bluuebunny/crossref_metadata_2025_split huggingface dataset:
Repeat expansion in a Fragile X model is independent of double strand break repair mediated by Pol θ, Rad52, Rad54l or Rad54b
Authors: Hayward Bruce E , Kim Geum-Yi , Miller Carson J , McCann Cai , Lowery Megan G. , Wood Richard D. , Usdin Karen 
Published: November 2024

AbstractMicrosatellite instability is responsible for the human Repeat Expansion Disorders. The mutation responsible differs from classical cancer-associated microsatellite instability (MSI) in that it requires the mismatch repair proteins that normally protect against MSI. LIG4, an enzyme essential for non-homologous end-joining (NHEJ), the major pathway for double-strand break repair (DSBR) in mammalian cells, protects against expansion in mouse models. Thus, NHEJ may compete with the expansion pathway for access to a common intermediate. This raises the possibility that expansion involves an NHEJ-independent form of DSBR. Pol θ, a polymerase involved in the theta-mediated end joining (TMEJ) DSBR pathway, has been proposed to play a role in repeat expansion. Here we examine the effect of the loss of Pol θ on expansion in FXD mouse embryonic stem cells (mESCs), along with the effects of mutations inRad52,Rad54landRad54b,genes important for multiple DSBR pathways. None of these mutations significantly affected repeat expansion. These observations put major constraints on what pathways are likely to drive expansion. Together with our previous demonstration of the protective effect of nucleases like EXO1 and FAN1, and the importance of Pol β, they suggest a plausible model for late steps in the expansion process.
Improved split prime editors enable efficient in vivo genome editing
Authors: Wei Rongwei , Yu Zhenxing , Ding Lihong , Lu Zhike , Yao Keyi , Zhang Heng , Huang Binglin , He Miao , Ma Lijia 
Published: November 2024

ABSTRACTPrime editor (PE) is a precise genome-editing tool that enables all 12 possible base-to-base conversions, as well as insertions and deletions, that does not require DSBs or donor DNA. The efficient delivery of prime editors in vivo is critical for realizing the full potential of prime editing in disease modeling and therapeutic correction. Although PE has been divided into two halves and delivered using dual adeno-associated viruses (AAVs), editing efficiency at different gene loci varies among split sites, and efficient split sites within Cas9 nickase are limited. In this study, by screening multiple split sites, we demonstrated that 1115 (Asn) is an efficient split site when delivering PE by dual-AAV. Besides, we utilized a feature reported by others recently that RNase could be detached from the Cas9n and designed split sites in the first half of Cas9n. We found that split-PE-367 enabled high prime editing efficiency with Rma intein. To test the editing efficiency in vivo, dual-AAV split-ePE3-367 was packaged in AAV9 and delivered by tail vein injection in mice, achieving 24.4% precise genome editing 4 weeks after injection. Our findings establish an alternative split-PE architecture that could achieve robust gene editing efficiency, facilitating the potential utility both in model organisms and as a therapeutic modality.
Pseudomonas aeruginosaessential gene perturbations that confer vulnerability to the mammalian host environment
Authors: Prasad Neha K. , Ward Ryan D. , Yu Michelle A. , Kwon Michael S. , Banta Amy B. , Rosenberg Oren S. , Peters Jason M. 
Published: November 2024

SummaryMultidrug-resistantPseudomonas aeruginosacauses highly morbid infections that are challenging to treat. While antibiotics reduce bacterial populations during infection, the host environment also plays a key role in inhibiting and eliminating pathogens. Identifying genetic targets that create vulnerabilities to the host environment may uncover strategies to synergize with nutrient limitation or inherent immune processes to clear bacterial infections. Here, we screened a partial knockdown library targetingP. aeruginosaessential and conditionally essential genes in a murine pneumonia model to identify genes with increased vulnerability in the host environment. We found that partial CRISPR interference (CRISPRi) knockdown of 178 genes showed significant fitness defects in mice relative to axenic culture. We validated two important outliers:ispD, encoding a key enzyme in isoprenoid precursor biosynthesis, andpgsA, encoding an enzyme involved in phospholipid synthesis that is strongly upregulated in human infections. Partial knockdown of both genes showed decreased virulence in a mouse survival assay but had little impact onin vitrogrowth. The use of CRISPRi screening to uncover genetic vulnerabilities represents a promising strategy to prioritize antibacterial targets that interact with the host environment.
Low dose of a non-urea selective GIRK channel activator improves hippocampal-dependent synaptic plasticity and memory disrupted by amyloid-β oligomers
Authors: Mulero-Franco Jaime , Jimenez-Herrera Raquel , Contreras Ana , Djebari Souhail , Jiménez-Díaz Lydia , Navarro-López Juan D. 
Published: November 2024

