bluuebunny/crossref_metadata_2025_split huggingface dataset:
Single-cell Transcriptomic Analysis of Salivary Gland Endothelial Cells
Authors: Altrieth Amber L. , Suarez Emily , Nelson Deirdre A. , Gabunia Sergo , Larsen Melinda 
Published: June 2023

AbstractVascular endothelial cells have important functions in fibrosis via direct and indirect methods and in regeneration through secretion of tissue-specific, paracrine-acting angiocrine factors. In the salivary gland, endothelial cells are required for proper development, but their roles within adult glands are largely unknown. The goal of this work was to identify ligand-receptor interactions between endothelial cells and other cell types that are important during homeostasis, fibrosis, and regeneration. To model salivary gland fibrosis and regeneration, we utilized a reversible ductal ligation. To induce injury, a clip was applied to the primary ducts for 14 days, and to induce a regenerative response, the clip was subsequently removed for 5 days. To identify endothelial cell-produced factors, we used single-cell RNA-sequencing of stromal-enriched cells from adult submandibular and sublingual salivary glands. Transcriptional profiles of homeostatic salivary gland endothelial cells were compared to endothelial cells of other organs. Salivary gland endothelial cells were found to express unique genes and displayed the highest overlap in gene expression with other fenestrated endothelial cells from the colon, small intestine, and kidney. Comparison of the 14-day ligated, mock ligated, and 5-day deligated stromal-enriched transcripts and lineage tracing were used to identify evidence for a partial endoMT phenotype, which was observed in a small number of endothelial cell subsets with ligation. CellChat was used to predict changes in ligand-receptor interactions in response to ligation and deligation. CellChat predicted that after ligation, endothelial cells are sources of protein tyrosine phosphatase receptor type m, tumor necrosis factor ligand superfamily member 13, and myelin protein zero signaling and targets for tumor necrosis factor signaling. Following deligation, CellChat predicted that endothelial cells are sources of chemokine (C-X-C motif) and EPH signaling to promote regenerative responses. These studies will inform future endothelial cell-based regenerative therapies.
Thermosensitivity of translation underlies the mammalian nocturnal-diurnal switch
Authors: Beale Andrew D. , Rzechorzek Nina M. , Mihut Andrei , Zeng Aiwei , Smyllie Nicola J. , Pilorz Violetta , Richardson Rose , Bertlesen Mads F , James Nathan R. , Fazal Shaline V. , Voysey Zanna , Pelletier Jerry , Crosby Priya , Peak-Chew Sew Y , Lancaster Madeline A. , Hut Roelof A. , O’Neill John S. 
Published: June 2023

AbstractEarly mammals were nocturnal until the Cretaceous-Paleogene extinction facilitated their rapid expansion into daytime niches. Diurnality subsequently evolved multiple times, independently, but the mechanisms facilitating this switch are unknown. We found that physiological daily temperature shifts oppositely affect circadian clock rhythms in nocturnal versus diurnal mammals. This occurs through a cell-intrinsic signal inverter, mediated by global differences in protein phosphorylation, and effected at the level of bulk protein synthesis rates, with diurnal translation rate being less thermosensitive than nocturnal. Perturbations that reduce translational initiation or mTOR activity are sufficient to trigger the nocturnal-to-diurnal switch at the cellular, tissue, and organismal scale. Our results suggest a convergent selection pressure to attain diurnality by reducing the effect of temperature-dependent changes in protein synthesis on circadian clocks.One sentence summaryRecalibrating the thermosensitivity of protein synthesis drives daytime-selective activity in mammals.
Structural and functional analysis of vaccinia viral fusion complex component protein A28 through NMR and molecular dynamic simulations
Authors: Kao Chi-Fei , Tsai Min-Hsin , Carillo Kathleen Joyce , Tzou Der-Lii , Chang Wen 
Published: June 2023

