed that loss of Clock activity leads to changes in activity of 31 transcription factors and causes a progressive decrease in global levels of chromatin accessibility in photoreceptors. Supporting a key role for Clock-dependent transcription in the eye, disruption of Clock activity in photoreceptors also induced light-dependent retinal degeneration and increased oxidative stress, independent of light exposure. Together, our data suggests that the circadian regulators Clock and cycle act as neuroprotective factors in the aging eye by directing gene regulatory networks that maintain expression of the phototransduction machinery and counteract oxidative stress.
An Attempt to Develop an Aptamer Lateral-Flow Assay (ALFA) for Easy, Rapid, and Sensitive Detection of Lethal Mushroom Toxin α-amanitin
Authors: Chen Qingchuan , Fan Chen , Huang Haozhe , Xu Binglin , Zong Yeqing
Published: September 2021

Amatoxins contribute to the majority of mushroom poisoning, most prominently, α-amanitin. Since mushroom is a common foodstuff worldwide, an easy, rapid, sensitive test for α-amanitin is needed. Several detection methods for α-amanitin have been developed, including HPLC, LC-MS, and ELISA, and LFIA. Aptamers have several advantages compared to antibodies: easy development via SELEX, longer shelf life, and higher temperature- and pH-tolerance. Aptamer Lateral Flow Assay (ALFA) is a similar technology compared to LFIA but incorporates aptamers as target-recognizing agents. This study attempted to develop an ALFA test strip for α-amanitin using a previously-developed aptamer, however failure of generating a colorimetric readout at the test line is persisted throughout all experiments, even though the concept is fully-proved and the control line functions normally. The failure is attributed to the small size of the molecule, leading to immobilization difficulties on the nitrocellulose membrane to form the test line, and the hindering of effective “surround” mechanism of aptamer-target binding (instead of “adhere”, when the target molecule is large, e.g. a protein). It is concluded that ALFAs for small-molecules whose aptamer-target interaction has not yet been studied and modelled in detail remains a challenge, despite ALFAs’ large potential.
Reconstructing Sample-Specific Networks using LIONESS
Authors: Kuijjer Marieke L. , Glass Kimberly
Published: September 2021

We recently developed LIONESS, a method to estimate sample-specific networks based on the output of an aggregate network reconstruction approach. In this manuscript, we describe how to apply LIONESS to different network reconstruction algorithms and data types. We highlight how decisions related to data preprocessing may affect the output networks, discuss expected outcomes, and give examples of how to analyze and compare single sample networks.
Ecological stochasticity and phage induction diversify bacterioplankton communities at the microscale
Authors: Szabo Rachel E. , Pontrelli Sammy , Grilli Jacopo , Schwartzman Julia A. , Pollak Shaul , Sauer Uwe , Cordero Otto X.
Published: September 2021

In many natural environments, microorganisms self-assemble around heterogeneously distributed resource patches. The growth and collapse of populations on resource patches can unfold within spatial ranges of a few hundred micrometers or less, making such microscale ecosystems hotspots of biological interactions and nutrient fluxes. Despite the potential importance of patch-level dynamics for the large-scale evolution and function of microbial communities, we have not yet been able to delineate the ecological processes that control natural populations at the microscale. Here, we addressed this challenge in the context of microbially-mediated degradation of particulate organic matter by characterizing the natural marine communities that assembled on over one thousand individual microscale chitin particles. Through shotgun metagenomics, we found significant variation in microscale community composition despite the similarity in initial species pools across replicates. Strikingly, a subset of particles was highly populated by rare chitin-degrading strains; we hypothesized that their conditional success reflected the impact of stochastic colonization and growth on community assembly. In contrast to the conserved functional structures that emerge in ecosystems at larger scales, this taxonomic variability translated to a wide range of predicted chitinolytic abilities and growth returns at the level of individual particles. We found that predation by temperate bacteriophages, especially of degrader strains, was a significant contributor to the variability in the bacterial compositions and yields observed across communities. Our study suggests that initial stochasticity in assembly states at the microscale, amplified through biotic interactions, may have significant consequences for the diversity and functionality of microbial communities at larger scales.