AbstractIncreased neural activity characterizes early Alzheimer’s disease (AD), serving as a prognostic indicator for disease progression and cognitive decline. Mechanisms that drive this hyperactivity and their behavioral effects remain mostly unrevealed, although normalizing altered excitability levels has been shown to reverse cognitive impairment in early AD, both in animals and humans. Soluble amyloid-βoligomers (oAβ)primary accumulate in limbic regions like hippocampus and induce neuronal hyperexcitability and subsequent cognitive deficits by impairing ion channel’s function. Indeed, G protein-gated inwardly rectifying K+ (GIRK) channels -that control neuronal excitability-are greatly affected and their selective pharmacological activation has already been shown very effective to counteract oAβ-induced hyperexcitability and hippocampal dysfunction. However, GIRK gain-of-function in healthy animals disrupts learning, memory and underlying synaptic plasticity, greatly limiting its therapeutic potential in preclinical asymptomatic early AD patients. Therefore, GIRK-based pharmacological treatment needs further investigation to overcome these limitations. Here we tested two doses of a novel, more potent, and neuronal selective GIRK activator, VU0810464, in healthy and early oAβ1-42-generated AD male and female mice. Both doses normalized hippocampal synaptic plasticity (long-term potentiation, LTP) and associated spatial object location memory (OLM) without sex dimorphism in AD animals. However, in healthy mice, low VU0810464 dose did not significantly alter LTP and OLM, whereas the high dose disrupted both. Our results suggest that the precise tuning of neural excitability with low dosing of VU0810464 might be a promising strategy to safely treat and prevent hippocampal overexcitation and upstreaming memory deficits in early preclinical asymptomatic phases of AD.
Effects of HA1 (a Probucol Analogue) and ApoC3 siRNA on Lipoprotein-Amyloid Metabolism, Neurovascular Integrity and Cognitive Function in db/db Mice
Authors: Sharif Arazu , Mamo John , Lam Virginie , Watts Gerald F , Luna Giuseppe , Nesbit Michael , Al-Salami Hani , Mooranian Armin , Takechi Ryu 
Published: November 2024

AbstractBackground and aimsChronically elevated levels of circulating lipoprotein-amyloid-β (Aβ) are implicated in the disruption of the blood-brain barrier and the initiation of a neurodegenerative cascade leading to Alzheimer’s disease (AD). Type 2 diabetes is associated with BBB dysfunction, dyslipidaemia, and an increased risk of AD. However, alterations in triglyceride-rich lipoproteins (TRL)-Aβ homeostasis and its downstream effects on the BBB in a diabetes context remain explored. This study aimed to 1) investigate, in a preclinical model of diabetes, the hypothesis that diabetes-induced impairments in TRL-Aβ metabolism might compromise BBB integrity and exacerbate cognitive function and behavioural changes, and 2) assess the efficacy of interventions that improve TRL catabolism, including probucol, HA1, and ApoC3 siRNA, to prevent disease progression by lowering circulating TRL-Aβ levels.MethodsFive-week-old db/db mice underwent a 9- or 23-weeks dietary interventions with probucol, a probucol prodrug HA1, or a standard diet with four-weekly injections of ApoC3 siRNA. Db/+ mice served as negative controls for each treatment duration. Blood levels of Aβ and ApoB were measured using ELISA. Immunofluorescence imaging was used to quantify enterocytic levels of Aβ and ApoB, and assess changes in neurovascular integrity (IgG, PDGFRβ, ZO1, occludin), neuroinflammation (GFAP, Iba1), and cerebral oxidative stress (8OHdG).ResultsOur results indicate that diabetes increased the abundance of plasma amyloid, specifically Aβ42, which correlated with enterocytic abundance, suggesting exaggerated postprandial excretion. Disrupted plasma amyloid homeostasis was associated with BBB breakdown, including diminished barrier function, and the loss of pericytes and astrocytes. Provision of the probucol analogue, HA1, normalised plasma and enterocytic amyloidemia concomitant with the preservation of the neurovascular junction. Treatment with ApoC3 siRNA attenuated plasma Aβ42 and modestly reduced neurovascular inflammation.ConclusionThe findings further support the hypothesis that aberrant peripheral metabolism of lipoprotein-Aβ is associated with microvascular corruption and the development of anxiety-like behaviour. HA1 is more effective than probucol or ApoC3 siRNA in positively modulating lipoprotein-amyloid homeostasis in db/db mice and maintaining central capillary integrity.HighlightsEnterocytic and plasma lipoprotein-Aβ levels are increased in diabetic db/db mice.Elevated plasma lipoprotein-Aβ levels correlate with BBB breakdown and anxiety-like behavior.HA1 lowers enterocytic and plasma Aβ levels, protects BBB, alleviates oxidative stress and anxiety.ApoC3 siRNA lowers plasma lipoprotein-Aβ, mitigates neuroinflammation and anxiety.
A cross-species analysis of neuroanatomical covariance sex difference in humans and mice
Authors: Pham Linh , Guma Elisa , Ellegood Jacob , Lerch Jason P. , Raznahan Armin 
Published: November 2024

