bluuebunny/crossref_metadata_2025 huggingface dataset:
Disrupted Sphingosine-1-Phosphate Homeostasis Drives Nephrotoxicity in Sphingosine-1-Phosphate Lyase Insufficiency Syndrome (SPLIS)
Authors: Majcher Adam , Buder Kathrin , Khan Ranjha , Saba Julie D. , Hornemann Thorsten
Published: January 2025

Sphingosine-1-phosphate lyase insufficiency syndrome (SPLIS), also known as nephrotic syndrome type 14 (NPHS14), is an autosomal recessive disorder characterized by renal, neurological, dermatological, endocrine, and immunological symptoms. This condition is caused by loss-of-function mutations in theSGPL1gene, which encodes sphingosine-1-phosphate lyase (SGPL1p/SPL), the enzyme responsible for the terminal degradation of sphingosine-1-phosphate (S1P) in sphingolipid catabolism.

We investigated a novel case of SPLIS associated with a recently reportedSGPL1mutation (c.1084T>A; p.Ser362Thr). Using stable isotope flux analyses, we demonstrated in patient-derived fibroblasts andHEK293TSGPL1 knockout models that SGPL1p deficiency does not consistently result in pathological S1P accumulation.

Instead, SPL-deficient cells are able to maintain steady-state S1P levels through two compensatory mechanisms:

Regulation of de novo sphingolipid synthesis via the ORMDL-ceramide axis.

Increased conversion of excess ceramides into glycosphingolipids.

However, when steady-state conditions are disrupted—either by external sphingolipid supplementation or by impairing homeostatic control—a pathological increase in intracellular S1P occurs in SPL-deficient cells.

In vivo,Sgpl1-/-mice exhibited significant urinary excretion of S1P and marked S1P enrichment in the kidneys. This pathological accumulation of S1P dysregulates cytoskeletal homeostasis, impairing renal epithelial formation. Based on these findings, we hypothesize that the reabsorption of urinary S1P contributes to toxic renal accumulation, providing an explanation for the nephrotoxicity observed in SPLIS and its association with nephrotic syndrome.

Importantly, we found that the cytoskeletal disruptions could be mitigated by inhibiting the Rho-ROCK signaling pathway using the clinically approved inhibitor Fasudil. These findings illuminate the pathophysiological basis of SPLIS nephrotoxicity and propose a promising pharmacological intervention strategy.

Graphical abstract
Tunable fluorogenic DNA probes drive fast and high-resolution single-molecule fluorescence imaging
Authors: Kümmerlin Mirjam , Zhao Qing , Hazra Jagadish , Hepp Christof , Farrar Alison , Turner Piers , Kapanidis Achillefs N.
Published: January 2025

A main limitation of single-molecule fluorescence (SMF) measurements is the “high concentration barrier”, describing the maximum concentration of fluorescent species tolerable for sufficient signal-to-noise ratio (SNR). To address this barrier in several SMF applications, we design fluorogenic probes based on short ssDNAs, fluorescing only upon hybridising to their complementary target sequence. We engineer the quenching efficiency and fluorescence enhancement upon duplex formation through screening several fluorophore-quencher combinations, label lengths, and sequence motifs, which we utilise as tuning screws to adapt our labels to different experimental designs. Using these fluorogenic probes, we can perform SMF experiments at concentrations of 10 µM fluorescent labels; this concentration is 100-fold higher than the operational limit for standard TIRF experiments. We demonstrate the ease of implementing these probes into existing protocols by performing super-resolution imaging with DNA-PAINT, employing a fluorogenic 6 nt-long imager; through the faster acquisition of binding events, the imaging of viral genome segments could be sped up significantly to achieve extraction of 20-nm structural features with only ∼150 s of imaging. The exceptional tunability of our probe design will overcome concentration barriers in SMF experiments and unlock new possibilities in super-resolution imaging, molecular tracking, and smFRET.

Abstract Figure
Distinct Colitis-Associated Macrophages Drive NOD2-Dependent Bacterial Sensing and Gut Homeostasis
Authors: Katkar Gajanan D. , Anandachar Mahitha Shree , Ibeawuchi Stella-Rita , Espinoza Celia R. , Coates Jane , Malhotra Yashaswat S. , Mullick Madhubanti , Castillo Vanessa , Vo Daniella T. , Sahoo Debashis , Sinha Saptarshi , Ghosh Pradipta
Published: January 2025