AbstractHost cell entry of vaccinia virus (a poxvirus) proceeds through multiple steps that involve many viral proteins to mediate cell infection. Upon binding to cells, vaccinia virus membrane fuses with host membranes via a viral entry fusion protein complex comprising 11 proteins: A16, A21, A28, F9, G3, G9, H2, J5, L1, L5 and O3. Despite vaccinia virus having two infectious forms, mature and enveloped, that have different membrane layers, both forms require an identical viral entry fusion complex for membrane fusion. Components of the poxvirus entry fusion complex that have been structurally assessed to date share no known homology with all other type I, II and III viral fusion proteins, and the large number of fusion protein components renders it a unique system to investigate poxvirus-mediated membrane fusion. Here, we determined the NMR structure of a truncated version of vaccinia A28 protein. We also expressed a soluble H2 protein and showed that A28 interacts with H2 protein at a 1:1 ratioin vitro. Furthermore, we performed extensivein vitroalanine mutagenesis to identify A28 protein residues that are critical for H2 binding, entry fusion complex formation, and virus-mediated membrane fusion. Finally, we used molecular dynamic simulations to model full-length A28-H2 subcomplex in membranes. In summary, we characterized vaccinia virus A28 protein and determines residues important in its interaction with H2 protein and membrane components. We also provide a structural model of the A28-H2 protein interaction to illustrate how it forms a 1:1 subcomplex on a modeled membrane.
poloaffects cell fate upon ionizing radiation inDrosophilahematopoietic progenitors by negatively regulatinglok
Authors: Nguyen Tram Thi Ngoc , Chen Guang-Chao , Shim Jiwon , Song Young-Han 
Published: June 2023

AbstractIn response to ionizing radiation (IR), stem cells undergo cell cycle arrest, senescence, premature differentiation, or cell death. The decision between survival and death is critical during tumorigenesis and effective killing of cancer cells. We used the larvalDrosophilalymph gland, a hematopoietic organ, as a model to understand the mechanism for cell fate decisions during stem cell development. The hematopoietic progenitors survived or died via apoptosis when larvae were irradiated in early or late third instar larval (L3) stages, respectively. In late L3 progenitors, the basal level ofpolo(DrosophilaPLK1) was low, enabling IR-induced activation oflok(DrosophilaCHK2), which was necessary and sufficient for inducing autophagy and reactive oxygen species (ROS) production resulting in cell death. The high level ofpoloin early L3 progenitors negatively regulatedlokresulting in significantly low or undetectable levels of ROS or autophagy, respectively. The surviving early L3 progenitors underwent cell cycle arrest followed by premature differentiation affected bytefu(DrosophilaATM) andlokmutation. These results provide clues for designing effective therapeutic strategies for cancer.Summary statementWe elucidated the mechanism underlying cell fate decisions during stem cell development in larvalDrosophila, which will help develop effective cancer treatment modalities.
Boosting class II HLA epitope presentation to T cells with endoplasmic reticulum transmembrane domain fusion proteins
Authors: Pinamonti Veronica , Fuentes Ana Mellado , Schmid Theresa , Cetin Miray , Lerner Taga , Hernandez Miguel Hernandez , Lindner John M. 
Published: June 2023

AbstractAntigen processing and presentation are crucial for T-cell receptor engagement and potent adaptive immune responses. While establishing an antigen presentation system to identify TCR-epitope pairs from large libraries of genetically templated, ectopically expressed antigens, we observed a longitudinal suppression of antigen processing in B cells, which we call antigen processing attenuation (APA). Beyond its potential role in regulating B cell-enforced peripheral tolerance, APA represents a bottleneck to sustained antigen presentation in screening and vaccination approaches. To overcome APA we screened several pathways which might enhance processing. Notably, we identified several sequences derived from transmembrane domains (TM) of endoplasmic reticulum-resident proteins which promote sustained minigene-encoded antigen presentation. In particular, the first TM domain ofSYT6, a protein associated with synaptic vesicles, greatly enhances processing and ameliorates APA. These findings suggest a new entry point into antigen processing pathways, enabling the discovery of novel immune epitopes and improving T-cell based immunization strategies.
Endosymbiont diversity across native and invasive brown widow spider populations
Authors: Mowery Monica A. , Rosenwald Laura C. , Chapman Eric , Lubin Yael , Segoli Michal , Khoza Thembile , Lyle Robin , White Jennifer A. 
Published: June 2023

