bluuebunny/crossref_metadata_2025_split huggingface dataset:
DNA methylation machinery is involved in development and reproduction in the viviparous pea aphid (Acyrthosiphon pisum)
Authors: Yoon Kane , Williams Stephanie , Duncan Elizabeth J. 
Published: February 2024

AbstractThe pea aphid (Acyrthosiphon pisum) like the majority of extant aphids displays cyclical parthenogenesis - the ability of mothers to switch the reproductive mode of their offspring from reproducing parthenogenetically to sexually in response to environmental cues. The pea aphid genome encodes two paralogs of thede novoDNA methyltransferase gene,dnmt3aanddnmt3x. Here we show, using phylogenetic analysis, that this gene duplication event occurred at least 106 million years ago, likely after the divergence of the lineage leading to the Aphidomorpha (aphids, phylloxera and adelgids) from that leading to the scale insects (Coccoidea) and that the two paralogs are maintained in the genomes of all aphids examined. We also show that the mRNA of bothdnmt3paralogs are maternally expressed in the viviparous aphid ovary. During development both paralogs are expressed in the germ cells of embryos beginning at stage 5 and persisting throughout development. Chemical inhibition of the DNA methylation machinery leads to defects of oocytes and early stage embryos, and causes a proportion of later stage embryos to be born dead or die soon after birth. These phenotypes suggest a role for DNA methyltransferases in reproduction, consistent with that seen in other insects. Taking the vast evolutionary history of thednmt3paralogs, and the localization of their mRNAs in the ovary, we suggest there is a role fordnmt3aand/ordnmt3xin early development, and a role for DNA methylation machinery in reproduction and development of the viviparous pea aphid.
Bending DNA increases its helical repeat
Authors: Chandrasekhar Soumya , Swope Thomas P. , Fadaei Fatemeh , Hollis Daniel R. , Bricker Rachel , Houser Draven , Portman John J. , Schmidt Thorsten L. 
Published: February 2024

AbstractIn all biological systems, DNA is under high mechanical stress from bending and twisting. For example, DNA is tightly bent in nucleosome complexes, virus capsids, bacterial chromosomes, or complexes with transcription factors that regulate gene expression. A structurally and mechanically accurate model of DNA is therefore necessary to understand some of the most fundamental molecular mechanisms in biology including DNA packaging, replication, transcription and gene regulation. An iconic feature of DNA is its double helical nature with an average repeath0of ∼10.45 base pairs per turn, which is commonly believed to be independent of curvature. We developed a ligation assay on nicked DNA circles of variable curvature that reveals a strong unwinding of DNA to over 11 bp/turn for radii around 3-4 nm. Our work constitutes a major modification of the standard mechanical model of DNA and requires reassessing the molecular mechanisms and energetics of all processes involving tightly bent DNA.
Axolotl mandible regeneration occurs through mechanical gap closure and a shared regenerative program with the limb
Authors: Kramer Julia , Aires Rita , Keeley Sean D. , Schröder Tom Alexander , Lauer Günter , Sandoval-Guzmán Tatiana 
Published: February 2024

AbstractThe mandible plays an essential part in human life and, thus, defects in this structure can dramatically impair the quality of life in patients. Axolotls, unlike humans, are capable of regenerating their lower jaws; however, the underlying mechanisms and their similarity to those in limb regeneration are unknown. In this work, we used morphological, histological, and transcriptomic approaches to analyze the regeneration of lateral resection defects in the axolotl mandible. We found that this structure can regenerate all missing tissues in 90 days through gap minimization, blastema formation, and finally tissue growth, differentiation, and integration. Moreover, transcriptomic comparisons of regenerating mandibles and limbs showed that they share molecular phases of regeneration, that these similarities peak during blastema stages, and that mandible regeneration occurs at a slower pacing.Altogether, our study demonstrates the existence of a shared regenerative program used in two different regenerating body structures with different embryonic origins in the axolotl, and contributes to our understanding of the minimum requirements for a successful regeneration in vertebrates, bringing us closer to understand similar lesions in human mandibles.
Biophysics of microRNA-34a targeting and its influence on down-regulation
Authors: Sweetapple Lara , Kosek David M , Banijamali Elnaz , Becker Walter , Müller Juliane , Karadiakos Christina , Baronti Lorenzo , Guzzetti Ileana , Schritt Dimitri , Chen Alan , Andersson Emma R , Petzold Katja 
Published: February 2024