Single-cell studies have revealed that intestinal macrophages maintain gut homeostasis through the balanced actions of reactive (inflammatory “accelerators”) and tolerant (non-inflammatory “brakes”) subpopulations. How these populations function in inflammatory bowel diseases (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), remains unresolved. Using multi-omics data and computational tools, here we define colon-specific macrophage states and reveal the critical role ofnon-inflammatorycolon-associatedmacrophages (ni-ColAMs) in IBD recovery. Through trans-scale analyses—integrating computational transcriptomics, proteomics, andin vivointerventional studies—we pinpointed GIV (CCDC88A), a heterotrimeric G-protein modulator, as a key regulator of ni-ColAMs. Notably, GIV emerged as the top-ranked gene in ni-ColAMs that physically and functionally interacts with nucleotide-binding oligomerization domain-containing protein 2 (NOD2), an innate immune sensor implicated in both CD and UC. Myeloid-specific depletion of GIV exacerbates infectious colitis, prolongs disease course, and abolishes the protective effects of muramyl dipeptide (MDP) in chemical colitis and sepsis models. GIV is required for NOD2 to dampen inflammation and efficiently clear microbes. GIV binds directly to NOD2’s terminal leucine-rich repeat (LRR#10) via its C-terminus. This interaction is disrupted in the CD-associated1007fsNOD2 variant, which lacks LRR#10, leading to impaired bacterial sensing and clearance. Findings illuminate how GIV and NOD2 collaborate to maintain gut homeostasis and reveal how derailment of such collaboration may drive dysbiosis and inflammation in IBD.
High-speed mapping of whole-mouse peripheral nerves at subcellular resolution
Authors: Shi Mei-Yu , Yao Yuchen , Wang Miao , Yang Qi , Ding Lufeng , Li Rui , Li Yuanyuan , Huang Haimeng , Yang Chao-Yu , Zhou Zhao , Zhu Zhenxiang , Wen Pengjie , Dai Fangling , Zeng Xiaohui , Zhang Ke-Ming , Guo Yuhong , Sun Zi-An , Xia Huanhuan , Ren Zhenhua , Cakmak Yusuf Ozgur , Zhang Ming , Xu Fuqiang , Qu Lei , Zhu Qingyuan , Lau Pak-Ming , Xu Cheng , Bi Guo-Qiang
Published: January 2025

In contrast to the rapid advancements in mesoscale connectomic mapping of the mammalian brain, similar mapping the peripheral nervous system has remained challenging due to the body’s size and complexity. Here, we present a high-speed blockface volumetric imaging system with an optimized workflow of whole-body clearing, capable of imaging the entire adult mouse at micrometer resolution within 40 hours. Three-dimensional reconstruction of individual spinal fibers in Thy1-EGFP mice reveals distinct morphological features of sensory and motor projections along the ventral and dorsal rami. Immunostaining facilitates body-wide mapping of sympathetic nerves and their branches, highlighting their perivascular patterns in limb muscles, bones, and most visceral organs. Viral tracing elucidates the fine architecture of vagus nerves and individual vagal fibers, revealing unexpected projection routes to various organs. Our approach offers an effective means to achieve a holistic understanding of cellular-level interactions among different systems that underlie body physiology and disease.

Highlights

Imaging of whole adult mouse at uniform subcellular resolution within 40 hours

Reconstruction of 191 thoracic spinal neuron projections for Thy1-EGFP mice

Mapping of immunolabeled sympathetic projections and their perivascular patterns

Resolving virus-labeled architecture of vagus nerves and complex single-fiber routes
Bioaugmentation in anaerobic digesters: A systematic review
Authors: Alipoursarbani Mozhdeh , Tideman Jeroen , López Mitzy , Abendroth Christian
Published: January 2025

Bioaugmentation, the intentional introduction of specific microorganisms into anaerobic digestion (AD) systems, has shown promise in enhancing methane production and in mitigating stressful conditions, particularly in systems operating below optimal performance. This review presents a systematic literature review (SLR) and meta-analysis to evaluate the efficacy of bioaugmentation strategies in AD. This review identified and analyzed studies meeting predefined eligibility criteria through a structured methodology involving research protocol, search, appraisal, synthesis, analysis, and reporting. A notable innovation of this review is its comprehensive critical comparison of different controls used in bioaugmentation studies, which has been inadequately addressed in previous literature. To facilitate the functional understanding, strains for bioaugmentation were grouped into the four phases of anaerobic digestion (hydrolysis, acidogenesis, acetogenesis and methanogenesis). A highly diverse set of microbes has been described for bioaugmentation, especially from the familiesClostridiaceae,PseudomonadaceaeandSyntrophomonadaceae. Most works are related to hydrolysis. The few works that address acidogenesis are mostly related to dark fermentation. Several studies used methanogenic archaea as well as syntrophic acetate oxidising bacteria, despite the difficulties in culturing them. On the other hand, studies applying strains for acetogenesis were largely underrepresented. Especially works on syntrophic propionate and butyrate oxidation (SPO and SBO) were missing.
Early-life paternal relationships predict adult female survival in wild baboons
Authors: Jansen David A.W.A.M. , Kinyua Warutere J. , Tung Jenny , Alberts Susan C. , Archie Elizabeth A.
Published: January 2025

Parent-offspring relationships can have profound effects on offspring behavior, health, and fitness in adulthood. These effects are strong when parents make heavy investments in offspring care. However, in some mammals, including several species of carnivores, rodents, and primates, fathers live and socialize with offspring, but paternal careper seis subtle or indirect. Do these limited father-offspring relationships also affect later-life outcomes for offspring? Working in a well-studied baboon population where males contribute little direct offspring care, we found that juvenile female baboons who had stronger paternal relationships, or who co-resided longer with their fathers, led adult lives that were 2-4 years longer than females with weak or short paternal relationships. This pattern did not differ between females who experienced high versus low levels of early-life adversity; hence, paternal relationships were not especially protective against harsh early environments. Males’ relationships were strongest with juvenile females they were most likely to have sired and when males had few current mating opportunities. Hence, father-daughter relationships may be constrained by male mating effort. Because survival predicts female fitness, fathers and their daughters may experience selection to engage socially and stay close in daughters’ early lives.
The brain dynamics of congenitally blind people seeing faces with sound
Authors: Matusz Pawel J. , Reich Lior , Stolz Louise A. , Retsa Chrysa , Tovar David A. , Striem-Amit Ella , Aggius-Vella Elena , Amedi Amir , Murray Micah M.
Published: January 2025