AbstractThe invasive brown widow spider,Latrodectus geometricus(Araneae: Theridiidae), has spread in multiple locations around the world and, along with it, brought associated organisms such as endosymbionts. We investigated endosymbiont diversity and prevalence across putative native and invasive populations of this spider, predicting lower endosymbiont diversity across the invasive range compared to the native range. First, we characterized the microbial community in the putative native (South Africa) and invasive (Israel and the United States) ranges via high throughput 16S sequencing of 103 adult females. All specimens were dominated by reads from only 1-3 amplicon sequence variants (ASV), and most individuals were infected with an apparently uniform strain ofRhabdochlamydia. We also foundRhabdochlamydiain spider eggs, indicating that it is a maternally-inherited endosymbiont. Relatively few other ASV were detected, but included two variantRhabdochlamydiastrains and severalWolbachia,Spiroplasmaand Enterobacteriaceae strains. We then diagnostically screened 118 adult female spiders from native and invasive populations specifically forRhabdochlamydiaandWolbachia.We foundRhabdochlamydiain 86% of individuals and represented in all populations, which suggests that it is a consistent and potentially important associate ofL. geometricus. Wolbachiawas found at lower overall prevalence (14%) and was represented in all countries, but not all populations. In addition, we found evidence for geographic variation in endosymbiont prevalence: spiders from Israel were more likely to carryRhabdochlamydiathan those from the US and South Africa, andWolbachiawas geographically clustered in both Israel and South Africa. Characterizing endosymbiont prevalence and diversity is a first step in understanding their function inside the host and may shed light on the process of spread and population variability in cosmopolitan invasive species.
Widespread gene regulator Psu inhibits transcription termination factor ρ by forced hyper-oligomerization
Authors: Gjorgjevikj Daniela , Kumar Naveen , Wang Bing , Hilal Tarek , Said Nelly , Loll Bernhard , Artsimovitch Irina , Sen Ranjan , Wahl Markus C. 
Published: June 2023

AbstractMany bacteriophages modulate the host transcription machinery for efficient expression of their own genomes. Phage P4 polarity suppression protein, Psu, is a building block of the viral capsid and inhibits the hexameric transcription termination factor, ρ, by presently unknown mechanisms. We elucidated cryogenic electron microscopy structures of ρ-Psu complexes, showing that Psu dimers laterally clamp two inactive, open ρ rings and promote their expansion to higher-oligomeric states. Systematic ATPase, nucleotide binding and nucleic acid binding studies revealed that Psu hinders ρ ring closure and traps nucleotides in their binding pockets on ρ. Structure-guided mutagenesis in combination with growth, pull-down and termination assays further delineated the functional ρ-Psu interfaces. Bioinformatic analyses suggested that, in addition to guarding its own genome against ρ, Psu enables expression of diverse phage-defense systems commonly found in P4-like mobile genetic elements across bacteria. Thus, Psu is a widespread gene regulator that inhibits ρviaforced hyper-oligomerization.
Structural Basis for Regulated Assembly of the Mitochondrial Fission GTPase Drp1
Authors: Rochon Kristy , Bauer Brianna L. , Roethler Nathaniel A. , Buckley Yuli , Su Chih-Chia , Huang Wei , Ramachandran Rajesh , Stoll Maria S. K. , Yu Edward W. , Taylor Derek J. , Mears Jason A. 
Published: June 2023

AbstractMitochondrial fission is crucial for distributing cellular energy throughout cells and for isolating damaged regions of the organelle that are targeted for degradation. This multistep process is initiated by the enhanced recruitment and oligomerization of dynamin-related protein 1 (Drp1) at the surface of mitochondria. As such, Drp1 is essential for inducing mitochondrial division in mammalian cells, and homologous proteins are found in all eukaryotes.De novomissense mutations in the Drp1 gene, DNM1L, are associated with severe neurodevelopmental diseases in patients, and no effective treatments are available. As a member of the dynamin superfamily of proteins (DSPs), controlled Drp1 self-assembly into large helical polymers stimulates its GTPase activity to promote membrane constriction. Still, little is known about the regulatory mechanisms that determine when and where Drp1 self-assembles, and proper mitochondrial dynamics requires correct spatial and temporal assembly of the fission machinery. Here we present a cryo-EM structure of a full-length, native Drp1 dimer in an auto-inhibited state. This dimer reveals two key conformational rearrangements that must be unlocked through intermolecular interactions to achieve the assembly competent state previously observed in crystal and filament structures. Specifically, the G domain is closed against the stalk domain and occludes intermolecular interactions necessary for self-assembly beyond a dimer. Similarly, adjacent stalks in the dimer form a more continuous interface that further occludes conserved intermolecular contact sites. This structural insight provides a novel mechanism for regulated self-assembly of the mitochondrial fission machinery.
RyR2 Binding of an Antiarrhythmic Cyclic Depsipeptide Mapped Using Confocal Fluorescence Lifetime Detection of FRET
Authors: Seflova Jaroslava , Schwarz Jacob A. , Smith Abigail N. , Svensson Bengt , Blackwell Daniel J. , Phillips Taylor A. , Nikolaienko Roman , Bovo Elisa , Rebbeck Robyn T. , Zima Aleksey V. , Thomas David D. , Petegem Filip Van , Knollmann Björn C. , Johnston Jeffrey N. , Robia Seth L. , Cornea Răzvan L. 
Published: June 2023