ABSTRACTStructural covariance in brain anatomy is thought to reflect inter-regional sharing of developmental influences - although this hypothesis has proved hard to causally test. Here, we use neuroimaging in humans and mice to study sex-differences in anatomical covariance - asking if regions that have developed shared sex differences in volume across species also show shared sex difference in volume covariance. This study design illuminates both the biology of sex-differences and theoretical models for anatomical covariance – benefitting from tests of inter-species convergence. We find that volumetric structural covariance is stronger in adult females compared to adult males for both wild-type mice and healthy human subjects: 98% of all comparisons with statistically significant covariance sex differences in mice are female-biased, while 76% of all such comparisons are female-biased in humans (q < 0.05). In both species, a region’s covariance and volumetric sex-biases have weak inverse relationships to each other: volumetrically male-biased regions contain more female-biased covariations, while volumetrically female-biased regions have more male-biased covariations (mice: r = −0.185, p = 0.002; humans: r = −0.189, p = 0.001). Our results identify a conserved tendency for females to show stronger neuroanatomical covariance than males, evident across species, which suggests that stronger structural covariance in females could be an evolutionarily conserved feature that is partially related to volumetric alterations through sex.SIGNIFICANCE STATEMENTStructural covariance is a potent readout of coordinated brain development, but hard to probe experimentally. Here we use sex differences as a naturally occurring test for developmental theories of structural covariance – adopting a cross-species approach for validation and translational benefit. Brain MRI reveals two conserved features of anatomical covariance across humans and mice: (i) tighter inter-regional coordination of brain development in females as evidenced by stronger volume covariance; (ii) a tendency for female-biased covariance to involve regions that are smaller in females – suggesting an unknown counterbalancing between these two distinct modes of sex-biased brain organization. These findings advance understanding of coordinated brain development and sex difference in a cross-species framework – facilitating future translational research on both topics.
Sufficiency of ITD for (biased) human auditory azimuthal perception
Authors: Azevedo Anna Carolina Pereira Fernandez Oka , Pellegrino Tomás Guimarães , Peña José Luis , Marin Bóris , Pavão Rodrigo 
Published: November 2024

AbstractExperiments on human auditory perception have shown that interaural time difference (ITD) is sufficient to generate spatial percepts, even though stimuli containing only the ITD cue are perceived as being emitted from inside the head instead of from external locations at specific azimuths. These experiments are thus interpreted as “lateralization” instead of “localization” tasks. In fact, lateralized spatial perception has been quantified using tasks in which participants have to report their estimates by selecting a putative location inside the head, or matching the perceived position to sounds with a given interaural level difference. Therefore, these estimates are made with respect to internal frames of reference, but it is unclear whether these percepts have any significance for the more ecological problem of locating an external sound source.In order to investigate the link between internalized spatial percepts and sound localization, we designed a new task in which subjects are instructed to report externalized azimuthal location for sounds containing only ITD cues. Despite the mismatch between an internalized percept having to be reported as emanating from an external location, subjects were able to estimate azimuths consistently. Furthermore, normalized estimates were indistinguishable from those obtained using traditional lateralization tasks. Our results revealed a direct relationship between perceived azimuths and ITD, which deviates from that obtained from acoustical analysis of binaural recordings, revealing estimation biases. Intriguingly, these results indicate that externalized percepts are not required for the generation of azimuthal percepts.
Structure-informed mutagenesis identifies a conserved region critical for mouse insulin receptor 5’UTR IRES function
Authors: Dahl William B. , Lan Tammy C.T. , Rouskin Silvia , Marr Michael T. 
Published: November 2024

