bluuebunny/crossref_metadata_2025_split huggingface dataset:
Recombinant B3 clade enterovirus D68 strains are efficiently rescued in 293T
cells and infect human spinal cord organoids
Authors: Jones Jennifer E. , Maya Sarah , Yovel Gal , Ciomperlik-Patton Jessica , Anstadt Jennifer , Freeman Megan Culler 
Published: January 2025

ABSTRACT
Enterovirus D68 (EV-D68) is associated with respiratory disease in children.
Between 2014 and 2018, biennial EV-D68 outbreaks coincided with peaks of a
polio-like neurologic condition called acute flaccid myelitis (AFM). We
hypothesized that specific mutations within the currently circulating B3 clade
of EV-D68 impacted neurovirulence. However, recovery of these strains from
infectious clones by published methods proved unsuccessful. Therefore, we tested
different cell lines, reagents, and conditions to enhance efficiency of
recombinant (r)EV-D68 rescue. In this study, we present a tractable rescue
system to define virulence determinants in B3 clade strains. Using this
approach, we successfully rescued historic and contemporary rEV-D68 strains to
high titer after limited cell culture passage. All strains in our study
replicated efficiently in the parental RD cell line and in human respiratory
epithelial cell lines, with BEAS-2B cells exhibiting greater permissivity than
A549 cells. While B2 and B3 clade strains could infect the SH-SY5Y neuroblastoma
cell line, the neurovirulent B2 clade strain rUSA/IL/2014-18952 was more
dependent on neuron differentiation than B3 clade strains and replication was
not sustained under multi-cycle growth conditions. Conversely, replication of
rUSA/IL/2014-18952 was more efficient in human spinal cord organoids, which
model the cellular heterogeneity of the spinal cord, while replication of B3
clade strains was more modest. Immunofluorescence staining confirmed infection,
with viral antigen colocalized with neurons. These findings suggest shifting
dynamics of EV-D68 neurovirulence and provide a critical platform for further
assessment of viral determinants of neurovirulence in B3 clade strains.

IMPORTANCE
Enterovirus D68 (EV-D68) can cause acute flaccid myelitis (AFM), a
debilitating neurological condition of the spinal cord in children.
Identifying viral determinants of EV- D68 neuropathogenesis is critical to
understanding recent shifts in AFM prevalence; however, these investigations
are limited to a small subset of infectious clones distantly related to
currently circulating B3 clade strains. In this study, we leverage improved
rescue strategies to characterize recombinant (r)EV-D68 strains from the
dominant B3 clade. While all rEV-D68 replicate efficiently in the parental
cell line and in human respiratory epithelial cells, B3 clade strains
achieved greater titers in cultured neurons than a neurovirulent B2 clade
strain, with less dependence on neuronal differentiation state. All B3 clade
strains established infection in human spinal cord organoids, but
replication varied between strains. Therefore, our study presents a
tractable rescue system to begin to dissect viral determinants of shifting
neurotropism within contemporary EV-D68 clades.
Retinal vascular dysfunction in theMthfr677C>Tmouse model of cerebrovascular disease
Authors: Reagan Alaina M. , MacLean Michael , Cossette Travis L. , Howell Gareth R. 
Published: January 2025

ABSTRACTIntroductionInvestigating retinal biomarkers for Alzheimer’s disease (AD) and related dementias (RD), has increased significantly. We examine retinal vascular health in a mouse containing the ADRD risk variantMthfr677C>Tto determine if changes in retina mirror similar changes in cerebrovasculature.MethodsMorphology and function of retinal vasculature and neurons were assessed usingin vivoimaging, immunohistochemistry and pattern electroretinography. RNAscope and proteomics were employed to determineMthfrgene expression and differential protein expression in mice carryingMthfr677C>T.ResultsMice show reduced retinal vascular density and reduced perfusion rate in aging mice, mirroring previously published brain data.Mthfris widely expressed and colocalizes with vascular cell markers. Proteomics identified common molecular signatures across brain and retina.DiscussionResults demonstrate thatMthfr-dependent vascular phenotypes occur in brain and retina similarly. These data suggest that assessing age and genetic-driven changes within retinal vasculature represents a minimally invasive method to predict AD-related cerebrovascular damage.
Sequestration of SerRS through LLPS Impairs Localized Translation and Contributes to Antibiotic Persistence
Authors: Zhang Ziyin , Li Daqian , Zheng Bo , Liu Jiafeng 
Published: January 2025