Sensory substitution devices (SSDs) convert images to sounds to equip blind individuals with nominally visual functions, like face or letter sensitivity. Prior studies showed that image-to-sound SSDs engage cortices ordinarily specialised for visual functions. However, the brain dynamics of SSD-supported perception remains unknown. Either visual cortices are the first locus of discrimination of SSD percepts, or their activation is a byproduct of perceptual processes unfurling elsewhere, such as in auditory cortices. Resolving this uncertainty is critical for understanding whether the blind truly “see” via SSDs. Using electrical neuroimaging of EEG data from congenitally blind adults, we show for the first time that inferotemporal visual cortices are the earliest site of face sensitivity when conveyed via SSDs. The blind do indeed “see” with SSDs. By providing the temporal dynamics of SSD perception, our findings provide unique evidence for the theory that cortices are characterised by task-contingent functional organisation.

Highlights

Sensory substitution devices (SSDs) convert information from one sense to another

SSDs can provide the blind with alternative access to visual information

Brain dynamics in visual cortices to SSD-conveyed object images are unknown

EEG electrical neuroimaging revealed face sensitivity within 500ms post-sound onset

Localized to fusiform regions, this face sensitivity shows the blind see with sound

eTOC blurb

We show for the first time that inferotemporal visual cortices are the earliest site of face sensitivity when conveyed via SSDs. The blind do indeed “see” with SSDs. Our findings of the temporal dynamics of SSD perception uniquely support the theory that cortices are characterised by task-selective sensory-independent functional organisation.
Dissecting the molecular basis underlying mycobacterial cell-wall hydrolysis by the catalytic domains ofD29LysA andDS6ALysA phage endolysins
Authors: Ceballos-Zúñiga Fernando , Gálvez-Larrosa Laura , Muñoz Inés G. , Infantes Lourdes , Pérez-Dorado Inmaculada
Published: January 2025

Mycobacteria encompass a broad range of microorganisms that cause infections with a significant impact on human health, resulting in millions of deaths each year. From tuberculosis and leprosy, caused byMycobacterium tuberculosisandMycobacterium leprae, respectively, to infections caused by emerging/opportunistic pathogens such asMycobacterium abscessus. The battle to combat this health burden is further challenged by limitations in the treatments currently available and the rise of antimicrobial resistance. This underscores the need for new therapeutic strategies to combat these infections. Mycobacteriophage LysA endolysins are complex, multi-domain peptidoglycan hydrolases with reported antimicrobial relevance and the potential to treat mycobacterial infections. However, despite the therapeutic prospects of LysAs, our understanding of their mechanism of action remains limited. This study provides a comprehensive structural-functional analysis of the catalytic domains of two LysA endolysins encoded by the bacteriophagesD29andDS6A, which are known to infect pathogenic mycobacteria, includingM. tuberculosis. As part of this work, we have characterized the four catalytic domains present in both endolysins (D29N4/D29GH19 andDS6AGH19/DS6AAmi2B) both alone and in complex with PG analogues. To achieve this, we combined protein engineering, X-ray crystallography, small-angle X-ray scattering, andin silicotools. To our knowledge, this has led to the first experimental structures reported for mycobacteriophage endolysins, which reveals key aspects of peptidoglycan binding and hydrolysis byD29LysA andDS6ALysA lysins, as well as other homologous LysAs, including the hydrolase domains similar to those examined here. Altogether, this represents a significant step forward in understanding how mycobacterial cell-wall hydrolysis occurs by this important class of endolysins and opens the door to their future use in therapeutic applications as enzybiotics. Information that will allow the rational design ofa la carteenzymes with optimized lytic properties against mycobacterial pathogens.
Functional recovery of adult brain tissue arrested in time during cryopreservation by vitrification
Authors: German Alexander , Akdaş Enes Yağız , Flügel-Koch Cassandra , Fejtova Anna , Winkler Jürgen , Alzheimer Christian , Zheng Fang
Published: January 2025

Cryopreserving adult brain tissue is challenging due to damage from ice formation, and traditional freezing methods fail to maintain neural architecture and function. Vitrification is a promising alternative to freezing, but its application to brain tissue has remained largely unexplored. Here, we demonstrate near-physiological recovery of adult mouse hippocampal slices after vitrification and rewarming. Key features of this cognitive hub are preserved, including structural integrity, metabolic responsiveness, neuronal excitability, and synaptic transmission and plasticity. Notably, hippocampal long-term potentiation was well preserved, indicating that the cellular machinery of learning and memory remains operational. These findings extend the knowledge of brain recovery after hypothermic shutdown to recovery after complete cessation of molecular mobility through vitrification. This suggests that brain tissue can be arrested in time and reactivated, opening avenues for potential clinical applications.

