bluuebunny/crossref_metadata_2025_split huggingface dataset:
Generation of a novel constitutive smooth muscle cell-specificMyh11-driven Cre mouse model
Authors: Dong Kunzhe , Bai Zhixia , He Xiangqin , Zhang Lu , Hu Guoqing , Yao Yali , Cai Chen-Leng , Zhou Jiliang 
Published: November 2024

AbstractDysfunction in either embryonic or postnatal vascular smooth muscle cells (SMCs) significantly contributes to the progression of various cardiovascular diseases. Therefore, elucidating the molecular mechanisms governing VSMC development and homeostasis is crucial.MYH11is the most reliable lineage gene for SMCs and has been utilized to develop tamoxifen-inducible Cre driver lines for achieving SMC-specific gene manipulation by crossing with mice carrying the loxP-flanked gene, particularly in adult mice. For studies involving SMCs during embryogenesis, the commonly used constitutive Cre driver is controlled by theTagln(Sm22α) promoter. However, this Cre driver exhibits activity in multiple non-SMC populations, including cardiomyocytes and skeletal muscle precursors, introducing confounding effects. Additionally, most existing SMC-specific Cre drivers are generated using a transgenic approach, raising concerns about random site integration and variable gene copy numbers. To address these limitations, we report a novel Cre mouse model generated by knock-in (KI) of a nuclear-localized Cre recombinase into theMyh11gene locus using homologous recombination. We confirmed that the Cre activity precisely recapitulates endogenousMyh11expression by crossing withRosa26mTmG or tdTomato reporter mice. Moreover,Myh11-driven Cre can efficiently delete the floxed allele of the transcription factorTead1specifically in SMCs. TheTead1SMC-specific knockout mice did not exhibit an overt phenotype, thereby circumventing the embryonic lethal phenotype mediated byTagln-driven Cre, as we previously reported. These findings establish this novel Cre driver line as a robust tool for tracing theMyh11-positive SMC lineage and manipulating gene function specifically in SMCs during embryonic development in mice.
Structural landscape of AAA+ ATPase motor states in the substrate-degrading human 26S proteasome reveals conformation-specific binding of TXNL1
Authors: Arkinson Connor , Gee Christine L. , Zhang Zeyuan , Dong Ken C. , Martin Andreas 
Published: November 2024

SummaryThe 26S proteasome targets many cellular proteins for degradation during general homeostasis, protein quality control, and the regulation of vital processes. A broad range of proteasome-interacting cofactors thereby modulates these functions and aids in substrate degradation. Here, we solved several high-resolution structures of the redox active cofactor TXNL1 bound to the human 26S proteasome at saturating and sub-stoichiometric concentrations by time resolved cryo-EM. We identified distinct binding modes of TXNL1 that depend on the proteasome conformational and ATPase motor states. Together with biophysical and biochemical experiments, our structural studies reveal that the resting-state proteasome prior to substrate engagement with the ATPase motor binds TXNL1 with low affinity and in variable positions on top of the Rpn11 deubiquitinase. In contrast, the actively degrading proteasome shows additional interactions leading to high-affinity TXNL1 binding, whereby TXNL1’s C-terminal tail covers the catalytic groove of the Rpn11 deubiquitinase and coordinates the active-site Zn2+. Furthermore, these cryo-EM structures of the degrading proteasome capture the ATPase hexamer in all registers of spiral-staircase arrangements and thus visualize the complete ATP-hydrolysis cycle of the AAA+ motor, indicating temporally asymmetric hydrolysis and conformational changes in bursts during mechanical substrate unfolding and translocation. Remarkably, we catch the proteasome in the act of unfolding the beta-barrel mEos3.2 substrate while the ATPase hexamer is in a particular spiral staircase register. Our findings challenge current models for protein translocation through hexameric AAA+ motors and reveal how the proteasome uses its distinct but broad range of conformational states to coordinate cofactor binding and substrate processing.HighlightsHigh resolution structures of the redox active cofactor TXNL1 in complex with the human 26S proteasome solved by time-resolved cryo-EM.TXNL1 binds the catalytic groove of the main proteasomal deubiquitinase Rpn11 and coordinates its active-site Zinc specifically in substrate-degrading states of the proteasome.Visualizing a partially unfolded intermediate of the mEos model substrate during processing.Structures of the actively degrading human proteasome reveal all spiral-staircase registers of the AAA+ ATPase hexamer with unexpected nucleotide occupancies that indicate asymmetric ATP hydrolysis mechanisms, conformational changes with burst phases, and thus new models for hand-over-hand substrate translocation.
Commonalities and differences in the Cryptococcus neoformans response to ingestion by protozoal and mammalian phagocytic cells suggests host cell metabolism as a target for intracellular survival
Authors: Dragotakes Quigly , Jacobs Ella , Gerbig Gracen , Greengo Seth , Jedlicka Anne , Dziedzic Amanda , Casadevall Arturo 
Published: November 2024

