bluuebunny/crossref_metadata_2025_split huggingface dataset:
Highly sensitive reverse transcription loop-mediated isothermal amplification for detecting and identifying groundnut ringspot and tomato spotted wilt orthotospoviruses
Authors: de Breuil Soledad , López Salvador , Dottori Carolina , Nome Claudia , Adkins Scott 
Published: December 2024

AbstractOrthotospovirus arachianuli(groundnut ringspot virus, GRSV) andOrthotospovirus tomatomaculae(tomato spotted wilt virus, TSWV) represent a threat to several agricultural crops worldwide. The management of the diseases they cause is particularly challenging due to their wide host range and transmission by thrips vectors. Accurate diagnosis and early detection are crucial, especially considering the limitations of conventional serological and molecular methods. This study introduces reverse transcription loop-mediated isothermal amplification (RT-LAMP) as a reliable alternative for detecting and identifying GRSV and TSWV in tomato, pepper, and peanut plants. The designed primer sets specifically targeted and amplified fragments of the viral RNA S segment. RT-LAMP reactions produced negative results in the healthy plants and exhibited no cross-reactivity with closely related orthotospoviruses or viruses from other genera. Compared to RT-PCR, RT-LAMP exhibited superior sensitivity, detecting viral RNA at concentrations 100 times lower. Field testing on symptomatic plants further confirmed the effectiveness of this technique. Overall, this study highlights RT-LAMP as a rapid, species-specific, and highly sensitive tool for the early detection of GRSV and TSWV in economically important crops, offering a promising solution for improved disease management.
Efficient synthesis of arabinosyl nucleosides via active site engineering of theLeishmania mexicanapurine 2′-deoxyribosyltranferase
Authors: Cecchini Davide A. , Acosta Javier , Saiz-Álvarez Laura P. , Ortega Carmen , Kaminski Pierre Alexandre , Fernández-Lucas Jesús , Hidalgo Aurelio 
Published: December 2024

AbstractEven though 2′-deoxyribosyltransferases (NDTs) have emerged as promising and eco-friendly catalysts for synthesizing different 2’-deoxynucleoside analogs, their limited activity towards substrates featuring modifications at the 2′ -C and 3′ -C positions of the ribose moiety significantly restrict their potential application in the pharmaceutical industry. Herein we report two engineered variants fromLeishmania mexicanapurine deoxyribosyltransferase (LmPDT) with the highest reported arabinosyltransferase activity for an NDT, achieved through a semi-rational design approach. The resulting variants,LmPDTN56L andLmPDTN56C, display up to 13.5- and 30-fold enhanced activity on the synthesis of vidarabine compared to the wild-type enzyme. Moreover, thermal unfolding and thermal challenge experiments revealed that the improvement in arabinosyl transferase activity did not result in a stability trade-off, withTmvalues similar to the wild-type enzyme and similar activity retention at 40 °C. After biochemical characterization, the optimal reaction parameters for variantLmPDTN56C were determined to be pH 6 and a temperature interval from 30 to 45 °C. Interestingly, under optimal operational conditions (40 °C, 50 mM MES buffer pH 6) the initial 30-fold enhancement observed forLmPDTN56C escalated to a remarkable 40-fold improvement. This comprehensive study elucidates the potential of the engineeredLmPDT variants for efficient and tailored biocatalytic synthesis of arabinosyl nucleosides, paving the way for enhanced applications in nucleoside analog production.
Sortase A-mediated farnesylation of Cdc42in vitro
Authors: Tschirpke Sophie , Hettema Nynke M. , Spitzbarth Benjamin , Eelkema Rienk , Laan Liedewij 
Published: December 2024