AbstractAntibiotic-tolerant persisters contribute to the emergence of resistance, posing a significant challenge to the efficacy of antibiotic therapies. Despite extensive research, the mechanisms underlying persistence remain inadequately understood. By tracking the evolution of exponential-phase bacterial populations subjected to intermittent high-dose ertapenem exposure, we characterized the evolved strains in terms of tolerance. Mutant strains, harboring mutations in the seryl-tRNA synthetase gene (serS), exhibited abrupt growth arrest upon serine depletion during exponential growth, resembling the persistence phenotype induced by serine hydroxamate (SHX). Under serine starvation, the mutated SerRS protein was sequestrated into liquid-liquid phase separation (LLPS)-driven condensates, disrupting their composition and impairing localized translation. This event precipitated growth arrest and dormancy in theSerSTstrain, triggering persistence. Our findings reveal an unrecognized role for aminoacyl-tRNA synthetases (aaRSs) in modulating bacterial condensates and provide insights into the molecular mechanisms underlying bacterial persistence.Graphic abstractSerRS Recruitment Disrupts Localized Translation in DeaD-marked Condensates.Upon serine starvation, SerRSTpartitions into LLPS-driven DeaD-marked condensates, impairing their localized translation activity and suggesting mechanisms underlying persistence arising during exponential phase. These evolutionarily conserved condensates orchestrate a robust translation program to enable stress responses and instruct cell fate decisions in bacterial populations.
Flaviviruses Hijack HuR to Suppress CNPY3: A Novel Pathway for Immune Evasion and Therapeutic Intervention
Authors: Ding Xiaoyan , He Jiuxiang , Zhao Jing , Zhou Yuxin , Chen Xiaozhong , Tong Jun , Quan Wenxuan , Qiu Minyue , Hua Dong , Liu Minchi , Guan Shan , Li Jintao 
Published: January 2025

AbstractFlaviviruses represent a serious global health threat, infecting millions annually with high rates of cross-infection among different flaviviruses, yet no broad-spectrum antiviral therapy currently targets these pathogens. Through biological big data analysis, we observed a significant downregulation of Canopy FGF Signaling Regulator 3 (CNPY3), a positive regulator of defense responses, in flavivirus-infected patients, with the level of downregulation correlating with infection severity. This observation was validated in cellular and animal models. Mechanistically, we demonstrate that flaviviruses hijack the RNA-binding protein Human antigen R (HuR) to destabilize CNPY3 mRNA via adenine-uridine-rich elements (AREs), thus downregulating CNPY3 expression and promoting viral replication. Our findings identify CNPY3 as a novel antiviral mediator essential for the TLR-mediated type I interferon (IFN-I) pathway in defense against flaviviruses. In vitro, CNPY3 overexpression reduced DENV and ZIKV replication, confirming its antiviral role. To explore therapeutic potential, we developed lipid nanoparticles (LNPs) encapsulating CNPY3 mRNA (LNP-CNPY3), and early administration in flavivirus-infected mice improved survival, reduced viral loads, and attenuated neuroinflammation by enhancing antiviral and interferon responses. Together, these findings establish CNPY3 as a critical antiviral target and support LNP-CNPY3 as a promising therapeutic approach against flavivirus infections.
Efficient searches in protein sequence space through AI-driven iterative learning
Authors: Suarez-Martin Ignacio , Risso Valeria A. , Romero-Zaliz Rocío , Sanchez-Ruiz Jose M. 
Published: January 2025

AbstractProtein sequence space is vast. This fact, together with the prevalence of epistasis, hampers the engineering of novel enzymes through library screening and is a major obstacle to any attempt to predict natural protein evolution. Recently, specialized methodologies have been used to determine fitness data on ∼260000 sequences for the gene of the enzyme dihydrofolate reductase and antibody affinity data for all combinations of the mutations present in the receptor binding domain (RBD) of the Omicron strain of SARS-CoV-2 (∼30000 variants). We show that, upon iterative training on a total of just a few hundred variants, various state-of-the-art AI tools (multi-layer perceptron, random forest and XGBoost algorithms) find very high fitness variants of the enzyme and predict the antibody-evasion patterns of the RBD. This work provides a basis for efficient, widely applicable, low-throughput experimental approaches to assess viral protein evolution and to engineer enzymes for biotechnological applications.
Non-nucleosomal (CENP-A/H4)2- DNA complexes as a possible platform for centromere organization
Authors: Ali-Ahmad Ahmad , Mors Mira , Carrer Manuel , Li Xinmeng , Bilokapić Silvija , Halić Mario , Cascella Michele , Sekulić Nikolina 
Published: January 2025