Significance Statement

The biogeochemical consequences of the degradation of particulate organic matter by microorganisms represent the cumulative effect of microbial activity on individual microscale resource patches. The ecological processes controlling community dynamics in these highly localized microenvironments remain poorly understood. Here, we find that complex marine communities growing on microscale resource particles diverge both taxonomically and functionally despite assembling under identical abiotic conditions from a common species pool. We show that this variability stems from bacteriophage predation and history-dependent factors in community assembly, which create stochastic dynamics that are spatially structured at the microscale. This microscale stochasticity may have significant consequences for the coexistence, evolution, and function of diverse bacterial and viral populations in the global ocean.
Modeling tissue-specific breakpoint proximity of structural variations from 2,382 whole-genomes to identify cancer drivers
Authors: Martinez-Fundichely Alexander , Dixon Austin , Khurana Ekta
Published: September 2021

Structural variations (SVs) in cancer cells often impact large genomic regions with functional consequences. However, little is known about the genomic features related to the breakpoint distribution of SVs in different cancers, a prerequisite to distinguish loci under positive selection from those with neutral evolution. We developed a method that uses a generalized additive model to investigate the breakpoint proximity curves from 2,382 whole-genomes of 32 cancer types. We find that a multivariate model, which includes linear and nonlinear partial contributions of various tissue-specific features and their interaction terms, can explain up to 57% of the observed deviance of breakpoint proximity. In particular, three-dimensional genomic features such as topologically associating domains (TADs), TAD-boundaries and their interaction with other features show significant contributions. The model is validated by identification of known cancer genes and revealed putative drivers in novel cancers that have previous evidence of therapeutic relevance in other cancers.
Horn trophy seized in illegal wildlife trade assigned to swamp buffalo using a combined morphometric and DNA based approach in wildlife forensics in India
Authors: Vipin  , Sharma Vinita , Sharma Chandra Prakash , Goyal Surendra Prakash , Gupta Sandeep Kumar
Published: September 2021

The illegal wildlife trade has threatened the existence of many extant wild animal species throughout the world. While dealing with the illegal wildlife trade of horns, we face problems of not having a proper protocol and lack of reference database to assign the species for proper implementation of wildlife laws. In one such condition, a horn trophy suspected to be of a wild buffalo was seized by authorities and sent to us for species identification. We used a combined approach of morphological and DNA analysis to ascertain the seized horn’s species. The two measurements, circumference at the base (CAB) and length on the front curve (LOFC) were measured for the seized and other horns of different bovid species, showing morphological resemblance with the seized horn. The 3-D scatter plot, generated by the values of CAB, LOFC and CAB/ LOFC, differentiated the different bovid species into distinct clusters and placed the seized horn in the proximity of domestic buffaloes. The Bayesian evolutionary analysis of the partial D-loop gene (521bp) placed the seized horn in a clade with swamp buffaloes. Since swamp buffaloes are domestic buffaloes, both these approaches concluded the same results. Hence, the current protocol developed may also be used to differentiate among wild buffalo, domestic buffalo, Cattle, Wild yak, Gaur and Takin using a combined approach of morphometric and DNA-based analysis, which may be used to deal with illegal wildlife trade of different bovid species at the world level.
A new sugar for an old phage: A c-di-GMP dependent polysaccharide pathway sensitizes E. coli for bacteriophage infection
Authors: Sellner Benjamin , Prakapaitė Rūta , van Berkum Margo , Heinemann Matthias , Harms Alexander , Jenal Urs
Published: September 2021

Bacteriophages are ubiquitous parasites of bacteria and major drivers of bacterial ecology and evolution. Despite an ever-growing interest in their biotechnological and therapeutic applications, detailed knowledge of the molecular mechanisms underlying phage-host interactions remains scarce. Here, we show that bacteriophage N4 exploits a novel surface glycan, NGR, as a receptor to infect its host Escherichia coli. We demonstrate that this process is regulated by the second messenger c-di-GMP and that N4 infection is specifically stimulated by the diguanylate cyclase DgcJ while the phosphodiesterase PdeL effectively protects E. coli from N4-mediated killing. PdeL-mediated protection requires its catalytic activity to reduce c-di-GMP and includes a secondary role as a transcriptional repressor. We demonstrate that PdeL binds to and represses the promoter of the wec operon, which encodes components of the ECA exopolysaccharide pathway. However, only the acetylglucosamine epimerase WecB but none of the other ECA components is required for N4 infection. Based on this, we postulate that NGR is an N-acetylmannosamine-based carbohydrate polymer that is produced and exported to the cell surface of E. coli in a c-di-GMP dependent manner where it serves as a receptor for N4. This novel carbohydrate pathway is conserved in E. coli and other bacterial pathogens, serves as the primary receptor for a range of N4-like bacteriophages, and is induced at elevated temperature and by specific amino acid-based nutrients. These studies provide an entry point into understanding how bacteria use specific regulatory mechanisms to balance costs and benefits of highly conserved surface structures.