AbstractThe outcome of the interaction between Cryptococcus neoformans and infected hosts can be determined by whether the fungal cell survives ingestion by phagocytic cells. This applies to both unicellular and multicellular hosts such as amoeba and animals, respectively. Ingestion by phagocytic cells results in the formation of the cryptococcal phagosome but this structure has proved difficult to isolate. In this study, we report the successful isolation of cryptococcal phagosomes from murine and human phagocytes, followed by their characterization using proteomic and transcriptional analysis. Comparison of cryptococcal proteins from Acanthamoeba castellanii, Mus musculus, and Homo sapiens phagocytes revealed the existence of a shared set suggesting a conserved fungal response to ingestion by phagocytic cells. Given that the cryptococcal intracellular pathogenic strategy is ancient, dating to at least to the cretaceous epoch, these results are consistent with the notion that the fungal response to ingestion reflects the result of selection pressures by environmental ameboid predators over eons of evolutionary time. We propose the existence of a conserved cryptococcal toolkit for intracellular survival that includes metabolic enzymes. We propose that host cell metabolic disruption provides a common strategy for the cryptococcal survival after ingestion by phylogenetically distant phagocytic hosts.
Drospondin, a novel glial-secreted glycoprotein, controls the development and function of theDrosophilanervous system
Authors: Rojo-Cortés Francisca , Roa-Siegfried Candy B. , Fuenzalida-Uribe Nicolás , Amado-Hinojosa Paula , González-Ramírez María-Constanza , Almonacid-Torres Isidora , Birman Serge , Goodman Lindsey D. , Kanca Oguz , Oliva Carlos , Marzolo María Paz , Campusano Jorge M. 
Published: November 2024

ABSTRACTReelin is a secreted glycoprotein with roles in the development of the mammalian neocortex, hippocampus and cerebellum. This vertebrate signaling molecule also contributes to adult brain function. Mammalian Reelin increases the complexity ofDrosophilaMushroom Body (MB) neurites, an effect mediated by LpR1 and LpR2, the orthologs of mammalian Reelin receptors. Paradoxically, to date, no Reelin ortholog has been described inDrosophila. Here, we report that the protein product of the uncharacterizedDrosophila CG17739gene, which we named Drospondin, shares sequence homology with vertebrate F-spondin and Reelin. We show that Drospondin is expressed in glial cells and is crucial for MB development. Our results also show that Drospondin genetically interacts with LpRs and that human Reelin rescues structural defects in Drospondin-deficient flies. Furthermore, Drospondin-deficient flies exhibit altered sleep homeostasis, locomotion, and social behaviors. Our results reveal that flies express a functional homolog of mammalian Reelin that controls the development and function of theDrosophilanervous system.
Identification of Covalent Cyclic Peptide Inhibitors Targeting Protein-Protein Interactions Using Phage Display
Authors: Wang Sijie , Faucher Franco F. , Bertolini Matilde , Kim Heeyoung , Yu Bingchen , Cao Li , Roeltgen Katharina , Lovell Scott , Shanker Varun , Boyd Scott D. , Wang Lei , Bartenschlager Ralf , Bogyo Matthew 
Published: November 2024