AbstractThe centromere is a part of the chromosome that is essential for the even segregation of duplicated chromosomes during cell division. It is epigenetically defined by the presence of the histone H3 variant CENP-A. CENP-A associates specifically with a group of 16 proteins that form the centromere-associated network of proteins (CCAN). In mitosis, the kinetochore forms on the CCAN to connect the duplicated chromosomes to the microtubules protruding from the cell poles. Previous studies have shown that CENP-A replaces H3 in nucleosomes, and recently the structures of CENP-A-containing nucleosomes in complex with CCANs have been revealed, but they show only a limited interaction between CCANs and CENP-A. Here, we report the cryoEM structure of 2x(CENP-A/H4)2-di-tetramers assembled on DNA in the absence of H2A/H2B histone dimer and speculate how (CENP-A/H4)2-tetramers and -di-tetramers might serve as a platform for CCAN organization.Main pointsCENP-A/H4 in the absence of H2A/H2B dimers associate on DNA to form di-tetrasomesCENP-A/H4 di-tetrasomes are stable but highly flexible structures resembling nucleosomesCENP-A/H4 tetrasomes bind CENP-N/L and can serve as platform for centromere organisation
A third type of PETase from the marineHalopseudomonaslineage
Authors: Turak Onur , Gagsteiger Andreas , Upadhyay Ashank , Kriegel Mark , Salein Peter , Agarwal Seema , Borchert Erik , Höcker Birte 
Published: January 2025

AbstractThe enzymatic degradation of polyethylene terephthalate (PET) offers a sustainable solution for PET recycling. Over the past two decades, more than 100 PETases have been characterized, primarily exhibiting similar sequences and structures. Here, we report new PET-degrading α/β hydrolases, includingHaloPETase1 from the marineHalopseudomonaslineage, thereby extending the narrow sequence space by novel features at the active site. The crystal structure ofHaloPETase1 was determined to a resolution of 1.16 Å, revealing a unique active site architecture and a lack of the canonical π-stacking clamp found in PETases so far. Further, variations in active site composition and loop structures were observed. Additionally, we found five more enzymes from the same lineage, two of which have a high similarity to type IIa bacterial PETases, while the other three resembleHaloPETase1. All these enzymes exhibited high salt tolerance ranging from 2.5 to 5 M NaCl leading to higher total product releases upon PET degradation at 40 or 50 °C. Based on these findings, we propose an extension of the existing PETase classification system to include type III PETases.
Persistent virulent phages exist in bacterial isolates
Authors: Dougherty Peter Erdmann , Bernard Charles , Carstens Alexander Byth , Stanford Kim , McAllister Tim A. , Rocha Eduardo P. C. , Hansen Lars Hestbjerg 
Published: January 2025

AbstractDespite the immense diversity of tailed bacteriophages, they are traditionally classified as either virulent or temperate, with only the latter thought capable of long-term persistence in bacterial cells through lysogeny. Virulent phages, characterized by their obligatory lytic cycle, are assumed to lack the ability to persist within bacterial colonies, and their infection is expected to decimate the host population underin-vitroconditions. Consequently, when bacterial isolates are cultured for sequencing, the resulting assemblies are not expected to contain virulent phage genomes. To test this assumption on a large scale, we analyzed over 267,000 publicly availableEscherichiaassemblies. Surprisingly, we identified 373 genomes corresponding to virulent phages within the bacterial genomes. These viral genomes are associated with specific phage groups and especially with jumbo phages with very large genomes (>200 kb). Chimallin was a core gene in two of these jumbo phage clusters, the major protein used by some jumbo phages to form a protective phage nucleus during infection. We found multiple lines of evidence suggesting that these virulent phage genomes in bacterial assemblies arise from persistent infections rather than contamination. Supporting this, we experimentally demonstrated the coexistence of non-temperate jumbo phages with their bacterial hosts. In a targeted follow-up search for three clades of persistent jumbo phages, we identified 285 additional jumbo phage genomes in bacterial taxa beyondEscherichia,highlighting that there are many more undiscovered persistent phages in bacterial assemblies. Our findings challenge the traditional virulent-temperate dichotomy, highlighting the overlooked diversity and prevalence of non-canonical phage lifestyles.
Protection of photosynthesis by UVR8 and cryptochromes in Arabidopsis under blue and UV radiation
Authors: Morales Luis Orlando , Shapiguzov Alexey , Rai Neha , Aphalo Pedro José , Brosché Mikael 
Published: January 2025