AbstractCdc42, a Rho-family GTPase, plays a pivotal role in establishing polarity inSaccharomyces cerevisiaeby accumulating on the membrane at the site of bud emergence. Cdc42’s ability to bind to membranes, mediated by prenylation, is essential for its function. Prenylation involves either the post-translational addition of a 15-carbon farnesyl group or a 20-carbon geranylgeranyl group to Cdc42’s C-terminus. One of the mayor challenges in studying the biophysical and biochemical interactions of Cdc42 at the polarity spotin vitrois obtaining prenylated Cdc42, due to labor-intensive and not easily reproducible traditional methods. Here, we present a streamlined, Sortase A-based approach to farnesylate Cdc42in vitro. This method leveragesE. coli-expressed Cdc42 with a Sortase A recognition motif, facilitating efficient and accessible farnesylation and purification using a purification tag-based strategy. The farnesylated Cdc42 retains functionality, as evidenced by GTP-dependent membrane binding, making it suitable for further biophysical and biochemical investigations. Additionally, our method can be easily adapted to yield geranyl-geranylated Cdc42.
Protection againstN. gonorrhoeaeinduced by OMV-based Meningococcal Vaccines are associated with cross-species directed humoral and cellular immune responses
Authors: Zhu Weiyan , Waltmann Andreea , Little Marguerite B. , Connolly Kristie L. , Matthias Kathryn A. , Thomas Keena S. , Gray Mary C. , Sikora Aleksandra E. , Criss Alison K. , Bash Margaret C. , Macintyre Andrew N. , Jerse Ann E. , Duncan Joseph A. 
Published: December 2024

AbstractLimited protective immunologic responses to naturalN. gonorrhoeaeinfection and a lack of knowledge about mechanisms of protection have hampered development of an effective vaccine. Recent studies in humans and mice have found meningococcal outer membrane vesicle-containing vaccines (OMV) induce cross species immune responses against gonococci and are associated with protection. The exact mechanisms or how humoral and cellular immunity are related to protection, remain unclear. To study this, we immunized mice with two meningococcal OMV-containing vaccines known to accelerate clearance ofN. gonorrhoeae, 4CMenB and OMV from an engineeredN. meningitidisstrain lacking major surface antigens PorA, PorB, and Rmp (MC58 ΔABR). We assessed serologic and cellular immune signatures associated with these immunizations and assessed bacterial clearance in the mice using a vaginal/cervical gonococcal infection model. Mice immunized with 4CMenB or MC58 ΔABR demonstrated shortened courses of recovery of vaginalN. gonorrhoeaecompared to control mice immunized with alum alone. Vaccination with 4CMenB or MC58ΔABR OMV elicited serum and vaginal cross-reactive anti-Ng-OMV antibody responses that were augmented after vaginal challenge withN. gonorrhoeae. Further, splenocytes in 4CMenB and MC58 ΔABR immunized mice exhibited elevated cytokine production after restimulation with heterologousN. gonorrhoeaeOMV when compared to splenocytes from Alum immunized mice. We further tested for correlations between bacterial burden and the measured anti-gonococcal immune responses within each vaccination group and found different immunologic parameters associated with reduced bacterial burden for each vaccine. Our findings suggest the cross-protection against gonococcal infection induced by different meningococcal OMV vaccines is likely multifactorial and mediated by different humoral and cellular immune responses induced by these two vaccines.
Sequence Composition Dictates Condensate Miscibility
Authors: Pei Gaofeng , Wang Xinxin , Geng Danqian , Chen Zhuo , Xu Weifan , Li Tingting , Li Pilong 
Published: December 2024

SummaryNumerous biomolecular condensates coexist within cells, yet the mechanisms governing their mixing and demixing remain elusive. To investigate how the amino acid composition of intrinsically disordered regions (IDRs) affects condensate coexistence, we paired and tested 28 IDRs in 378 different combinations. Our results reveal that IDRs enriched in serine or aromatic amino acids tend to form miscible condensates. Conversely, IDRs with high charge levels form immiscible condensates, even when the serine levels are artificially boosted. Additionally, phosphorylation acts as a switch, modulating condensate miscibility. We also observed that the miscibility between transcription factor condensates and Pol II condensates profoundly influences transcription. We engineered transcription factors by increasing serine content, enhancing both their miscibility with Pol II and their transcriptional activity. However, introducing high levels of charge had the opposite effect. These findings shed light on the fundamental mechanisms controlling condensate miscibility and offer insights for designing specific functional condensates.
Renovating the Barnes maze for mouse models of Dementia with STARR FIELD: A 4-day protocol that probes learning rate, retention and cognitive flexibility
Authors: Bertolli Aimee , Halhouli Oday , Liu-Martínez Yiming , Blaine Brianna , Thangavel Ramasamy , Zhang Qiang , Emmons Eric , Narayanan Nandakumar S. , Gumusoglu Serena B , Geerling Joel C. , Aldridge Georgina M 
Published: December 2024