ABSTRACTmicroRNAs (miRNAs) regulate target mRNA expression post-transcriptionally through their association with Argonaute 2 (AGO2) proteins. Predicting the efficiency of mRNA repression by miRNA has been limited by our comprehension of the structure-function relationship within miRNA binding sites. Using a combination of EMSA, luciferase reporter assays, and structural probing, we investigated the interaction between the human tumour suppressormiR-34aand 12 mRNA targets. Comparison of direct RNA:RNA interactions and those within the functional AGO2 protein revealed that the isolated mRNA:miRNA duplex serves as a strong predictor for duplex affinity and structure within AGO2. Our findings reveal that AGO2 has a bidirectional capacity to modulate affinity; weakening tight RNA:RNA binders while strengthening weak ones. We identified three distinct structural groups that form uponmiR-34abinding and reveal a novel structural group that exhibits a guide strand bulge. MD simulations indicate a conceivable fit of this miRNA-bulge structure within AGO2. Our results demonstrate that the structural characteristics of mRNA:miRNA duplexes could serve as contributing determinants of repression efficacy.
LXA4 promotes the browning of white adipose through miR-133a-3p/SIRT1 pathway
Authors: Yu Dan , Ruan Yuan , Wang Yisu , Chen Xiaopei , Wang Dan , Wu Tianfeng 
Published: February 2024

AbstractLipoxin A4 (LXA4) promotes the browning of white adipose and energy consumption. The specific mechanism of which involved in white adipose browning is less clear. A high-fat diet (HFD) mouse model was constructed. Different groups of mice were treated with LXA4 accordingly. The body weight of mouse, subcutaneous and visceral fat, and food intake were recorded. The effect of LXA4 was examined by observing changes in pathology, serum insulin and lipid accumulation indices. The effects of LXA4/miR-133a-3p/Sirtuin1 on lipid droplet formation, fat browning-related genes, and the insulin receptor-AKT pathway in cells were examined after induction of adipocyte differentiation in 3T3-L1 precursors. At the cellular level, LXA4 promoted lipid droplet formation, expressions of fat browning genes and activation of the insulin receptor-AKT pathway in differentiated 3T3-L1 cells. MiR-133a-3p agomir partially offset the effects of LXA4. SIRT1 was a downstream target gene of miR-133a-3p, participating in the promotive effects of LXA4 on fat browning. LXA4 promotes white adipose browning and relieves insulin resistance through miR-133a-3p/SIRT1 pathway.
T cell interaction partners of DHHC20
Authors: Haxhiraj Deana , Kuropka Benno , Brencher Eva , Rosenberger David , Brandt Helena , Speck David , Scheerer Patrick , Brügger Britta , Clementi Cecilia , Freund Christian 
Published: February 2024

AbstractReversible palmitoylation of proteins at cysteine residues represents a post-translational modification that can alter the cellular localization of proteins, change their distribution within lipid membranes or modulate their conformation and molecular interaction patterns. DHHC enzymes catalyze protein palmitoylation, while thioesterases or hydrolases can rapidly remove the acyl-chain. In human T cells, DHHC proteins have been shown to modify proteins that are involved in major signaling pathways such as Ca2+-signaling or kinase-dependent activation of transcription factors for cytokines. For DHHC20, a role in the palmitoylation of the Orai1 Ca2+-channel has been demonstrated, but otherwise its T cell interaction partners are largely unknown. Here, we show that recombinantly expressed DHHC20 robustly interacts with 28 proteins from Jurkat T cells, as shown by affinity enrichment combined with mass spectrometric analysis. We find a robust interaction between DHHC20 and the β-subunit of the trimeric G protein Gαsβ1γ2, while the typically palmitoylated Ga subunit is not identified. Cross-linking mass spectrometry with purified DHHC20 and Gαsβ1γ2 then confirms a direct interaction between the β1γ2 domains and the enzyme, while rigid docking offers structural poses that are in agreement with the observed intermolecular cross-linking constraints. Thus, we suggest a model where the β1γ2 subunits of a trimeric G protein serve as a stable interaction partner of a DHHC enzyme, presumably acting as a landing platform for the Gα subunit that is subsequently palmitoylated by the enzyme.
Klumpy: A Tool to Evaluate the Integrity of Long-Read Genome Assemblies and Illusive Sequence Motifs
Authors: Madrigal Giovanni , Minhas Bushra Fazal , Catchen Julian 
Published: February 2024