AbstractPhotosynthesis in plants is negatively affected by high light intensity and UV radiation. The photoreceptors UV RESISTANCE LOCUS 8 (UVR8) and CRYPTOCHROMES (CRYs) mediate perception and acclimation of plants to UV-B/UV-A2 (290–340 nm) and UV-A1/blue light (350–500 nm), respectively. However, their roles in photoprotection of photosynthesis across different wavebands of the spectrum remain unclear. Using chlorophyll fluorescence and LED lighting we studied the roles of UVR8 and CRYs in maintaining photosynthetic capacity in Arabidopsis exposed to UV-B, UV-A1, and blue light. Analysis of quantum yield of Photosystem II, non-photochemical quenching, and LHCII phosphorylation demonstrated that CRYs preserve photosynthetic performance in plants exposed to UV-B, UV-A1, and blue light. UVR8 and CRYs exhibit partially redundant functions in maintaining photosynthetic activity under UV-B, UV-A1, and blue light, and in preventing photodamage under high UV-A1 irradiance. Impaired UVR8 and CRY signaling reduced epidermal flavonol accumulation in leaves, which further compromised photoprotection. These findings provide valuable insights into how UV and blue light perception contribute to photoprotection, with broad implications for plant performance both in natural and managed environments.
Mechanisms of chromatin remodeling by an Snf2-type ATPase
Authors: Malik Deepshikha , Deshmukh Ashish , Bilokapic Silvija , Halic Mario 
Published: January 2025

AbstractChromatin remodeling enzymes play a crucial role in the organization of chromatin, enabling both stability and plasticity of genome regulation. These enzymes use a Snf2-type ATPase motor to move nucleosomes, but how they translocate DNA around the histone octamer is unclear. Here we use cryo-EM to visualize the continuous motion of nucleosomal DNA induced by human chromatin remodeler SNF2H, an ISWI family member. Our work reveals conformational changes in SNF2H, DNA and histones during nucleosome sliding and provides the structural basis for DNA translocation. ATP hydrolysis induces conformational changes in SNF2H that pull the DNA tracking strand, distorting DNA and histones at SHL2. This is followed by SNF2H rotation on the nucleosome, which first pulls the DNA guide strand and creates one-base pair bulge at SHL2, and then releases the pulled DNA. Given the high conservation of the catalytic motors among ATP-dependent chromatin remodelers, the mechanisms we describe likely apply to other families.
Transcription Machinery Anchors ecDNAs to Mitotic Chromosomes for Segregation
Authors: Xie Yipeng , Lim Jun Yi Stanley , Liu Wenyue , Gilbreath Collin , Kim Yoon Jung , Wu Sihan 
Published: January 2025

AbstractExtrachromosomal DNA (ecDNA) is a prevalent driver of cancer, whose random segregation promotes aggressive tumors. Acentric ecDNAs attach to chromosomes during mitosis for segregation. However, the molecular mechanism governing ecDNA-chromosome mitotic interactions remains poorly understood. This study shows that ecDNAs attach to histone 3 lysine 27 acetylation (H3K27ac)-marked chromatin during mitosis. H3K27ac depletion resulted in ecDNA detachment from mitotic chromosomes. Diverse bromodomain proteins, which are known readers of H3K27ac, stabilize ecDNAs’ mitotic interaction, exhibiting context-dependent and mutually complementary roles. Furthermore, disruptions of the Mediator complex and RNA polymerase II transcription activity both dissociate ecDNAs from mitotic chromosomes, suggesting that the transcription machinery mediates ecDNA segregation. Mis-segregated ecDNAs were expelled into the cytosol and degraded, leading to diminished oncogene expression and a reversal of therapy resistance. Our research provides new insights into the interplay between RNA transcription and acentric ecDNA inheritance in cancer, offering a novel avenue for disrupting ecDNA-driven oncogenesis.
LIN-67 functionally interacts with heterochronic miRNAs and regulates developmental timing inCaenorhabditis elegans
Authors: Medley Jeffrey C. , Perales Roberto , Humada Belén Gaete , Doerfel Katja , Levine Ariana , Panzade Ganesh , Hammell Christopher M. , Zinovyeva Anna 
Published: January 2025