ABSTRACTPeptide macrocycles are promising therapeutics for a variety of disease indications due to their overall metabolic stability and potential to make highly selective binding interactions with targets. Recent advances in covalent macrocycle peptide discovery, driven by phage and mRNA display methods, have enabled the rapid identification of highly potent and selective molecules from large libraires of diverse macrocycles. However, there are currently limited examples of macrocycles that can be used to disrupt protein-protein interactions and even fewer examples that function by formation of a covalent bond to a target protein. In this work, we describe a directed counter-selection method that enables identification of covalent macrocyclic ligands targeting a protein-protein interaction using a phage display screening platform. This method utilizes binary and ternary screenings of a chemically modified phage display library, employing the stable and weakly reactive aryl fluorosulfate electrophile. We demonstrate the utility of this approach using the SARS-CoV-2 Spike-ACE2 protein-protein interaction and identify multiple covalent macrocyclic inhibitors that disrupt this interaction. The resulting compounds displayed antiviral activity against live virus that was irreversible after washout due to the covalent binding mechanism. These results highlight the potential of this screening platform for developing covalent macrocyclic drugs that disrupt protein-protein interactions with long lasting effects.
Carotenoid pigments enhance rhodopsin-mediated phototrophy by light-harvesting and photocycle-accelerating
Authors: Fujiwara Takayoshi , Hosaka Toshiaki , Hasegawa-Takano Masumi , Nishimura Yosuke , Tominaga Kento , Mori Kaho , Nishino Satoshi , Takahashi Yuno , Uchikubo-Kamo Tomomi , Hanada Kazuharu , Maoka Takashi , Takaichi Shinichi , Inoue Keiichi , Shirouzu Mikako , Yoshizawa Susumu 
Published: November 2024

SummaryMicrobial rhodopsins, photoreceptor proteins widely distributed in marine microorganisms, receive large amount of light energy that sustains marine ecosystems. Although rhodopsins generally harbor retinal as their only chromophore, a recent study reported that carotenoid antennae transfer light energy to the retinal in proteorhodopsins, proton pump rhodopsins abundant in marine environments. Here, using marine bacterial isolates, we detected energy transfer from a carotenoid (myxol) to retinal not only in proteorhodopsin but also in the chloride ion-pumping rhodopsin. Carotenoid binding improved the light utilization efficiency of the proteorhodopsin by accelerating the photocycle, together with facilitating light-harvesting. The carotenoid-binding ability is conserved in rhodopsins of the phylumBacteroidota, which are widely transcribed in the photic zone. These findings suggest that the distribution of carotenoid-binding rhodopsins is more taxon-specific than previously thought, thus underscoring the importance of carotenoid-binding rhodopsins that provide an extended light utilization strategy in the environmental adaptation of marine bacteria.
Drugs Commonly Used in Fungal Infection Risk Conditions Promote Drug Tolerance or Resistance inCandida albicans
Authors: Obermeier Mariella , Esparza Mora Margy Alejandra , Heese Olivia , Cohen Nir , Varma Sreejith Jayasree , Tober-Lau Pinkus , Hartl Johannes , Kurth Florian , Berman Judith , Ralser Markus 
Published: November 2024

AbstractBackgroundFungal infections are an increasing concern, particularly among immunocompromised patients and those with comorbidities who require multiple medications. However, the effects of drugs targeting human pathways on fungal cells, and whether they influence antifungal drug responses, are poorly understood.MethodsWe systematically analyzed clinical guidelines to shortlist non-antifungal drugs commonly used in conditions that increase the likelihood of fungal infections. Focusing on the most prevalent fungal pathogen, we then tested how these drugs affected the antifungal response ofCandida albicansto two commonly used antifungals, fluconazole and anidulafungin. Drug interactions identified were further assessed using checkerboard and disk diffusion assays. Finally, antifungal treatment efficacy of fluconazole in combination with negatively interacting drugs was evaluated in anin vivo Galleria mellonellamodel of disseminatedC. albicansinfection.FindingsOut of 119 drugs frequently co-administered with antifungals in 40 conditions associated with a high risk of fungal infections, 34 compounds affected the antifungal drug response inC. albicans, with most drugs reducing or antagonising antifungal efficacy, several through increasing resistance or tolerance. Notably, fluconazole combinations with carvedilol and loperamide promoted antifungal resistance in both fungal cultures and inGalleria mellonella.InterpretationOur findings suggest that medications frequently taken by patients at risk of fungal infections regularly act on the fungal pathogens and can affect the effectiveness of antifungals. We propose that human drugs acting on fungal pathogens may be an underestimated factor contributing to the evolution of antifungal tolerance and resistance.
A recurrent vision transformer shows signatures of primate visual attention
Authors: Morgan Jonathan , Albanna Badr , Herman James P. 
Published: November 2024