Importance

Because bacterial surface glycans are in direct contact with the environment they can provide essential protective functions during infections or against competing bacteria. But such structures are also ‘Achilles heels’ as they can serve as primary receptors for bacteriophages. Bacteria thus need to carefully control the exposure of conserved surface glycans to balance costs and benefits. Here, we identify a novel exopolysaccharide that is widely conserved in E. coli and is used by N4 and related bacteriophages as primary receptor. We demonstrate that the synthesis of NGR (N4 glycan receptor) is tightly controlled by the second messenger c-di-GMP in a highly specific manner and by a single diguanylate cyclase. These studies provide an example of how bacteria can alleviate the strong selective pressure imposed on them by bacteriophages entering through conserved surface structures by carefully regulating their synthesis and secretion.
BMP7 increases UCP1-dependent and independent thermogenesis with a unique gene expression program in human neck area derived adipocytes
Authors: Shaw Abhirup , Tóth Beáta B , Arianti Rini , Csomós István , Póliska Szilárd , Vámos Attila , Bacso Zsolt , Győry Ferenc , Fésüs László , Kristóf Endre
Published: September 2021

White adipocytes contribute to energy storage accumulating lipid droplets, whereas brown and beige adipocytes mainly function in dissipating energy as heat primarily via the action of uncoupling protein 1 (UCP1). Bone morphogenic protein 7 (BMP7) has been shown to drive brown adipocyte differentiation in mice. In this study, we have performed global RNA-sequencing and functional assays on adipocytes obtained from subcutaneous (SC) and deep-neck (DN) depots of human neck and differentiated with or without BMP7. We found that BMP7 did not influence differentiation but upregulated browning markers, including UCP1 mRNA and protein. BMP7 also enhanced mitochondrial DNA content, fragmentation, and levels of oxidative phosphorylation complex subunits along with PGC1α and p-CREB upregulation. Furthermore, both UCP1-dependent proton leak and UCP1-independent, creatine driven substrate cycle coupled thermogenesis were augmented upon BMP7 treatment in SC and DN derived adipocytes. The gene expression analysis shed light also on possible role of genes unrelated to thermogenesis so far, including ACAN, CRYAB, and ID1, which were amongst the highest upregulated ones by BMP7 treatment in both types of adipocytes. Together, our study shows that BMP7 strongly upregulates thermogenesis in human neck area derived adipocytes, along with genes, which might have a supporting role in energy expenditure.
Small extracellular vesicles from iPSC-MSC lose their regenerative potential upon UV-C irradiation
Authors: Biani M.C. , Lombardi A. , Norris A. , Bucci P. , La Greca A. , Waisman A. , Moro L.N. , Sevlever G. , Montanari J. , Miriuka S. , Luzzani C.
Published: September 2021

Mesenchymal Stem Cells derived from induced Pluripotent Stem cells (iPSC-MSC) have become a promising alternative to classical Mesenchymal Stem Cells in regenerative medicine. Their properties -as immunomodulatory and regenerative capacities-are in part due to the secretion of Extracellular Vesicles (EVs). Small EVs (sEVs) with sizes that range from 50 to 120 nm contain proteins, lipids, and nucleic acids that exert a role in cellular communication. Their content will depend on the cell of origin and its physiological state, thus the message they convey might change in response to changes in cellular conditions. In particular, the DNA damage response (DDR) has been reported to modulate sEVs secretion. In this work, we analyze how UV-C radiation upon iPSC-MSC alter sEVs secretion, cargo and bystander effect. Here, we confirm that UV-C radiation causes DDR in a dose dependent manner. In addition, we found that UV-C induced stress did not modulate the expression of genes that participate in sEVs biogenesis pathway. Consequently, we found that the amount of sEVs secreted by radiated and non-irradiated cells remained stable. However, sEVs from radiated cells were unable to promote cell migration in their target cells. Moreover, a label-free proteomic analysis revealed that UV-C induced DDR produces sEVs with an altered cargo, rich in migration-inhibiting proteins, and resulting in a less stromal-oriented repertoire.
The transcriptomic signature of cyclical parthenogenesis
Authors: Huynh Trung Viet , Hall Alexander S. , Xu Sen
Published: September 2021