Teaser

Brain tissue is arrested in time by vitrification and reactivated, with neuronal and synaptic function preserved.
Old World alphaviruses use distinct mechanisms to infect brain microvascular endothelial cells for neuroinvasion
Authors: Alvarez Pablo A. , Tang Ashley , Winters Declan M. , Kaushal Prashant , Medina Angelica , Kaczor-Urbanowicz Karolina E. , Ramirez Reyes Bryan , Kaake Robyn M. , Fregoso Oliver I. , Pyle April D. , Bouhaddou Mehdi , Tang Hengli , Li Melody M.H.
Published: January 2025

Several alphaviruses bypass the blood-brain barrier (BBB), causing debilitating or fatal encephalitis. Sindbis virus (SINV) has been extensively studiedin vivoto understand alphavirus neuropathogenesis; yet the molecular details of neuroinvasion at the BBB remain poorly understood. We investigated alphavirus-BBB interactions by pairing a physiologically relevant, human pluripotent stem cell derived model of brain microvascular endothelial cells (BMECs) with SINV strains of opposite neuroinvasiveness. Our system demonstrates that SINV neuroinvasion correlates with robust infection of the BBB. Specifically, SINV genetic determinants of neuroinvasion enhance viral entry into BMECs. We also identify solute carrier family 2 member 3 (SLC2A3, also named GLUT3) as a potential BMEC-specific entry factor exploited for neuroinvasion. Strikingly, efficient BBB infection is a conserved phenotype that correlates with the neuroinvasive capacity of several Old World alphaviruses, including chikungunya virus. Here, we reveal BBB infection as a shared pathway for alphavirus neuroinvasion that can be targeted for preventing alphavirus-induced encephalitis.
Causal modeling of gene effects from regulators to programs to traits: integration of genetic associations and Perturb-seq
Authors: Ota Mineto , Spence Jeffrey P. , Zeng Tony , Dann Emma , Marson Alexander , Pritchard Jonathan K.
Published: January 2025

Abstract

Genetic association studies provide a unique tool for identifying causal links from genes to human traits and diseases. However, it is challenging to determine the biological mechanisms underlying most associations, and we lack genome-scale approaches for inferring causal mechanistic pathways from genes to cellular functions to traits. Here we propose new approaches to bridge this gap by combining quantitative estimates of gene-trait relationships from loss-of-function burden tests with gene-regulatory connections inferred from Perturb-seq experiments in relevant cell types. By combining these two forms of data, we aim to build causal graphs in which the directional associations of genes with a trait can be explained by their regulatory effects on biological programs or direct effects on the trait. As a proof-of-concept, we constructed a causal graph of the gene regulatory hierarchy that jointly controls three partially co-regulated blood traits. We propose that perturbation studies in trait-relevant cell types, coupled with gene-level effect sizes for traits, can bridge the gap between genetics and biology.
Protocol for a qualitative study exploring older adults’ experience of mental health and wellbeing in work-related later life transitions
Authors: Woodhouse Rebecca , McMillan Dean , Wadman Ruth
Published: January 2025

Later life transitions occur in the later years of life and can cause significant disruption to a person’s usual daily routine and lifestyle. Transitions that are specifically related to work, such as retirement, can cause sudden changes to an individual’s financial, social and psychological environment and for some, may negatively impact on mental health and wellbeing. This protocol outlines a qualitative study to explore the experiences of adults who have experienced a negative impact on their mental health or wellbeing due to a work-related transition in later life. Semi-structed interviews will be completed with people aged 50 and over to gain a deeper understanding of 1) the mental health and wellbeing of people undertaking later life transitions; 2) the context surrounding the transitions (such as due to ill-health or redundancy); and 3) the types of support they received or may have benefitted from. Research in this area is scarce and this study will help us to better understand the experience of those who have encountered such difficulties to help develop appropriate interventions and support in this area.
The essential role of 2,4-dienoyl-CoA reductase for degradation of complex fatty acid mixtures
Authors: Schiaffi Veronica , Pavoncello Viola , Barras Frédéric , Bouveret Emmanuelle
Published: January 2025

Fatty acids (FAs) can be used as carbon and energy source by most bacteria. FAs are very diverse and show variations in aliphatic chain length, degree and kind of branching, and number of double bonds. After their import in the cell and their activation by a thioester link to Coenzyme A, fatty acids are degraded by the β-oxidation machinery constituted of the Fad proteins. The core enzymes of the β-oxidation machinery can degrade most FAs, except for those FAs that bear an unsaturation at an even-numbered carbon. Such FAs include arachidonic acid or linoleic acid, which are essential FAs in the mammalian diet. We studied the role of the 2,4-dienoyl-CoA reductase FadH inE. colimetabolism of FAs. We showed thatfadHis essential for growth on linoleic acid and that Cys residues acting as ligands for FadH-bound [Fe-S] cluster are essential for activityin vivo. Moreover, we showed that when it is mixed with other FAs, linoleic acid prevents growth of thefadHmutant. These results suggest a specific metabolic burden imposed by linoleic acid degradation, which might prove important in the context of the natural gut environment with mixed FA composition. Eukaryotes also use 2,4-dienoyl-CoA reductases for β-oxidation of FAs in mitochondria but these eukaryotic enzymes belong to a different family than FadH, with different co-factors equipment and mechanism. Yet, we showed that eukaryotic 2,4-dienoyl-CoA reductases can complement theE. coli fadHmutant for growth on linoleic acid and for relief of linoleate mediated jamming of the β-oxidation. Altogether these studies demonstrate the key role of prokaryotic and eukaryotic 2,4-dienoyl-CoA reductases in a complex environment containing mixtures of saturated and unsaturated FAs, as found in the gut or in the surrounding of cancer cells.
Polygenic viral factors enable efficient mosquito-borne transmission of African Zika virus
Authors: Torii Shiho , Lord Jennifer S. , Lavina Morgane , Prot Matthieu , Lecuyer Alicia , Diagne Cheikh T. , Faye Oumar , Faye Ousmane , Sall Amadou A. , Bonsall Michael B. , Simon-Lorière Etienne , Montagutelli Xavier , Lambrechts Louis
Published: January 2025