AbstractHyperactivity of cardiac sarcoplasmic reticulum (SR) ryanodine receptor (RyR2) Ca2+-release channels contributes to heart failure and arrhythmias. Reducing RyR2 activity, particularly during cardiac relaxation (diastole), is a desirable therapeutic goal. We previously reported that the unnatural enantiomer (ent) of an insect-RyR activator, verticilide, inhibits porcine and mouse RyR2 at diastolic (nanomolar) Ca2+and has in vivo efficacy against atrial and ventricular arrhythmia. To determine theent-verticilide structural mode of action on RyR2 and guide its further development via medicinal chemistry structure-activity relationship studies, here we used fluorescence lifetime (FLT)-measurements of Förster resonance energy transfer (FRET) in HEK293 cells expressing human RyR2. For these studies, we used an RyR-specific FRET molecular-toolkit and computational methods for trilateration (i.e., using distances to locate a point of interest). Multi-exponential analysis of FLT-FRET measurements between four donor-labeled FKBP12.6 variants and acceptor-labeledent-verticilide, yielded distance relationships placing the acceptor probe at two candidate loci within the RyR2 cryo-EM map. One locus is within the Ry12 domain (at the corner periphery of the RyR2 tetrameric complex). The other locus is sandwiched at the interface between helical domain 1 and the SPRY3 domain. These findings document RyR2-target engagement byent-verticilide, reveal new insight into the mechanism of action of this new class of RyR2-targeting drug candidate, and can serve as input in future computational determinations of theent-verticilide binding site on RyR2 that will inform structure-activity studies for lead optimization.
ER-Mitochondria Contact Sites expand during mitosis
Authors: Yu Fang , Courjaret Raphael , Elmi Asha , Hammad Ayat , Fisher Melanie , Terasaki Mark , Machaca Khaled 
Published: June 2023

AbstractMembrane contact sites between various organelles define specialized spatially defined signaling hubs, they are of great interest to better understand inter-organelle communication and its implications on cellular physiology. ER-mitochondria contact sites (ERMCS) are one of the best studied and mediate Ca2+signaling that regulates mitochondrial bioenergetics. However, little is known about ERMCS during mitosis. Here we show that ERMCS expand during mitosis using transmission electron microscopy, serial EM coupled to 3D reconstruction, and ERMCS markers. ERMCS expansion in mitosis is functionally significant as it is associated with enhanced Ca2+coupling between the ER and mitochondria resulting in heightened activation of mitochondrial dehydrogenases. Our data suggest that ERMCS remodeling in mitosis is important to meet the increased energy needs during cell division.
Cell-based assay to determine Type 3 Secretion System translocon assembly inPseudomonas aeruginosausing split luciferase
Authors: Guo Hanling , Geddes Emily , Opperman Timothy J. , Heuck Alejandro P. 
Published: June 2023

AbstractMultidrug resistantPseudomonas aeruginosaposes a serious threat to hospitalized patients. This organism expresses an arsenal of virulence factors that enables it to readily establish infections and to disseminate in the host. The Type III secretion System (T3SS) and its associated effectors play a crucial role in the pathogenesis ofP. aeruginosa,making them attractive targets for the development of novel therapeutic agents. The T3SS translocon, comprised of PopD and PopB, is an essential component of the T3SS secretion apparatus. In the properly assembled translocon, the N-terminus of PopD protrudes into the cytoplasm of the target mammalian cell, which can be exploited as a molecular indicator of functional translocon assembly. In this manuscript, we describe a novel whole-cell-based assay that employs the split NanoLuc luciferase detection system to provide a readout for translocon assembly.The assay demonstrates a favorable signal/noise ratio (17.9) and robustness (z’=0.73), making it highly suitable for high-throughput screening of small molecule inhibitors targeting T3SS translocon assembly.
Multifaceted roles of rice ABA/stress-induced intrinsically disordered proteins in augmenting drought resistance
Authors: Lin Meng-Chun , Tseng I-Chieh , Wang Ching-Lan , Hsiao Wen-Rong , Shih Yun-Jhih , Lin Wen-Dar , Yu Su-May , Ho Tuan-Hua David 
Published: June 2023