AbstractThe improvement and decreasing costs of third-generation sequencing technologies has widened the scope of biological questions researchers can address with de novo genome assemblies. With the increasing number of reference genomes, validating their integrity with minimal overhead is vital for establishing confident results in their applications. Here, we present Klumpy, a tool for detecting and visualizing both misassembled regions in a genome assembly and genetic elements (e.g., genes, promotors, or transposable elements) of interest in a set of sequences. By leveraging the initial raw reads in combination with their respective genome assembly, we illustrate Klumpy’s utility by investigating antifreeze glycoprotein (afgp) loci across two icefishes, by searching for a reported absent gene in the northern snakehead fish, and by scanning the reference genomes of a mudskipper and bumblebee for misassembled regions. In the two former cases, we were able to provide support for the noncanonical placement of an afgp locus in the icefishes and locate the missing snakehead gene. Furthermore, our genome scans were able to identify an cryptic locus in the mudskipper reference genome, and identify a putative repetitive element shared amongst several species of bees.
Decoding functional proteome information in model organisms using protein language models
Authors: Barrios-Núñez Israel , Martínez-Redondo Gemma I. , Medina-Burgos Patricia , Cases Ildefonso , Fernández Rosa , Rojas Ana M. 
Published: February 2024

ABSTRACTProtein language models have been tested and proved to be reliable when used on curated datasets but have not yet been applied to full proteomes. Accordingly, we tested how two different machine learning based methods performed when decoding functional information from the proteomes of selected model organisms. We found that protein Language Models are more precise and informative than Deep Learning methods for all the species tested and across the three gene ontologies studied, and that they better recover functional information from transcriptomics experiments. The results obtained indicate that these Language Models are likely to be suitable for large scale annotation and downstream analyses, and we recommend a guide for their use.
TheBacillus subtilis yqgC-sodAoperon protects magnesium-dependent enzymes by supporting manganese efflux
Authors: Sachla Ankita J. , Soni Vijay , Piñeros Miguel , Luo Yuanchan , Im Janice J. , Rhee Kyu Y. , Helmann John D. 
Published: February 2024

AbstractMicrobes encounter a myriad of stresses during their life cycle. Dysregulation of metal ion homeostasis is increasingly recognized as a key factor in host-microbe interactions. Bacterial metal ion homeostasis is tightly regulated by dedicated metalloregulators that control uptake, sequestration, trafficking, and efflux. Here, we demonstrate that deletion of theBacillus subtilis yqgC-sodA(YS) complex operon, but not deletion of the individual genes, causes hypersensitivity to manganese (Mn). YqgC is an integral membrane protein of unknown function and SodA is a Mn-dependent superoxide dismutase (MnSOD). The YS strain has reduced expression of two Mn efflux proteins, MneP and MneS, consistent with the observed Mn sensitivity. The YS strain accumulated high levels of Mn, had increased reactive radical species (RRS), and had broad metabolic alterations that can be partially explained by the inhibition of Mg-dependent enzymes. Although the YS operon deletion strain and an efflux-deficientmneP mneSdouble mutant both accumulate Mn and have similar metabolic perturbations they also display phenotypic differences. Several mutations that suppressed Mn intoxication of themneP mneSefflux mutant did not benefit the YS mutant. Further, Mn intoxication in the YS mutant, but not themneP mneSstrain, was alleviated by expression of Mg-dependent, chorismate-utilizing enzymes of themenaquinone,siderophore, and tryptophan (MST) family. Therefore, despite their phenotypic similarities, the Mn sensitivity in themneP mneSand theyqgC-sodAdeletion mutants results from distinct enzymatic vulnerabilities.ImportanceBacteria require multiple trace metal ions for survival. Metal homeostasis relies on the tightly regulated expression of metal uptake, storage, and efflux proteins. Metal intoxication occurs when metal homeostasis is perturbed and often results from enzyme mis-metalation. InBacillus subtilis, MnSOD is the most abundant Mn-containing protein and is important for oxidative stress resistance. Here, we report novel roles for MnSOD and a co-regulated membrane protein, YqgC, in Mn homeostasis. Loss of both MnSOD and YqgC (but not the individual mutations) prevents the efficient expression of Mn efflux proteins and leads to a large-scale perturbation of the metabolome due to inhibition of Mg-dependent enzymes, including key chorismate-utilizing MST (menaquinone, siderophore, and tryptophan) family enzymes.
The human Shu complex promotes RAD51 activity by modulating RPA dynamics on ssDNA
Authors: Hengel Sarah R. , Oppenheimer Katherine , Smith Chelsea , Schaich Matthew A. , Rein Hayley L. , Martino Julieta , Darrah Kristie , Ezekwenna Oluchi , Burton Kyle , Van Houten Bennett , Spies Maria , Bernstein Kara A. 
Published: February 2024