Zika virus (ZIKV) is a mosquito-borne flavivirus primarily transmitted among humans byAedes aegypti. Over the past two decades, it has caused significant outbreaks associated with birth defects and neurological disorders. Phylogenetically, ZIKV consists of two main genotypes referred to as the African and Asian lineages, each exhibiting distinct biological properties. African lineage strains are transmitted more efficiently by mosquitoes, but pinpointing the genetic basis of this difference has remained challenging. Here, we address this question by comparing recent African and Asian strains using chimeric viruses, in which segments of the parental genomes are swapped. Our results show that the structural genes from the African strain enhance viral internalization, while the non-structural genes improve genome replication and infectious particle production in mosquito cells.In vivomosquito transmission is most significantly influenced by the structural genes, although no single viral gene alone determines this effect. Additionally, we develop a stochastic model ofin vivoviral dynamics in mosquitoes that mirrors the observed patterns, suggesting that the primary difference between the African and Asian strains lies in their ability to traverse the mosquito salivary glands. Overall, our findings suggest that the polygenic nature of ZIKV transmissibility has prevented Asian lineage strains from achieving the same epidemic potential as African lineage strains, underscoring the importance of lineage-specific adaptive landscapes in shaping ZIKV evolution and emergence.
Rdh10-mediated Retinoic Acid Signaling Regulates the Neural Crest Cell Microenvironment During ENS Formation
Authors: Butler Tjaden Naomi E. , Shannon Stephen R. , Seidel Christopher W. , Childers Melissa , Aoto Kazushi , Sandell Lisa L. , Trainor Paul A.
Published: January 2025

The enteric nervous system (ENS) is formed from vagal neural crest cells (NCC), which generate most of the neurons and glia that regulate gastrointestinal function. Defects in the migration or differentiation of NCC in the gut can result in gastrointestinal disorders such as Hirschsprung disease (HSCR). Although mutations in many genes have been associated with the etiology of HSCR, a significant proportion of affected individuals have an undetermined genetic diagnosis. Therefore, it’s important to identify new genes, modifiers and environmental factors that regulate ENS development and disease. Rdh10 catalyzes the first oxidative step in the metabolism of vitamin A to its active metabolite, RA, and is therefore a central regulator of vitamin A metabolism and retinoic acid (RA) synthesis during embryogenesis. We discovered thatretinol dehydrogenase 10(Rdh10) loss-of-function mouse embryos exhibit intestinal aganglionosis, characteristic of HSCR. Vagal NCC form and migrate inRdh10mutant embryos but fail to invade the foregut.Rdh10is highly expressed in the mesenchyme surrounding the entrance to the foregut and is essential between E7.5-E9.5 for NCC invasion into the gut. Comparative RNA-sequencing revealed downregulation of theRet-Gdnf-Gfrα1gene signaling network inRdh10mutants, which is critical for vagal NCC chemotaxis. Furthermore, the composition of the extracellular matrix through which NCC migrate is also altered, in part by increased collagen deposition. Collectively this restricts NCC entry into the gut, demonstrating thatRdh10-mediated vitamin A metabolism and RA signaling pleiotropically regulates the NCC microenvironment during ENS formation and in the pathogenesis of intestinal aganglionosis.
Large-scale genomic analysis reveals the distribution and diversity of Type VI Secretion Systems inEscherichia coli
Authors: Nesporova Kristina , Govers Sander K.
Published: January 2025

The type VI secretion system (T6SS) is a versatile nanomachine that injects effectors into target cells, playing a role in bacterial competition and virulence. While widespread in Gram-negative bacteria, T6SS prevalence varies across species and strains, and its distribution inEscherichia coliremains underexplored despiteE. colibeing an important intestinal (IPEC) and extraintestinal (ExPEC) pathogen. Here, we examined the prevalence and distribution of T6SS subclasses (T6SSi) across 131610E. coligenomes, which we annotated for clinical relevance. T6SS genes were identified while focusing on the three subclasses of T6SSigene clusters known to be present inE. coli:T6SSi1, T6SSi2, and T6SSi4b. Across phylogenetic groups, T6SSi1was broadly present, while T6SSi2showed associations with B1, B2, and G, and T6SSi4bwas rare. T6SSi1was primarily associated with IPEC, and T6SSi2with ExPEC. Even clearer patterns emerged at the sequence type (ST) level or when strains were further categorized into pathovars. For example, both the most dominant ExPEC and IPEC ST (ST131 and ST11, respectively) displayed niche-specific T6SS trends, with non-complete T6SSs being more associated with humans. In a final step, we evaluated the co-occurrence of T6SSs with other virulence-associated genes (VAGs) and multidrug resistance (MDR). This analysis confirmed the association of T6SSi1and T6SSi2with IPEC- and ExPEC-associated VAGs, respectively, and revealed a negative correlation between complete T6SSisubclasses and MDR. Together, our work provides a comprehensive overview of the diversity of T6SSs acrossE. coli, shedding more light on their potential contribution to pathogenicity in this species.