AbstractWater deficit stress causes devastating loss of crop yield worldwide. Improving crop drought resistance has become an urgent issue. Here we report that a group of abscisic acid (ABA)/drought stress-induced monocot-specific, intrinsically disordered, and highly proline-rich proteins, REPETITIVE PROLINE-RICH PROTEINS (RePRPs), play pivotal roles in drought resistance in rice seedlings. Rice ectopically expressingRePRPs outlived wild-type rice under extreme drought conditions primarily due to two underlying mechanisms. First,RePRPreduces water loss by decreasing stomata conductance in shoot. In addition,RePRPoverexpression enhances the levels of extracellular water barriers such as lignin and suberin, primarily in the root vascular bundle. Several groups of genes involved in lignin biosynthesis, especially the wall-bound peroxidase responsible for the final assembly of the lignin network, were induced byRePRP. Second, overexpression ofRePRPleads to lowered root osmotic potential. Root cell osmotic pressure was more negative in rice plants overexpressingRePRP2than wild-type plants, and the concentration of a key osmolyte, proline, was enhanced. Furthermore, the protein levels of two aquaporins that are important for drought stress tolerance were elevated. Hence, ABA/stress-inducedRePRPexpression leads to several beneficial traits of drought resistance, including lower water loss rate upon dehydration and higher root water use efficiency under drought conditions. This group of unique stress proteins may be an important target for technology development in enhancing drought stress resistance in cereals.
Contrasting genes conferring short and long-term biofilm adaptation inListeria
Authors: Monteith William , Pascoe Ben , Mourkas Evangelos , Clark Jack , Hakim Maliha , Hitchings Matthew D. , McCarthy Noel , Yahara Koji , Asakura Hiroshi , Sheppard Samuel K. 
Published: June 2023

AbstractListeria monocytogenesis an opportunistic food-borne bacterium that is capable of infecting humans with high rates of hospitalisation and mortality. Natural populations are genotypically and phenotypically variable, with some lineages being responsible for most human infections. The success ofL. monocytogenesis linked to its capacity to persist on food and in the environment. Biofilms are an important feature that allow these bacteria to persist and infect humans, therefore, understanding the genetic basis of biofilm formation is key to understanding transmission. We sought to investigate the biofilm forming ability ofL. monocytogenesby identifying genetic variation that underlies biofilm formation in natural populations using genome-wide association studies. Changes in gene expression of specific strains during biofilm formation were then investigated using RNAseq. Genetic variation associated with enhanced biofilm formation was identified in 273 genes by GWAS and differential expression in 220 genes by RNAseq. Statistical analyses show that number of overlapping genes flagged by either type of experiment is less than expected by random sampling. This is consistent with an evolutionary scenario where rapid adaptation is driven by variation in gene expression of pioneer genes, and this is followed by slower adaptation driven by nucleotide changes within the core genome.Impact statementListeria monocytogenesis a problematic food-borne bacterium that can cause severe illness and even death in humans. Some strains are known to be more common in disease and biofilms are crucial for survival in the environment and transmission to humans. To unravel the genetic basis of biofilm formation, we undertook a study employing genome-wide association studies (GWAS) and gene transcription profiling. We identified 273 genes associated with robust biofilm formation through GWAS and discovered differential expression in 220 genes through RNAseq. Statistical analysis revealed fewer overlapping genes than expected by chance, supporting an evolutionary scenario where initial adaptation relies on gene expression variation, followed by slower adaptation through genetic changes within the core genome.Data summaryShort read genome data are available from the NCBI (National Center for Biotechnology Information) SRA (Sequence Read Archive), associated with BioProject PRJNA971143 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA971143). Assembled genomes and supplementary material are available from FigShare: doi: 10.6084/m9.figshare.23148029. RNA sequence data and differential gene expression profiles have been deposited in the NCBI Gene Expression Omnibus.