ABSTRACTCells under stress shift their proteome by repressing cap-dependent translation initiation. RNA elements called internal ribosome entry sites (IRES) can allow key cellular transcripts to remain efficiently translated to support an effective stress response. Well- characterized IRESes depend on RNA structures that reduce the protein requirements for translation initiation, thus circumventing translation inhibition. We have previously determined that the insulin receptor 5’ untranslated region (5’UTR) possesses a capacity for IRES activity that is conserved from insects to mammals. There are several prominent examples of viral IRES structures solved in solution; however, the RNA secondary structures of cellular IRESes remain mostly elusive, especiallyin vivo. Here we probe the secondary structure of the Insr 5’UTR IRES in tandem with two well-studied viral IRESes from Hepatitis C virus (HCV) and Encephalomyocarditis virus (EMCV) using dimethyl sulfate mutational profiling by sequencing (DMS-MaPseq) in cells andin vitro. We find that the viral IRES structures in cells are consistent with their knownin vitrostructures and that significant linearization of these well-studied IRESes occurs in the region surrounding their translation start codon in cells. Using the concurrent DMS- MaPseq probing as a constraint, we present a model of the mouse insulin receptor (Insr) 5’UTR. With this model as a guide, we employed a mutation strategy which allowed us to identify a conserved segment distal from the translation start codon as critical for Insr IRES function. This knowledge informed the design of a minimal IRES element with equivalent activity to the full-length Insr 5’UTR across translation contexts.BackgroundThe RNA structural requirements for cap-independent translation initiation facilitated by theMus musculusinsulin receptor (Insr) 5’ untranslated region (5’UTR) are unknown.ResultsRNA secondary structure probing of the Insr 5’UTR in cells provides a folding model used to identify elements required for cap-independent translation initiation.ConclusionA small region of the Insr 5’UTR distal to the translation start codon is necessary for cap-independent translation initiation and is fully sufficient in the proper structural context.
DNA replication fork speed Acts as a Pacer in Cortical Neurogenesis
Authors: Wang Jianhong , Kong Yifan , Li Xuezhuang , Xiang Kun , Tan Yulian , Shi Lei 
Published: November 2024

AbstractDNA replication fork speed, which controls the rate of genome duplication, has emerged as a key regulator of cellular plasticity. However, its role in neurogenesis remains unexplored. Mini-chromosome maintenance complex (MCMs)-binding protein (MCMBP) functions as a chaperone for newly synthesized MCMs, increasing chromatin coverage to restrain fork speed. We demonstrate that selective deletion of MCMBP in neural progenitor radial glial cells (RGCs) accelerates fork speed, triggering widespread apoptosis, DNA damage, and micronuclei formation, ultimately activating p53 and causing microcephaly. Unexpectedly, concurrent deletion ofTrp53andMcmbpfurther increased fork speed, leading to extensive RGC detachment from the ventricular zone and acquisition of outer-RGC characteristics. Mechanistically, we found that the MCM3 subunit coordinates DNA and centrosome duplication, thereby mediating RGC attachment. Behavioral analysis revealed that disruption of fork speed results in anxiety-like behavior in mice. These findings unveil a previously unrecognized role for replication fork speed in neurogenesis.
ANKZF1 helps to eliminate the stress-damaged mitochondria by LC3-mediated mitophagy
Authors: Ali Mudassar , Mapa Koyeli 
Published: November 2024