ABSTRACTLand-based mazes that require spatial cues to identify the location of a hiding-place are a low-stress method to evaluate learning rate and memory retention in mice. One version, the Barnes maze, allows quantification of naturalistic exploratory behaviors not evident in water-based tasks. As the task relies on innate behaviors, it does not require overtraining, making it more feasible to examine early learning and non-memory executive functions that are characteristic of some non-amnestic dementias. However, because it is difficult to hide odor cues in the traditional version of the maze, learning rate during individual trials can be difficult to interpret. We designed and tested the use of 3D-printed escape shuttles that can be made in duplicate, as well as a docking tunnel that allows mice to self-exit the maze to improve reproducibility and limit experimenter influence. In combination with maze turning and escape tunnel caps, we show our shuttles mitigate the possibility of undesired cues. We then compare use of our 4-day protocol across several mouse models of cognitive impairment. We demonstrate an additional stage, the STARR protocol (Spatial Training and Rapid Reversal), to better challenge executive functions such as working memory and behavioral flexibility. We examine commonly used outcome measures across mice with and without access to spatial cues, as well as across mouse models of cognitive impairment to demonstrate the use of our 4-day protocol. Overall, this protocol provides detailed instructions to build and perform a robust spatial maze that can help expand the range of deficits identified. Our findings will aid in interpretation of traditional protocols, as well as provide an updated method to screen for both amnestic and non-amnestic cognitive changes.
Hypocretin Receptor 1 Blockade Early in Abstinence Prevents Incubation of Cocaine Seeking and Normalizes Dopamine Transmission
Authors: Clark Philip J. , Migovich Volodar M. , Das Sanjay , Xi Wei , Kortagere Sandhya , España Rodrigo A. 
Published: December 2024

AbstractAbstinence from cocaine use has been shown to elicit a progressive intensification or incubation of cocaine craving/seeking that is posited to contribute to propensity for relapse. While the mechanisms underlying incubation of cocaine seeking remain elusive, considerable evidence suggests that abstinence from cocaine promotes mesolimbic dopamine adaptations that contribute to exaggerated cocaine seeking. Consequently, preventing these dopamine adaptations may reduce incubation of cocaine seeking and thereby reduce the likelihood of relapse. In the present studies, we first examined if incubation of cocaine seeking was associated with aberrant dopamine transmission in the nucleus accumbens after seven days of abstinence from intermittent access to cocaine. Given the extensive evidence that hypocretins/orexins regulate motivation for cocaine, we then examined to what extent hypocretin receptor 1 antagonism on the first day of abstinence prevented incubation of cocaine seeking and dopamine adaptations later in abstinence. Results indicated that abstinence from intermittent access to cocaine engendered robust incubation of cocaine seeking in both female and male rats. We also observed aberrant dopamine transmission, but only in rats that displayed incubation of cocaine seeking. Further, we showed that a single injection of the hypocretin receptor 1 antagonist, RTIOX-276, on the first day of abstinence prevented incubation of cocaine seeking and aberrant dopamine transmission. These findings suggest that hypocretin receptor 1 antagonism may serve as a viable therapeutic for reducing cocaine craving/seeking, thus reducing the likelihood of relapse.
Circadian Activity Predicts Breeding Phenology in the Asian Burying Beetle Nicrophorus nepalensis
Authors: Chen Hao , Rubenstein Dustin R. , Mai Guan-Shuo , Chang Chung-Fan , Shen Sheng-Feng 
Published: December 2024