Importance

Our study represents a large-scale analysis of T6SSs across one of the most comprehensive collections ofE. coligenomes to date. In doing so, we updated several misconceptions on T6SSs distribution and other genomic properties ofE. colistrains, which originated from smaller scale studies and were subsequently extrapolated in the literature. This includes the prevalence and distribution of T6SSisubclasses across phylogenetic groups (e.g., T6SSi2is not prevalent in phylogroup D), the association of specific virulence factors with IPEC and/or ExPEC (e.g., hemolysin A is more often associated with IPEC and not a typical ExPEC characteristic), and characteristics of pathogenic STs (e.g., ST131 displays distinct genomic properties based on its environmental niche). As such, this study not only advances our understanding of T6SS inE. coli, but also serves as a valuable resource for future studies on the clinical relevance and distribution of other genetic elements.
Convergent Agonist and Heat Activation of Nociceptor TRPM3
Authors: Kumar Sushant , Jin Fei , Park Sung Jin , Choi Wooyoung , Keuning Sarah I. , Massimino Richard P. , Vu Simon , Lü Wei , Du Juan
Published: January 2025

Detecting noxious heat is vital for survival, triggering pain responses that protect against harm1,2. The TRPM3 channel is a key nociceptor for sensing noxious heat and a promising therapeutic target for pain treatment and neurological disorders such as epilepsy3–11. Here, we functionally and structurally characterized TRPM3 in response to diverse stimuli: the synthetic superagonist CIM0216Ref12, the anticonvulsant antagonist primidone13,14, and heat1,10,15. Our findings reveal that TRPM3 is intrinsically dynamic, with its intracellular domain (ICD) sampling both resting and activated states, though strongly favoring the resting state without stimulation. CIM0216 binds to the S1–S4 domain, inducing conformational changes in the ICD and shifting the equilibrium toward activation. Remarkably, heat induces similar ICD rearrangements, revealing a converged activation mechanism driven by chemical compounds and temperature. This mechanism is supported by functional data showing that mutations facilitating the ICD movement markedly increase the sensitivity of TRPM3 to both chemical and thermal signals. These findings establish a critical role of the ICD in temperature sensing in TRPM3, a mechanism likely conserved across the TRPM family. Finally, we show that primidone binds to the same site as CIM0216 but acts as an antagonist. This study provides a framework for understanding the thermal sensing mechanisms of temperature-sensitive ion channels and offers a structural foundation for developing TRPM3-target therapeutics for pain and neurological disorders.
Single cell Edit Detection and Identification Tool (scEDIT): computational workflow for efficient and economical single cell analysis of CRISPR edited cells
Authors: Suryawanshi Gajendra W
Published: January 2025

Recent advances in single-cell DNA sequencing (scDNA-seq) and CRISPR technology have revolutionized gene therapy and drug discovery. However, data analysis requires expensive high-performance computing (HPC) clusters or large data servers, limiting reanalysis due to the lack of open-source software. To address this, we present scEDIT, a fast, lightweight, portable, and standalone software for pre- and post-processing CRISPR editing data from the Tapestri single-cell DNA-seq platform. scEDIT is memory-efficient, multithreaded, and compatible with most UNIX based systems. Tests using a low-cost desktop and public single cell CRISPR data demonstrate that the tool can efficiently process raw sequences, identify cell barcodes, count unedited and edited amplicons per cell, and outputs detailed filtered reads. Analysis of the single cell CRISPR data reveals indel patterns shared between in vitro experiments and unique indel profiles detected for in vivo study. Results further demonstrate the ability of single cell analysis in providing quantitative insights into the true zygosity of edited cell population. Although data shows a linear relation between indel frequencies by read count and cell count details of indel share between difference cells can only be truly explored with single cell data. The efficiency, stability, and portability of scEDIT makes it an invaluable tool for uncovering new insights into the single cell data without requiring expensive computational resources.
The microbiome of Calasterella californica is shaped mostly by the genetics of the host thalli
Authors: González-Ramírez Ixchel , Song Michael J. , Mishler Brent D.
Published: January 2025

Premise of the study: Plant microbiomes have been shown to affect essential life processes like plant growth, defense mechanisms, and stress responses. Much of the work done on this topic has focused on vascular plants and model organisms. Liverwort biology is very different from that of vascular plants, making generalizations difficult. Here we characterize the microbiome of the liverwort Calasterella californica and explore whether climatic, geographic, and genetic factors are potential drivers of the microbiome composition. Methods: We utilized the genomic data obtained through the California Conservation Genomics Project, which densely sampled individuals of C. californica across California. We selected 34 samples and utilized the MetaPhlAn pipeline to characterize the composition of the microbial communities for each sample. Key results: We characterize the core microbiome of Calasterella, which is dominated by Methylobacterium. There is a weak association between the relative abundance of bacteria in the Hyphomicrobiales and liverwort size. Genetic distance was the most important factor in shaping beta diversity of the Calasterella californica microbiome. Environmental distance was only weakly predictive and geographic distance was not significant. Conclusions: Leveraging genomic data obtained from field samples of a liverwort species allowed us to characterize the microbial diversity of C. californica and its variation across space. There is a group of taxa that are consistently associated with this plant. And the variation observed across samples seems to be linked to the genetic similarity of the liverworts.
An Open-Source Joystick Platform for Investigating Forelimb Motor Control, Auditory-Motor Integration, and Value-Based Decision-Making in Head-Fixed Mice
Authors: Linares-García Ivan , Iliakis Evan A. , Juliani Sofia E. , Ramirez Alexandra N. , Woolley Joel , Díaz-Hernández Edgar , Fuccillo Marc V. , Margolis David J.
Published: January 2025