Cyclical parthenogenesis, where females can engage in sexual or asexual reproduction depending on environmental conditions, represents a novel reproductive phenotype that emerged during eukaryotic evolution. The fact that environmental conditions can trigger cyclically parthenogens to engage in distinct reproductive modes strongly suggests that gene expression plays a key role in the origin of cyclical parthenogenesis. However, the genetic basis underlying cyclical parthenogenesis remains understudied. In this study we characterize the female transcriptomic signature of sexual vs. asexual reproduction in the cyclically parthenogenetic microcrustacean Daphnia pulex and D. pulicaria. Our analyses of differentially expressed genes, pathway enrichment, and GO term enrichment clearly show that compared to sexual reproduction the asexual reproductive stage is characterized by both the under-regulation of meiosis and cell-cycle genes and the up-regulation of metabolic genes. We suggest that the under-regulation of meiosis and cell-cycle genes is responsible for the origin of parthenogenesis from meiosis, whereas differentially expressed metabolic genes may mediate choice of asexual vs. sexual reproductive pathway. Furthermore, our analyses identify some cases of divergent expression among gene family members (e.g., doublesex, NOTCH2) associated with asexual or sexual reproductive stage, suggesting potential functional divergence among gene family members.
Temporal Changes in Transcripts of MITE Transposable Elements during Rice Endosperm Development
Authors: Nagata Hiroki , Ono Akemi , Tonosaki Kaoru , Kawakatsu Taiji , Yano Kentaro , Kishima Yuji , Kinoshita Tetsu
Published: September 2021

The repression of transcription from transposable elements (TEs) by DNA methylation is necessary to maintain genome integrity and prevent harmful mutations. However, under certain circumstances, TEs are thought to escape from the host defense system and reactivate their transcription. In Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa), DNA demethylases target the sequences derived from TEs in the central cell, the progenitor cell for the endosperm in the female gametophyte. This genome-wide DNA demethylation is also observed in the endosperm after fertilization. In this study, we used a custom microarray to survey the transcripts generated from TEs during the rice endosperm development and at selected timepoints in the embryo as a control. The expression patterns of TE transcripts are dynamically up- and downregulated during endosperm development, especially for miniature inverted-repeat transposable elements (MITEs). Surprisingly, some TE transcripts were directionally controlled, while the other DNA transposons and retrotransposons were not. We also discovered the NF-Y binding motif, CCAAT, in the region near the 5′ terminal inverted repeat of Youren, one of the transcribed MITEs in the endosperm. Our results uncover dynamic changes in TE activity during endosperm development in rice.
miPEP858/miR858-MYB3-PSK4 module regulates growth and development in Arabidopsis
Authors: Badola Poorwa Kamal , Sharma Ashish , Gautam Himanshi , Trivedi Prabodh Kumar
Published: September 2021

Small molecules, peptides and miRNAs, are the crucial regulators of plant growth. Here, we show the importance of cross-talk between miPEP858a/miR858a and Phytosulfokine (PSK4) in regulating plant growth and development in Arabidopsis. Genome-wide expression analysis suggested modulated expression of PSK4 in miR858 mutant and overexpression, miR858OX, plants. The silencing of PSK4 in miR858OX plants compromised the growth, whereas over-expression of PSK4 in miR858 mutant rescued the developmental defects. The exogenous application of synthetic PSK4 further complemented the plant development in mutant plants. Exogenous treatment of synthetic miPEP858a in PSK4 mutant led to clathrin-mediated internalization of the peptide however did not enhance growth as in the case of wild-type plants. We also demonstrate that the MYB3 is an important molecular component participating in miPEP858a/miR858a-PSK4 module. Finally, our work highlights the signalling between miR858/miPEP858-MYB3-PSK4 in modulating the expression of key elements involved in auxin responses leading to the regulation of growth.