AbstractClimate change continues to alter breeding phenology in a range of plant and animal species across the globe. Traditional methods for assessing when organisms reproduce often rely on time-intensive field observations or destructive sampling, creating an urgent need for efficient, non-invasive approaches to assess reproductive timing. Here, we examined three populations of the Asian burying beetleNicrophorus nepalensisfrom subtropical Okinawa (500 m) and Taiwan mountains (1100-3200 m) that were reared under contrasting photoperiods in order to develop a predictive framework linking circadian activity to breeding phenology. Using automated activity monitors, we quantified adult circadian rhythms and employed machine learning to predict breeding phenology (seasonal versus year-round breeders) from behavior alone. Our model achieved 95% accuracy under long-day conditions using just three behavioural features, and notably, maintained 76% accuracy under short-day conditions when both types are reproductively active, revealing persistent behavioural differences between breeding strategies. These results demonstrate how integrating behavioural monitoring with machine learning can provide both a rapid, scalable method for tracking population responses to climate change and novel insights into species’ adaptive responses to shifting seasonal cues across different elevational gradients in their native range.
Deep conservation of cis-regulatory elements and chromatin organization in echinoderms uncover ancestral regulatory features of animal genomes
Authors: Magri Marta S. , Voronov Danila , Foley Saoirse , Martínez-García Pedro Manuel , Franke Martin , Cary Gregory A. , Santos-Pereira José M. , Cuomo Claudia , Fernández-Moreno Manuel , Gil-Galvez Alejandro , Acemel Rafael D. , Paganos Periklis , Ku Carolyn , Ranđelović Jovana , Rusciano Maria Lorenza , Firbas Panos N. , Gómez-Skarmeta José Luis , Hinman Veronica F. , Arnone Maria Ina , Maeso Ignacio 
Published: December 2024

AbstractDespite the growing abundance of sequenced animal genomes, we only have detailed knowledge of regulatory organization for a handful of lineages, particularly flies and vertebrates. These two groups of taxa show contrasting trends in the molecular mechanisms of 3D chromatin organization and long-term evolutionary dynamics of cis-regulatory element (CREs) conservation. To help us identify shared versus derived features that could be responsible for the evolution of these different regulatory architectures in animals, we studied the evolution and organization of the regulatory genome of echinoderms, a lineage whose phylogenetic position and relatively slow molecular evolution has proven particularly useful for evolutionary studies. First, using PacBio and HiC data, we generated new reference genome assemblies for two species belonging to two different echinoderm classes: the purple sea urchinStrongylocentrotus purpuratusand the bat sea starPatiria miniata. Second, we characterized their 3D chromatin architecture, identifying TAD-like domains in echinoderms that, like in flies, do not seem to be associated with CTCF motif orientation. Third, we systematically profiled CREs during sea star and sea urchin development using ATAC-seq, comparing their regulatory logic and dynamics over multiple developmental stages. Finally, we investigated sea urchin and sea star CRE evolution across multiple evolutionary distances and timescales, from closely related species to other echinoderm classes and deuterostome lineages. This showed the presence of several thousand elements conserved for hundreds of millions of years, revealing a vertebrate-like pattern of CRE evolution that probably constitutes an ancestral property of the regulatory evolution of animals.
Gene mobility elements mediate cell type specific genome organization and radial gene movementin vivo
Authors: Lucas Tanguy , Wang Lin-Ing , Glass-Klaiber Juniper , Quiroz Elvis , Patra Sofiya , Molotkova Natalia , Kohwi Minoree 
Published: December 2024

ABSTRACTUnderstanding the level of genome organization that governs gene regulation remains a challenge despite advancements in chromatin profiling techniques. Cell type specific chromatin architectures may be obscured by averaging heterogeneous cell populations. Here we took a reductionist perspective, starting with the relocation of thehunchbackgene to the nuclear lamina inDrosophilaneuroblasts. We previously found that this event terminates competence to produce early-born neurons and is mediated by an intronic 250 base-pair element, which we term gene mobility element (GME). Here we found over 800 putative GMEs globally that are chromatin accessible and are Polycomb (PcG) target sites. GMEs appear to be distinct from PcG response elements, however, which are largely chromatin inaccessible in neuroblasts. Performingin situHi-C of purified neuroblasts, we found that GMEs form megabase-scale chromatin interactions, spanning multiple topologically associated domain borders, preferentially contacting other GMEs. These interactions are cell type and stage-specific. Notably, GMEs undergo developmentally- timed mobilization to/from the neuroblast nuclear lamina, and domain swapping a GFP reporter transgene intron with a GME relocates the transgene to the nuclear lamina in embryos. We propose that GMEs constitute a genome organizational framework and mediate gene-to-lamina mobilization during progenitor competence state transitionsin vivo.
TTF2 promotes replisome eviction from stalled forks in mitosis
Authors: Can Geylani , Shyian Maksym , Krishnamoorthy Archana , Lim Yang , Wu R. Alex , Zaher Manal S. , Raschle Markus , Walter Johannes C. , Pellman David S. 
Published: December 2024