AbstractTemplated DNA repair that occurs during homologous recombination and replication stress relies on RAD51. RAD51 activity is positively regulated by BRCA2 and the RAD51 paralogs. The Shu complex is a RAD51 paralog-containing complex consisting of SWSAP1 and SWS1. We demonstrate that SWSAP1-SWS1 binds RAD51, maintains RAD51 filament stability, and enables strand exchange. Using single molecule confocal fluorescence microscopy combined with optical tweezers, we show that SWSAP1-SWS1 decorates RAD51 filaments proficient for homologous recombination. We also find SWSAP1-SWS1 enhances RPA diffusion on ssDNA. Importantly, we show humansgSWSAP1andsgSWS1knockout cells are sensitive to pharmacological inhibition of PARP and APE1. Lastly, we identify cancer variants in SWSAP1 that alter SWS1 complex formation. Together, we show that SWSAP1-SWS1 stimulates RAD51-dependent high-fidelity repair and may be an important new cancer therapeutic target.
Performance characteristics of highly automated HSV-1 and HSV-2 IgG testing
Authors: Crawford Katharine H. D. , Selke Stacy , Pepper Gregory , Goecker Erin , Sobel Aniela , Wald Anna , Johnston Christine , Greninger Alexander L. 
Published: February 2024

ABSTRACTHerpes simplex virus (HSV) infections are one of the most common and stigmatized infections of humankind, affecting more than 4 billion people around the world and more than 100 million Americans. Yet most people do not know their infection status and antibody testing is not recommended, partly due to poor test performance. Here, we compared the test performance of the Roche Elecsys HSV-1 IgG and HSV-2 IgG, DiaSorin LIAISON HSV-1/2 IgG, and Bio-Rad BioPlex 2200 HSV-1 & HSV-2 IgG assays with the gold-standard HSV western blot in 1994 persons, including 1017 persons with PCR or culture-confirmed HSV-1 and/or HSV-2 infection. Across all samples, the Bio-Rad and Roche assays had similar performance metrics with low sensitivity (<85%), but high specificity (>97%) for detecting HSV-1 IgG and both high sensitivity (>97%) and high specificity (>98%) for detecting HSV-2 IgG. The DiaSorin assay had a higher sensitivity (92.1%) but much lower specificity (88.7%) for detecting HSV-1 IgG and comparatively poor sensitivity (94.5%) and specificity (94.2%) for detecting HSV-2 IgG. The DiaSorin assay performed poorly at low-positive index values with 60.9% of DiaSorin HSV-1 results and 20.8% of DiaSorin HSV-2 results with positive index values <3.0 yielding false positive results. Based on an estimated HSV-2 seroprevalence of 12% in the United States, positive predictive values for HSV-2 IgG were 96.1% for Roche, 87.4% for Bio-Rad, and 69.0% for DiaSorin, meaning nearly 1 of every 3 positive DiaSorin HSV-2 IgG results would be falsely positive. Further development in HSV antibody diagnostics is needed to provide appropriate patient care.
Adeno-Associated Viral Tools to Trace Neural Development and Connectivity Across Amphibians
Authors: Jaeger Eliza C.B. , Vijatovic David , Deryckere Astrid , Zorin Nikol , Nguyen Akemi L. , Ivanian Georgiy , Woych Jamie , Arnold Rebecca C. , Ortega Gurrola Alonso , Shvartsman Arik , Barbieri Francesca , Toma Florina A. , Gorbsky Gary J. , Horb Marko E. , Cline Hollis T. , Shay Timothy F. , Kelley Darcy B. , Yamaguchi Ayako , Shein-Idelson Mark , Tosches Maria Antonietta , Sweeney Lora B. 
Published: February 2024