AbstractMitochondria, the double membrane-bound organelles of endosymbiotic origin, are crucial centers for cellular energy production and several essential metabolic pathways. Recent studies reveal that mitochondria become dysfunctional following numerous cellular stresses, and during pathologies, demanding an extensive investigation of mitochondrial turnover mechanisms. Apart from the specific response pathways to tackle different stresses, mitophagy or degradation of mitochondria by autophagy is a critical quality control mechanism that clears irreversibly damaged mitochondria. Mitophagy is majorly executed either by receptor-mediated or PINK-Parkin-dependent pathways. Here, we show that the human orthologue of yeast Vms1, ANKZF1, participates in PINK-Parkin-mediated mitophagy. We show that ANKZF1 is extensively recruited to damaged mitochondria along with Parkin during mitochondrial proteotoxic stress induced by the expression of a single misfolded/aggregated protein or during uncoupler-induced membrane depolarization. Importantly, ANKZF1 recruitment to damaged mitochondria is significantly enhanced in the presence of Parkin, and ANKZF1 physically interacts with Parkin and LC3 during mitochondrial proteotoxic or depolarization stresses. ANKZF1 harbors six putative LC3-interacting regions (LIRs), LIR4 present at residues 333-336 is particularly important for ANKZF1-LC3 interaction. Furthermore, we show thatANKZF1knockout cells are compromised in clearing stress-damaged mitochondria by mitophagy, indicating an important role of ANKZF1 in mitochondrial turnover during stress. In summary, we show a new role of ANKZF1 in eliminating the stress-damaged mitochondria, reiterating the mito-protective role of Vms1/ANKZF1 during mitochondrial stresses.
CDR2 is a dynein adaptor recruited by kinectin to regulate ER sheet organization
Authors: Teixeira Vanessa , Singh Kashish , Gama José B. , Celestino Ricardo , Carvalho Ana Xavier , Pereira Paulo , Abreu Carla M.C. , Dantas Tiago J. , Carter Andrew P. , Gassmann Reto 
Published: November 2024

ABSTRACTThe endoplasmic reticulum (ER) relies on the microtubule cytoskeleton for distribution and re-modelling of its extended membrane network, but how microtubule-based motors contribute to ER organization remains unclear. Using biochemical and cell-based assays, we identify cerebellar degeneration-related protein 2 (CDR2) and its paralog CDR2-like (CDR2L), onconeural antigens with poorly understood functions, as ER adaptors for cytoplasmic dynein-1 (dynein). We demonstrate that CDR2 is recruited by the integral ER membrane protein kinectin (KTN1) and that double knockout of CDR2 and CDR2L enhances KTN1-dependent ER sheet stacking, reversal of which by exogenous CDR2 requires its dynein-binding CC1 box motif. Exogenous CDR2 expression additionally promotes CC1 box-dependent clustering of ER sheets near centrosomes. CDR2 competes with the eEF1Bβ subunit of translation elongation factor 1 for binding to KTN1, and eEF1Bβ knockdown increases endogenous CDR2 levels on ER sheets, inducing their centrosome-proximal clustering. Our study describes a novel molecular pathway that implicates dynein in ER sheet organization and may be involved in the pathogenesis of paraneoplastic cerebellar degeneration.
Female Moths Incorporate Plant Acoustic Emissions into Their Oviposition Decision-Making Process
Authors: Seltzer Rya , Zer Eshel Guy , Yinon Omer , Afani Ahmed , Eitan Ofri , Matveev Sabina , Levedev Galina , Davidovitz Michael , Ben Tov Tal , Sharabi Gayl , Shapira Yuval , Shvil Neta , Atallah Ireen , Hadad Sahar , Ment Dana , Hadany Lilach , Yovel Yossi 
Published: November 2024

AbstractInsects rely on plants’ visual, chemical, tactile, and electrical cues when making various decisions. Recently it has been found that plants emit ultrasonic sounds, which are in the hearing range of many moths, especially under dehydration stress. In this study, we sought to determine whether insects also rely on plant acoustic signals when making decisions. We investigated whether female moths rely on ultrasonic clicks which are typically produced by dehydrated plants when deciding where to oviposit. In the absence of an actual plant, the moths indeed preferred to lay their eggs in proximity to acoustic signals which represent dehydrating plants. Tracking the moths’ behavior prior to the decision showed that they examined both sides of the arena and gradually spent more time on the acoustic-playback side. Interestingly, when actual plants were added to the arena, the oviposition preference was reversed and the moths preferred silent plants, which is in accordance with their a-priori preference for hydrated plants. Deafening the moths eliminated their preference, confirming that the choice was based on hearing. Moreover, male moth signals did not affect female oviposition decision, suggesting that the response was specific to plant sound emissions. We reveal evidence for a first acoustic interaction between moths and plants, but as plants emit various sounds, our findings hint to the existence of more currently unknown insect-plant acoustic interactions.