bluuebunny/crossref_metadata_2025 huggingface dataset:
The Lipocone Superfamily: A Unifying Theme In Metabolism Of Lipids, Peptidoglycan And Exopolysaccharides, Inter-Organismal Conflicts And Immunity
Authors: Burroughs A. Maxwell , Nicastro Gianlucca G. , Aravind L
Published: January 2025

Wnt proteins are critical signaling molecules in developmental processes across animals. Despite intense study, their evolutionary roots have remained enigmatic. Using sensitive sequence analysis and structure modeling, we establish that the Wnts are part of a vast assemblage of domains, the Lipocone superfamily, defined here for the first time. It includes previously studied enzymatic domains like the phosphatidylserine synthases (PTDSS1/2) and the TelC toxin domain fromStreptococcus intermedius, the enigmatic VanZ proteins, the animal Serum Amyloid A (SAA) and a further host of uncharacterized proteins in a total of 30 families. Though the metazoan Wnts are catalytically inactive, we present evidence for a conserved active site across this superfamily, versions of which are consistently predicted to operate on head groups of either phospholipids or polyisoprenoid lipids, catalyzing transesterification and phosphate-containing head group severance reactions. We argue that this superfamily originated as membrane proteins, with one branch (including Wnt and SAA) evolving into soluble versions. By comprehensively analyzing contextual information networks derived from comparative genomics, we establish that they act in varied functional contexts, including regulation of membrane lipid composition, extracellular polysaccharide biosynthesis, and biogenesis of bacterial outer-membrane components, like lipopolysaccharides. On multiple occasions, members of this superfamily, including the bacterial progenitors of Wnt and SAA, have been recruited as effectors in biological conflicts spanning inter-organismal interactions and anti-viral immunity in both prokaryotes and eukaryotes. These findings establish a unifying theme in lipid biochemistry, explain the origins of Wnt signaling and provide new leads regarding immunity across the tree of life.
Decoding MASLD Progression: A Molecular Trajectory-Based Framework for Modelling Disease Dynamics
Authors: Kamzolas Ioannis , Koutsandreas Thodoris , Barker Charlie George , Vathrakokoili Pournara Anna , Weston Harry , Vacca Michele , Papatheodorou Irene , Vidal-Puig Antonio , Petsalaki Evangelia
Published: January 2025

Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD) has emerged as a silent pandemic, affecting nearly one-third of the global population. This condition encompasses a spectrum of liver disorders, ranging from simple steatosis to Metabolic Dysfunction-Associated Steatohepatitis (MASH), which is characterised by liver steatosis, lipotoxicity, hepatocellular damage, inflammation, and fibrosis. Left unchecked, MASLD/MASH can progress to cirrhosis and hepatocellular carcinoma. Despite the progressive nature of MASLD/MASH, current research primarily relies on static, histopathologically defined stages, which fail to capture the dynamic disease continuum.

In this study, we present an integrative stratification approach that combines patient pseudo-temporal ordering, network analysis, and cell-type deconvolution to map the continuous disease trajectory. By analyzing transcriptomic profiles, we predict patients’ positions along this trajectory, moving beyond conventional stage-based classifications. This approach reveals the sequence of critical molecular events driving MASLD/MASH progression, providing new insights into the disease’s pathophysiology. Furthermore, we identify novel trajectory-specific biomarkers that support a more refined, personalised strategy for managing MASLD. This work highlights the potential of trajectory-based frameworks in advancing our understanding and treatment of complex metabolic diseases.
Cell Type-Specific and Diabetic Kidney Disease-Associated Expression of Long Non-Coding RNAs in Human Kidneys
Authors: de Klerk Juliette A , Slieker Roderick C , Parker Wilson C , Wu Haojia , Muto Yoshiharu , Postma Rudmer J , ’t Hart Leen M , Peerlings Janneke HD , Herrewijnen Floris , Song Heein , Spijker H Siebe , Dumas Sébastien J , Koning Marije , van der Pluijm Loïs AK , Baelde Hans J , Gerrits Tessa , Rotmans Joris I , van Zonneveld Anton Jan , van Solingen Coen , Humphreys Benjamin D , Bijkerk R
Published: January 2025

Background

Long non-coding RNAs (lncRNAs) play essential roles in cellular processes, often exhibiting cell type-specific expression and influencing kidney function. While single-cell RNA sequencing (scRNA-seq) has advanced our understanding of cellular specificity, past studies focus solely on protein-coding genes. We hypothesize that lncRNAs, due to their cell-specific nature, have crucial functions within particular renal cells and thereby play essential roles in renal cell function and disease.