AbstractWhen cells enter mitosis with under-replicated DNA, sister chromosome segregation is compromised, which can lead to massive genome instability. The replisome-associated E3 ubiquitin ligase TRAIP mitigates this threat by ubiquitylating the CMG helicase in mitosis, leading to disassembly of stalled replisomes, fork cleavage, and restoration of chromosome structure by alternative end-joining. Here, we show that replisome disassembly requires TRAIP phosphorylation by the mitotic Cyclin B-CDK1 kinase, as well as TTF2, a SWI/SNF ATPase previously implicated in the eviction of RNA polymerase from mitotic chromosomes. We find that TTF2 tethers TRAIP to replisomes using an N-terminal Zinc finger that binds to phosphorylated TRAIP and an adjacent TTF2 peptide that contacts the CMG-associated leading strand DNA polymerase ε. This TRAIP-TTF2-pol ε bridge, which forms independently of the TTF2 ATPase domain, is essential to promote CMG unloading and stalled fork breakage. Conversely, RNAPII eviction from mitotic chromosomes requires the ATPase activity of TTF2. We conclude that in mitosis, replisomes undergo a CDK- and TTF2-dependent structural reorganization that underlies the cellular response to incompletely replicated DNA.
A tool to automate assessment of regional brain atrophy in mouse models of neurodegenerative disease
Authors: Moldenhauer Samuel , Potluri Nalini , Xie Yuanyun , Southwell Amber L 
Published: December 2024

AbstractAs life expectancy rises, so too does the prevalence of neurodegenerative diseases. Neurodegeneration causes progressive regional brain atrophy, typically initiating prior to symptom onset. Researchers measure the impact of potential treatments on atrophy in mouse models to assess their effectiveness. This is important because treatments designed to combat neuropathology are more likely to modify the disease, per contra to symptom management. Magnetic resonance imaging, while accurate in measurement of brain region structure volumes, is prohibitively expensive. Conversely, stereological volume assessment, the process of estimating the volume of individual 3D brain regions from imaged 2D brain sections, is more commonly used. This involves manually tracing brain region(s) of interest in regularly spaced imaged cross-sections to determine their 2D area, followed by application of the Cavalieri principle to estimate the volume. The pertinent caveats of this approach are the labor-intensive manual tracing process, and potential inaccuracies that arise due to human variation. To overcome these challenges, we have created a Neuropathology Assessment Tool (NAT) to automate regional brain tracing and identification using artificial intelligence (AI) and concepts from topological data analysis. The NAT was validated by comparing manual and NAT analysis of striatal volume in Huntington disease model mice. The NAT detected striatal atrophy with higher efficiency, 93.8% agreement with manual measurements, and lower inter-group variability. The NAT will increase efficiency of preclinical neuropathology assessment, allowing for a greater number of experimental therapies to be tested and facilitating drug discovery intractable neurodegenerative diseases.
Mechanisms of Social Behavior in the Anti-Social Blind Cavefish (Astyanax mexicanus)
Authors: Sekulovski Britney , Miller Noam 
Published: December 2024

AbstractThe evolution of social behavior inAstyanax mexicanus, which exists as a sighted, surface-dwelling morph and a blind, cave-dwelling morph, provides a model for understanding how environmental pressures shape social behaviors. We compared the shoaling behavior of blind and surfaceA. mexicanusto that of zebrafish (Danio rerio), and examined the effects of nutritional state and the neuropeptides isotocin (IT) and arginine vasotocin (AVT) on their social behavior. Blind cavefish not only fail to form shoals, but actively avoid conspecifics, with hunger further diminishing their social cohesion. Administration of low doses of AVT and an IT antagonist partially restored social behavior in blind cavefish, reducing distances between individuals, whereas surface fish exhibited minimal or opposite responses to these hormonal manipulations. Our findings suggest that the loss of schooling behavior in blind cavefish is not a consequence of visual impairment alone, as they remain capable of detecting and responding to others. Instead, this behavior likely reflects an adaptive response to their resource-poor, predator-free cave environment, where shoaling may be disadvantageous. The differing responses to social hormones between the morphs indicate that blind cavefish may have lost the motivation to shoal rather than the ability, highlighting how ecological pressures can shape social behavior.