Investigation of neural processes underlying motor control requires behavioral readouts that capture the richness of actions, including both categorical (choice-based) information and motor execution (kinematics). We present an open-source platform for behavioral training of head-fixed mice that combines a stationary or retractable forelimb-based joystick, sound-presentation system, capacitive lick sensor, and water reward dispenser. The setup allows for the creation of multiple behavioral paradigms, two of which are highlighted here: a two-alternative forced-choice auditory-motor discrimination paradigm, and a two-armed bandit value-based decision-making task. In the auditory-motor paradigm, mice learn to report high or low frequency tones by pushing or pulling the joystick. In the value-based paradigm, mice learn to push or pull the joystick based on the history of rewarded trials. In addition to reporting categorical choices, this setup provides a rich dataset of motor parameters that reflect components of the underlying learning and decision processes in both of these tasks. These kinematic parameters (including joystick speed and displacement, Fréchet similarity of trajectories, tortuosity, angular standard deviation, and movement vigor) provide key additional insights into the motor execution of choices that are not as readily assessed in other paradigms. The system’s flexibility of task design, joystick readout, and ease of construction represent an advance compared to currently available manipulandum tasks in mice. We provide detailed schematics for constructing the setup and protocols for behavioral training using both paradigms, with the hope that this open-source resource is readily adopted by neuroscientists interested in mechanisms of sensorimotor integration, motor control, and choice behavior.

Significance Statement

Behavioral paradigms for experiments in head-restrained mice are important for investigating the relationship between neural activity and behavior. However, behavioral setups are often constrained by high cost, design complexity, and implementation challenges. Here, we present an open-source platform for behavioral training of head-fixed mice using a joystick manipulandum. The setup allows for the creation of multiple behavioral paradigms, including an auditory-motor discrimination paradigm, and a value-based decision-making task. We include detailed instructions for construction and implementation of the entire open-source behavioral platform.
The mechanism of Micafungin action to Pteropine orthoreovirus infection in human cell line
Authors: Bubphasook Wirayatida , Iida Atsuo , Hondo Eiichi
Published: January 2025

Pteropine orthoreovirus (PRV) is a fusogenic virus carried by bats that causes respiratory illnesses in humans. PRVs are transmitted from bats to humans and among humans. Micafungin (MCFG), an approved drug for fungal treatment, shows potential in inhibiting PRV propagation, although its precise mechanism of action remains unclear. This study aimed to investigate molecular mechanisms of MCFG during PRV propagation. Initially, molecular docking was employed to predict the primary target of MCFG. It was the p17 protein of PRV. Prior to examining the effect of MCFG on p17 suppression, differential gene expression analysis was performed to compare PRV-infected host cells with MCFG-treated cells, and it was found that IL-6 should be the main regulator induced by MCFG. Silencing IL-6 with siRNA markedly induced PRV release, syncytial formation, and marginally enhanced PRV RNA replication, corresponding to the main suppressive effects of MCFG against PRV. The p17, the presumed suppression target to inhibit syncytial formation by MCFG, markedly reduced syncytial formation but did not influence viral RNA replication. In contrast, MCFG significantly suppressed syncytial formation and slightly reduced PRV RNA replication, although both MCFG and anti-p17 antibody increased IL-6 mRNA expression. MCFG may inhibit other PRV proteins, like nonstructural replication protein (sigmaNS), which were found by molecular docking study. In conclusion, MCFG primarily targets p17 and modulates host immunity through IL-6, which likely interferes directly with syncytial formation.
Evaluating effects of antibiotics across preclinical models of the human gastrointestinal microbiota
Authors: Auchtung Thomas A. , Lerma Armando I. , Schuchart Keegan , Auchtung Jennifer M.
Published: January 2025

While antibiotics play important roles in treating infections, disruption of the gastrointestinal microbiota during antibiotic treatment can lead to negative health consequences. However, for many antibiotics, the spectrum of activity has been determined for select isolates rather than for the range of microbes that populate the gastrointestinal tract. Here, we examined the response of communities of gastrointestinal microbes to antibiotics using two different model systems, human fecal minibioreactors and human microbiota associated mice. Communities established in minibioreactors using 12 different fecal donors were exposed to 12 different classes of antibiotics. Samples from three fecal donors were used to colonize germ-free mice from three different genetic backgrounds; progeny mice were then exposed to 6 of 12 antibiotics tested in minibioreactors. Initial bacterial community diversity was dependent on both the fecal donor and model system. Antibiotics affected a wide range of taxa across the phylogenetic spectrum, with many taxa similarly affected across treatments with different classes of antibiotics. Vancomycin, typically administered to treat Gram-positive bacterial infections, decreased the abundance of diverse taxa, including Gram-negative Bacteroidota species. Effects on some taxa were restricted by model system, indicating the importance of environmental context on antibiotic susceptibility. Altogether, these results indicate the complex interrelationships between microbiota composition and environmental context on antibiotic susceptibility and demonstrate strengths and weaknesses of each pre-clinical model system for evaluating effects of new antibiotics and other compounds with potential to disrupt the microbiota.
HIV-1 subtype C LTR Sp1IIIT5A mutant enhances transcription activity and Sp1 binding affinity
Authors: Msthali Nomcebo , Kehinde Ibrahim , Khan Rene , Soliman Mahmoud , Ndung’u Thumbi , Madlala Paradise
Published: January 2025

Background

Genetic variation within HIV-1 subtype C (HIV-1C) long terminal repeat (LTR) transmitted/founder viruses influences transcription activation and clinical disease outcomes. The role of specific mutations such as thymine-to-adenine (T5A) mutation at position five of the Specificity protein 1 (Sp1) III motif (Sp1IIIT5A) remains underexplored. This study investigates the impact of Sp1IIIT5A on HIV-1C LTR transcription activity and Sp1 binding affinity.