AbstractWe present a Recurrent Vision Transformer (Recurrent ViT) that integrates a capacity-limited spatial memory module with self-attention to emulate primate-like visual attention. Trained via reinforcement learning on a spatially cued orientation-change detection task, our model exhibits hallmark behavioral signatures of primate attention—including improved detection accuracy and faster reaction times for cued stimuli that scale with cue validity. Analysis of its self-attention maps reveals rich temporal dynamics: spatial biases induced by cues are maintained during blank intervals and reactivated prior to anticipated stimulus changes, mirroring the top–down modulation observed in primate studies. Moreover, targeted manipulations of internal attention weights yield performance changes analogous to those produced by microstimulation in attentional control regions such as the frontal eye fields and superior colliculus. These findings demonstrate that embedding recurrent, memory-driven mechanisms within transformer architectures may provide a computational framework for linking artificial and biological attention
Sex differences in central amygdala glutamate responses to calcitonin gene-related peptide
Authors: Lorsung Rebecca , Cramer Nathan , Alipio Jason Bondoc , Ji Yadong , Han Sung , Masri Radi , Keller Asaf 
Published: November 2024

AbstractWomen are disproportionately affected by chronic pain compared to men. While societal and environmental factors contribute to this disparity, sex-based biological differences in the processing of pain are also believed to play significant roles. The central lateral nucleus of the amygdala (CeLC) is a key region for the emotional-affective dimension of pain, and a prime target for exploring sex differences in pain processing since a recent study demonstrated sex differences in CGRP actions in this region. Inputs to CeLC from the parabrachial nucleus (PB) play a causal role in aversive processing, and release both glutamate and calcitonin gene-related peptide (CGRP). CGRP is thought to play a crucial role in chronic pain by potentiating glutamatergic signaling in CeLC.However, it is not known if this CGRP-mediated synaptic plasticity occurs similarly in males and females. Here, we tested the hypothesis that female CeLC neurons experience greater potentiation of glutamatergic signaling than males followingendogenousCGRP exposure. Using trains of optical stimuli to evoke transient CGRP release from PB terminals in CeLC, we find that subsequent glutamatergic responses are preferentially potentiated in CeLC neurons from female mice. This potentiation was CGRP-dependent and involved a postsynaptic mechanism. This sex difference in CGRP sensitivity may explain sex differences in affective pain processing.Significance statementThe central lateral nucleus of the amygdala (CeLC) receives a dense projection from parabrachial nucleus (PB) neurons that corelease calcitonin gene-related peptide (CGRP) and glutamate following aversive stimuli. This PBCGRP→CeLC projection plays a causal role in chronic pain. We show that endogenous CGRP release potentiates glutamate signaling in female, but not male, CeLC neurons. In the context of previous work in male CeLC, this suggests that that females are more sensitive to even transient CGRP release events. Understanding how this sex difference in CGRP sensitivity arises could enhance strategies for treating chronic pain in both women and men.
Comprehensive benchmarking of tools for nanopore-based detection of DNA methylation
Authors: Kulkarni Onkar , Mathew Reuben Jacob , Zaveri Lamuk , Jana Rhea , Singh Nitesh Kumar , Ara Sreenivas , Tallapaka Karthik Bharadwaj , Sowpati Divya Tej 
Published: November 2024

AbstractLong read sequencing technologies such as Oxford Nanopore (ONT) offer direct detection of DNA base modifications. While several tools and models have been developed to identify DNA methylation from nanopore data, they are often restricted to 5-methylcytosine (5mC) and older flowcell (FC) chemistry. The performance and accuracy of newer models, including those developed by ONT, remain unknown, particularly for their new FC chemistry (R10.4.1) and sampling rate (5kHz). Here, using a variety of bacterial and human datasets, we systematically evaluate the performance of existing methylation models for 5mC (both CpG and non-CpG contexts), 6-methyladenine, and 4-methylcytosine. We also demonstrate the effect of other parameters such as sequencing depth, read quality, basecalling mode, and more importantly, the presence of neighboring DNA modifications. Our work thus provides important information to researchers utilizing nanopore sequencing to study DNA modifications, and highlights the lacunae in current generation methylation detection models.
Crystal structure of plant γ-glutamyl peptidase 1 with dual roles in sulfur metabolism and implications for oxidative stress regulation
Authors: Sone Kosei , Ito Takehiro , Yamada Chihaya , Kashima Toma , Miyanaga Akimasa , Ohkama-Ohtsu Naoko , Fushinobu Shinya 
Published: November 2024