SummaryThe development, evolution, and function of the vertebrate central nervous system (CNS) can be best studied using diverse model organisms. Amphibians, with their unique phylogenetic position at the transition between aquatic and terrestrial lifestyles, are valuable for understanding the origin and evolution of the tetrapod brain and spinal cord. Their metamorphic developmental transitions and unique regenerative abilities also facilitate the discovery of mechanisms for neural circuit remodeling and replacement. The genetic toolkit for amphibians, however, remains limited, with only a few species having sequenced genomes and a small number of transgenic lines available. In mammals, recombinant adeno-associated viral vectors (AAVs) have become a powerful alternative to genome modification for visualizing and perturbing the nervous system. AAVs are DNA viruses that enable neuronal transduction in both developing and adult animals with low toxicity and spatial, temporal, and cell-type specificity. However, AAVs have never been shown to transduce amphibian cells efficiently. To bridge this gap, we established a simple, scalable, and robust strategy to screen AAV serotypes in three distantly-related amphibian species: the frogsXenopus laevisandPelophylax bedriagae,and the salamanderPleurodeles waltl,in both developing larval tadpoles and post-metamorphic animals. For each species, we successfully identified at least two AAV serotypes capable of infecting the CNS; however, no pan-amphibian serotype was identified, indicating rapid evolution of AAV tropism. In addition, we developed an AAV-based strategy that targets isochronic cohorts of developing neurons – a critical tool for parsing neural circuit assembly. Finally, to enable visualization and manipulation of neural circuits, we identified AAV variants for retrograde tracing of neuronal projections in adult animals. Our findings expand the toolkit for amphibians to include AAVs, establish a generalizable workflow for AAV screening in non-canonical research organisms, generate testable hypotheses for the evolution of AAV tropism, and lay the foundation for modern cross-species comparisons of vertebrate CNS development, function, and evolution.
Lipid A double bond position determination using ozone and laser-induced dissociation
Authors: Mikhael Abanoub , Pětrošová Helena , Smith Derek , Ernst Robert K. , Goodlett David R. 
Published: February 2024

AbstractOzone coupled with tandem MS is a traditional tool to assign double bond position in lipids. However, few studies were reported on the use of ozone for double bond determination of bacterial lipids. Most MS-based double bond localization methods used in bacterial lipidomics focused on low molecular weight phospholipids, but never tested for assigning double bond position in complex glycolipids such as Lipid A. Here, we present a simple approach to identify unsaturated lipid A double bond position using on-MALDI plate ozone treatment of intact Lipid A.
Discovery of Alkenyl Oxindole as a Novel PROTAC Moiety for Targeted Protein Degradation via CRL4DCAF11Recruitment
Authors: Wanga Ying , Wei Tianzi , Zhao Man , Huang Aima , Sun Fan , Chen Lu , Lin Risheng , Xie Yubao , Zhang Ming , Xu Shiyu , Sun Zhihui , Hong Liang , Wang Rui , Tian Ruilin , Li Guofeng 
Published: February 2024

AbstractAlkenyl oxindoles have been characterized as autophagosome-tethering compounds (ATTECs), which can target mutant huntingtin protein (mHTT) for lysosomal degradation. In order to expand the application of alkenyl oxindoles for targeted protein degradation, we designed and synthesized a series of hetero-bifunctional compounds by conjugating different alkenyl oxindoles with the BRD4 inhibitor JQ1. Through structure-activity relationship study, we successfully developed JQ1-alkenyl oxindole conjugates that potently degrade BRD4. Unexpectedly, we found that these molecules degrade BRD4 through the ubiquitin-proteasome system, rather than the autophagy-lysosomal pathway. Using pooled CRISPR interference (CRISPRi) screening, we revealed that JQ1-alkenyl oxindole conjugates recruit the E3 ubiquitin ligase complex CRL4DCAF11for substrate degradation. Furthermore, we validated the most potent hetero-bifunctional molecule HL435 as a promising drug-like lead compound to exert antitumor activity bothin vitroandin vivo. Our research provides new employable PROTAC moieties for targeted protein degradation, providing new possibilities for drug discovery.
Structural basis for activation and mechanism of the bacterial hexameric motors RadA and ComM in DNA branch migration during homologous recombination
Authors: Rosa Leonardo Talachia , Vernhes Emeline , Soulet Anne-Lise , Polard Patrice , Fronzes Rémi 
Published: February 2024

AbstractSome DNA helicases play key roles in genome maintenance and plasticity through their branch migration activity in distinct pathways of homologous recombination. RadA is a highly conserved bacterial helicase involved in both DNA repair in all species and in natural transformation in Gram positive firmicutes. In others, ComM is the natural transformation-specific helicase. Both RadA and ComM form hexameric rings and use ATP hydrolysis as a source of energy. In this work we present the cryoEM structures of RadA and ComM interacting with DNA and ATP analogues. This is the first structure reported for ComM, and the first structure of RadA in its active state. In particular, this study unveiled a molecular switch for ATP hydrolysis and DNA binding coupling in RadA and the role of the Lon protease-like domain, shared by RadA and ComM. Together our results provide new insights on the branch migration mechanism at the molecular level in the context of homologous recombination.