One-sentence summary

Signaling network between small molecules, miPEP858a/miR858a and phytosulfokine, regulates plant growth in Arabidopsis.
Distinct cell adhesion signature defines glioblastoma myeloid-derived suppressor cell subsets
Authors: Bayik Defne , Bartels Cynthia F. , Lovrenert Katreya , Watson Dionysios C. , Zhang Duo , Kay Kristen , Lauko Adam , Johnson Sadie , Lo Alice , McGraw Mary , Grabowski Matthew , Mohammadi Alireza M. , Veglia Filippo , Fan Yi , Vogelbaum Michael A. , Scacheri Peter , Lathia Justin D.
Published: September 2021

Increased myeloid-derived suppressor cell (MDSC) frequency is associated with worse outcomes and poor therapeutic response in glioblastoma (GBM). Monocytic (m) MDSCs represent the predominant subset in the GBM microenvironment. However, the molecular basis of mMDSC enrichment in the tumor microenvironment compared to granulocytic (g) MDSCs has yet to be determined. Here, we report that mMDSCs and gMDSCs display differences in their tumoraccelerating ability, with mMDSCs driving tumor growth in GBM models. Epigenetic assessments indicate enhanced gene accessibility for cell adhesion programs in mMDSCs and higher cellsurface integrin expression in mouse and human mMDSCs. Integrin β1 blockage abrogated the tumor-promoting phenotype of mMDSCs and altered the immune profile in the tumor microenvironment. These findings suggest that integrin β1 expression underlies the enrichment of mMDSCs in tumors and represents a putative immunotherapy target to attenuate myeloid cell-driven immune suppression in GBM.

Myeloid-derived suppressor cells (MDSCs) drive glioblastoma growth; however, the function of specific MDSCs subsets is unclear. Bayik et al. demonstrate that adhesion programs are enhanced in monocytic MDSCs and responsible for their GBM-promoting function.
LORE receptor homomerization is required for 3-hydroxydecanoic acid-induced immune signaling and determines the natural variation of immunosensitivity within the Arabidopsis genus
Authors: Eschrig Sabine , Schäffer Milena , Shu Lin-Jie , Illig Tina , Eibel Sonja , Fernandez Atiara , Ranf Stefanie
Published: September 2021

The S-domain-type receptor-like kinase (SD-RLK) LIPOOLIGOSACCHARIDE-SPECIFIC REDUCED ELICITATION (LORE) fromArabidopsis thalianais a pattern recognition receptor that senses medium-chain 3-hydroxy fatty acids, such as 3-hydroxydecanoic acid (3-OH-C10:0), to activate pattern-triggered immunity. Here, we show that LORE homomerization is required to activate 3-OH-C10:0-induced immune signaling.

Fluorescence lifetime imaging inNicotiana benthamianademonstrated thatAtLORE homomerizes via the extracellular and transmembrane domains. Co-expression ofAtLORE truncations lacking the intracellular domain exerts a dominant negative effect onAtLORE signaling in bothN. benthamianaandA. thaliana, highlighting that homomerization is essential for signaling.

Screening for 3-OH-C10:0-induced reactive oxygen species production revealed natural variation within the Arabidopsis genus.Arabidopsis lyrataandArabidopsis hallerido not respond to 3-OH-C10:0, although both possess a putative LORE orthologue. Both LORE orthologues have defective extracellular domains that bind 3-OH-C10:0 to a similar level but lack the ability to homomerize. Thus, ligand binding is independent of LORE homomerization. Analysis ofAtLORE andAlyrLORE chimera suggests that the loss ofAlyrLORE homomerization is caused by several amino acid polymorphisms across the extracellular domain.