Methods

The Sp1IIIT5A mutant and consensus HIV-1C LTR sequences were cloned into the pGL3 Luciferase Basic reporter vector, sequenced, and transfected into SVG and Jurkat cell lines, independently. Transcription activity and Sp1 expression were assessed via luciferase assays and Western blot. Structural models of Sp1IIIT5A, consensus LTRs and Sp1 were generated, and docking scores calculated using HDOCK, HADDOCK, and pyDockDNA. Molecular dynamics simulations analyzed stability and interactions of Sp1IIIT5A LTR-Sp1 complexes.

Results and Discussion

The Sp1III5A mutant significantly increased basal (SVG: p<0.0001; Jurkat: p=0.0052) and Tat-mediated (SVG and Jurkat: p<0.0001) HIV-1C LTR transcription activity in both cell lines, with stronger effects in SVG cells. Sp1 expression levels remained similar across cell lines (p=0.0814). Sp1III5A exhibited higher binding affinity (-332.7, -174.6, and -279.2 kcal/mol) than the canonical sequence (-311.4, -157.0, and -247.3 kcal/mol).

Conclusion

The Sp1IIIT5A mutation significantly enhances HIV-1C LTR transcription activity and Sp1 binding affinity, indicating its potential tole in modulating HIV-1C transcription and pathogenesis. Further investigation is needed to elucidate its impact on HIV-1C latency.

Importance

In this study we show that the thymine-to-adenine (T5A) mutation at position five of the Sp1 III motif (Sp1IIIT5A) within the HIV-1 subtype C (HIV-1C) long terminal repeat (LTR) increases viral transcription. This mutation enhances the interaction between HIV-1C and the cellular transcription factor Sp1, promoting the viral strain’s ability to replicate. Our findings provide insight into why certain HIV-1C strains behave differently, potentially leading to heterogenous rates of disease progression. Understanding the Sp1IIIT5A mutation could lead to improved strategies for controlling HIV-1C and developing cure strategies to clear the infection or result in virus remission.
Unravel the start element and promoter architecture across the domain Bacteria
Authors: Kuo Syue-Ting , Chang Joshua Kevin , Chang Clara , Shen Wei-Yi , Hsu Christine , Lai Sheng-Wen , Chou Hsin-Hung David
Published: January 2025

Promoters comprise multiple elements to modulate transcription. Despite decades of research, significant sequence and length variation of promoter elements renders elucidation of promoter function and the evolutionary diversity of transcriptional regulation challenging. Combining massively parallel assays with biophysical modeling, we mapped each promoter element across 49 phylogenetically diverse bacterial genomes (GC%: 27.8-72.1). We discovered a conserved promoter element, ‘start,’ that dictates transcription start site selection. We uncovered broad sequence conservation in the underappreciated spacer element, affecting transcription by up to 600-fold. We showed the discriminator element conserved in major bacterial clades Terrabacteria but variable in Gracilicutes. High sequence variation in Gracilicutes’ discriminator element resulted from diversifying evolution, enabling coordination of global gene expression with cellular growth rate through differential regulation of individual promoters. Identifying sequence and functional similarity between bacterial promoter elements and their archaeal and eukaryotic counterparts strongly suggests a shared evolutionary origin of promoter architecture.
Direct free energy calculation from
ab initio
path integral Monte Carlo simulations of warm dense matter
Authors: Dornheim Tobias , Moldabekov Zhandos A. , Schwalbe Sebastian , Vorberger Jan
Published: January 2025

We carry out highly accurate path integral Monte Carlo simulations to directly estimate the free energy of various warm dense matter systems including the uniform electron gas and hydrogen without any nodal restrictions or other approximations. Since our approach is based on an effective ensemble in a bosonic configuration space, it does not increase the computational complexity beyond the usual fermion sign problem. Its application to inhomogeneous cases such as an electronic system in a fixed external ion potential is straightforward and opens up the enticing possibility to benchmark density functional theory and other existing methods. Finally, it is not limited to warm dense matter and can be applied to a gamut of other systems such as ultracold atoms and electrons in quantum dots.

Published by the American Physical Society
2025
Implication of preheating on gravity assisted baryogenesis in
R2
-Higgs inflation
Authors: Cado Yann , Englert Christoph , Modak Tanmoy , Quirós Mariano
Published: January 2025

We investigate the impact of preheating on baryogenesis in R2-Higgs inflation. In this scenario, the inclusion of a dimension-six operator (R/Λ2)BμνB˜μν abundantly generates helical hypermagnetic fields during inflation, leading to a baryon asymmetric Universe at the electroweak crossover. Focusing on the R2-like regime, we first derive the relevant dynamics of preheating using a doubly covariant formalism. We find that preheating can happen for the Higgs, transverse gauge, and Goldstone bosons, however, it is dependent on the value of the nonminimal coupling ξH between the Standard Model Higgs field and the Ricci scalar. We identify the preheating temperature to determine the appropriate scale Λ for driving baryogenesis, which is around Λ∼2.2(2.6)×10−5MP for ξH∼1(10). Our results represent the most accurate estimation of the scale of gravity induced baryogenesis in R2-Higgs inflation to date. Areas for further improvement are identified.

Published by the American Physical Society
2025