Abstractγ-Glutamyl peptidase 1 (GGP1) plays a dual role in primary and secondary sulfur metabolism inArabidopsis thaliana. During glutathione (GSH) turnover, GGP1 hydrolyzes the isopeptide bond of GSH to degrade the tripeptide into Glu and Cys-Gly. During glucosinolate and camalexin biosynthesis, GGP1 processes GSH conjugates, which have a large substituent at the thiol side chain, by hydrolyzing the same isopeptide bond of γ-Glu. In the present study, we determined the crystal structures of the following GGP1 forms: ligand-free, Glu complex, covalent γ-Glu intermediate, and disulfide-linked S-S inactive forms. The intermediate structure, in which γ-Glu is covalently linked to the nucleophile C100, was trapped by mutating the catalytic His to Asn (H192N). In the Glu complex and γ-Glu intermediate structures, Glu bound to the S1 subsite is extensively recognized by several hydrogen bonds. The substrate recognition of the Cys-Gly moiety at the S1’ and S2’ subsites was revealed by modeling GSH in the active site. Mutational analysis indicated that R206 plays an important role in substrate binding by forming a salt bridge with Gly at the S2’ subsite. An open pocket is present beyond the thiol side chain of Cys in the S1’ subsite, which contributed to the dual activity of GGP1 toward GSH and GSH conjugates. The S-S inactive structure was obtained by soaking GGP1 crystals in Cys-Gly, and the catalytic cysteine (C100) partially formed a disulfide bond with a neighboring C154 residue. The partial inactivation of GGP1 in the presence of a pro-oxidant (Cys-Gly) has revealed its possible role in oxidative stress regulation inArabidopsis.
Anterior cingulate neurons display subregion-specific interaction with frontal eye fields as revealed by combined antidromic stimulation and resting state imaging
Authors: Babapoor-Farrokhran Sahand , Major Alex J. , Johnston Kevin D. , Miller Earl K. , Camprodon Joan A. , Everling Stefan 
Published: November 2024

AbstractThe anterior cingulate cortex (ACC) is thought to exert cognitive control over saccade generation in the frontal eye fields (FEF), but the nature of this interaction remains unclear. Although prior imaging studies have suggested ACC interacts with FEF, few studies have confirmed this by electrophysiological recordings. This study aimed to characterize the functional connectivity between ACC and medial and lateral FEF during cognitive saccade tasks. We combined resting-state functional MRI (rs-fMRI) with single-unit electrophysiology in two macaque monkeys performing memory-guided saccade and pro-/anti-saccade tasks. Anti- and ortho-dromic stimulation was used to electrophysiologically identify ACC neurons mono- and polysynaptically connected to FEF. We analyzed ACC neuronal selectivity for different task aspects and correlated these properties with both positive and negative rs-fMRI functional connectivity between ACC and FEF subregions. Anti- and ortho-dromically identified ACC neurons were predominantly connected to medial FEF, which showed stronger positive functional connectivity with ACC compared to lateral FEF. Sites with higher proportions of task-selective neurons yielded stronger functional connectivity with FEF. This stronger functional connectivity was particularly related to the delay and saccadic periods of different cognitive saccade tasks. Using combined imaging and electrophysiology, our findings provide converging evidence for functional interactions between ACC and FEF, predominantly medial FEF regions which encode large amplitude saccades. The correlation between functional connectivity and task-related neuronal selectivity supports ACC’s interaction with FEF in the modulation of saccade generation and cognitive control. Additionally, we report suggestive evidence that mono- and poly-synaptic connections may be related to positive functional connectivity, but we found no such relationship for negative functional connectivity (anticorrelations). These results advance our understanding of prefrontal cortical interactions in oculomotor control and the electrophysiological mechanisms of positive and negative resting-state functional connectivity.