Our findings shed light on the activation mechanism of LORE and the loss of 3-OH-C10:0 perception within the Arabidopsis genus.
Temperature and nutrient conditions modify the effects of phenological shifts in predator-prey communities
Authors: Rudolf V.H.W.
Published: September 2021

While there is mounting evidence indicating that the relative timing of predator and prey phenologies shapes the outcome of trophic interactions, we still lack a comprehensive understanding of how important the environmental context (e.g. abiotic conditions) is for shaping this relationship. Environmental conditions not only frequently drive shifts in phenologies, but they can also affect the very same processes that mediate the effects of phenological shifts on species interactions. Thus, identifying how environmental conditions shape the effects of phenological shifts is key to predict community dynamics across a heterogenous landscape and how they will change with ongoing climate change in the future. Here I tested how environmental conditions shape effects of phenological shifts by experimentally manipulating temperature, nutrient availability, and relative phenologies in two predator-prey freshwater systems (mole salamander-bronze frog vs dragonfly larvae-leopard frog). This allowed me to (1) isolate the effect of phenological shifts and different environmental conditions, (2) determine how they interact, and (3) how consistent these patterns are across different species and environments. I found that delaying prey arrival dramatically increased predation rates, but these effects were contingent on environmental conditions and predator system. While both nutrient addition and warming significantly enhanced the effect of arrival time, their effect was qualitatively different: Nutrient addition enhanced the positive effect of early arrival while warming enhanced the negative effect of arriving late. Predator responses varied qualitatively across predator-prey systems. Only in the system with strong gape-limitation were predators (salamanders) significantly affected by prey arrival time and this effect varied with environmental context. Correlations between predator and prey demographic rates suggest that this was driven by shifts in initial predator-prey size ratios and a positive feedback between size-specific predation rates and predator growth rates. These results highlight the importance of accounting for temporal and spatial correlation of local environmental conditions and gape-limitation in predator-prey systems when predicting the effects of phenological shifts and climate change on predator-prey systems.
Deficiency of Irx5 protects mice from diet-induced obesity and associated metabolic abnormalities
Authors: Son Joe Eun , Kim Kyoung-Han , Hui Chi-chung
Published: September 2021

Obesity, a leading cause of several metabolic abnormalities, is mainly due to an imbalance of energy homeostasis. IRX3 and IRX5 have been suggested as determinants of obesity in connection with the intronic variants of FTO, the strongest genetic risk factor of polygenic obesity in humans. Although the causal effects of Irx3 on obesity and its related metabolic consequences have been demonstrated in vivo, the metabolic function of Irx5 remains unclear. In this study, using mice homozygous for an Irx5-knockout (Irx5KO) allele, we show a direct link between Irx5 expression and regulation of body mass/composition and energy homeostasis. Irx5KO mice are leaner and resistant to diet-induced obesity and associated metabolic abnormalities, primarily through the loss of adiposity with an increase in basal metabolic rate with adipose thermogenesis and lower food intake. Furthermore, our long-term feeding analysis found that Irx3 mutant mouse lines also have less food intake, indicating that lower caloric intake also contributes to their lean phenotype. Together, these results demonstrate that Irx5 is critical for energy homeostasis and regulation of body mass/composition and suggest that it likely acts in other tissues beyond adipocytes.
Convergent coding of recent and remote fear memory in the basolateral amygdala
Authors: Liu Jianfeng , Totty Michael S. , Melissari Laila , Bayer Hugo , Maren Stephen
Published: September 2021

Background

In both rodents and humans, the basolateral amygdala (BLA) is essential for encoding and retrieving conditioned fear memories. Although the BLA is a putative storage site for these memories, recent evidence suggests that they become independent of the BLA with the passage of time.

Methods

We systematically examined the role for the BLA in the retrieval of recent (1 day) and remote (2 weeks) fear memory using optogenetic, electrophysiological, and calcium imaging methods in male and female Long-Evans rats. Critically, we used a behavioral design that permits within-subjects comparison of recent and remote memory at the same time point; freezing behavior served as the index of learned fear.

Results

We found that BLA c-Fos expression was similar after the retrieval of recent or remote fear memories. Extracellular single-unit recordings in awake, behaving animals revealed that single BLA neurons exhibit robust increases in spike firing to both recent and remote conditional stimuli (CSs). Fiber photometry recordings revealed that these patterns of activity emerge from principal neurons. Consistent with these results, optogenetic inhibition of BLA principal neurons impaired conditioned freezing to both recent and remote CSs. There were no sex differences in any of the measures or manipulations.

Conclusions

These data reveal that BLA neurons encode both recent and remote fear memories, suggesting substantial overlap in the allocation of temporally distinct events. This may underlie the broad generalization of fear memories across both space and time. Ultimately, these results provide evidence that the BLA is a long-term storage site for emotional memories.