AbstractTemporal regulation of gene expression is required for developmental transitions, including differentiation, proliferation, and morphogenesis. In the nematodeCaenorhabditis elegans, heterochronic microRNAs (miRNAs) regulate the temporal expression of genes that promote animal development. The heterochronic miRNAs lin-4 and let-7 are required during different stages of larval development and are associated with the miRNA-specific Argonaute ALG-1. In this study, we have identifiedlin-67as a heterochronic gene that negatively regulateslin-4,let-7,andalg-1. Loss oflin-67function restores proper developmental timing and stage-specific gene expression to hypomorphiclin-4andlet-7mutants. We found that loss oflin-67resulted in a reduced number of seam cells, defects in alae formation, precocious expression of an adult-specific gene reporter, and sterility. LIN-67 contains a K homology (KH) RNA-binding domain and is a homolog of the Sam68-like splicing factor KHDRBS2. We show that LIN-67 localizes to the nucleus throughout animal development and is enriched in nuclear foci. Mutating the KH domain of LIN-67 abolished the nuclear localization of LIN-67, suggesting that the localization of LIN-67 is likely dependent on RNA-binding activity. We show that LIN-67 negatively regulateslin-4miRNA levels and restores normal levels oflet-7toalg-1mutants, which can, at least in part, explain howlin-67suppressesalg-1. Our data indicate thatlin-67is a novel heterochronic gene that regulates developmental timing and miRNA-dependent gene regulation inC. elegans.
SSARP: An R Package for Easily Creating Species- and Speciation- Area Relationships Using Web Databases
Authors: Martinet Kristen M. , Román-Palacios Cristian , Harmon Luke J. 
Published: January 2025

AbstractA universal method of quantifying patterns of biodiversity on islands is the species-area relationship (SAR). SARs visualize the relationship between species richness (the number of species) and the area of the land mass on which they live. An extension of this visualization, the speciation-area relationship (SpAR), helps researchers determine trends in speciation rate over a set of land masses. Comparing these relationships across island systems globally is an extremely difficult task because gathering and processing a large amount of species occurrence data and island data often requires researchers to conduct lengthy literature searches and combine datasets from several different sources. Here we presentSSARP(Species/Speciation-Area Relationship Projector), an R package that provides a simple workflow for creating SARs and SpARs. TheSSARPworkflow allows users to gather occurrence data from GBIF, use mapping tools to determine whether the GPS points in the occurrence data refer to valid land masses, associate those land masses with their areas using a built-in dataset of island names and areas, and create SARs using linear and segmented regression.SSARPalso provides multiple functions for estimating speciation rates for use in creating a SpAR. UsingSSARPallows researchers to dramatically increase the scope of their biodiversity research through the creation of SARs and SpARs with data from island systems across the globe.
Identification and characterization of shifted G•U wobble pairs resulting from alternative protonation of RNA
Authors: Saon Md. Sharear , Douds Catherine A. , Veenis Andrew J. , Pearson Ashley N. , Yennawar Neela H. , Bevilacqua Philip C. 
Published: January 2025

AbstractRNA can serve as an enzyme, small molecule sensor, and vaccine, and it may have been a conduit for the origin of life. Despite these profound functions, RNA is thought to have quite limited molecular diversity. A pressing question, therefore, is whether RNA can adopt novel molecular states that enhance its function. Covalent modifications of RNA have been demonstrated to augment biological function, but much less is known about non-covalent alterations such as novel protonated or tautomeric forms. Conventionally, a G•U wobble has the U shifted into the major groove. We used a cheminformatic approach to identify four structural families of shifted G•U wobbles in which the G instead resides in the major groove of RNA, which requires alternative tautomeric states of either base, or an anionic state of the U. We provide experimental support for these shifted G•U wobbles via the potent, and unconventional,in vivoreactivity of the U with dimethylsulfate (DMS) in three organisms. These shifted wobbles may play important functional roles and could serve as drug targets. Our cheminformatics approach is general and can be applied to identify alternative protonation states in other RNA motifs, as well as in DNA and proteins.Graphical Abstract