Methods

Using single-nucleus RNA sequencing (snRNA-seq) data from kidney samples of five healthy individuals and six DKD patients, we explored the non-coding transcriptome. Cell type-specific lncRNAs were identified, and their differential expression in DKD was assessed. Integrative analyses included expression quantitative trait loci (eQTL), genome-wide association studies (GWAS) for estimated glomerular filtration rate (eGFR), and gene regulatory networks. Functional studies focused onTARID, a lncRNA with podocyte-specific expression, to elucidate its role in podocyte health.

Results

we identified 349 lncRNAs with cell type-specific expression across kidney cell types. Of these, 104 lncRNAs were differentially expressed in DKD. Integrative analyses, including eQTL data, GWAS results for eGFR and gene regulatory networks, pinpointedTARID, a podocyte-specific lncRNA, as a key candidate upregulated in DKD. Functional studies confirmedTARID’s podocyte-specific expression and revealed its central role in actin cytoskeleton reorganization, a critical process in podocyte health.

Conclusions

Our study provides a comprehensive resource of single-cell lncRNA expression in the human kidney and highlights the importance of cell type-specific lncRNAs in kidney function and disease. Specifically, we demonstrate the functional relevance ofTARIDin podocyte health. This work underscores the utility of integrating scRNA-seq with functional genomics to uncover novel regulatory mechanisms in kidney biology.

Key points

This provides a resource for kidney (cell type-specific) lncRNA expression and demonstrates the importance of lncRNAs in renal health.

We identified 349 cell type-specific lncRNAs in the human kidney, with 104 showing altered expression in diabetic kidney disease (DKD).

TARID, a podocyte-specific lncRNA upregulated in DKD, is crucial for actin cytoskeleton reorganization in podocytes.
Spatial map of native duct cell populations in human pancreas and their representation in pancreatic cancers
Authors: Van den Bossche JL , Van der Vliet M , Michiels E , Senar O Azurmendi , Madran Z , Coolens K , Van Lint S , Nacher M , Arsenijevic T , Messaoudi N , Lefesvre P , Montanya E , Rovira M , Van Laethem J , Baldan J , Houbracken I , Rooman I
Published: January 2025

Background & Aims

Epithelial tumors generally resemble the cellular architecture of their tissue of origin. However, this link remains largely unexplored in the pancreas.

Methods

Using Nanostring GeoMx DSP®, Resolve Molecular Cartography® and Nanostring CosMx®, and integration with single cell RNAseq datasets, we mapped the human pancreatic ductal epithelium in non-neoplastic pancreas, and compared it to pancreatic cancer subtypes.

Results

Groups of Keratin-5+cells among the Pan-Cytokeratin+cells in the duct have a gene signature reminiscent of stem cells and (supra)basal cells from other tissues. In spatial analysis at single cell resolution, the pancreatic duct manifests as a stratified epithelium comprising a basal and four luminal populations; In large ducts, KRT5+basal (BAS) cells express ΔNp63 while KRT5+luminal (LUM)-B cells reside supra-basally and are distinct from the common KRT5-LUM-A cells. LUM-C and LUM-D cells pertain to intercalated ducts and ductal glands, respectively. LUM-A and -C cells express gel-forming mucins while LUM-B cells have membrane-bound MUC4 and MUC16. In cancer, BAS and LUM-B signatures associate with the basal-like pancreatic ductal adenocarcinoma (PDAC) and correlate with lower survival but exhibit a mixed spatial pattern with a diffracted gene signature. In contrast, adenosquamous cancers of the pancreas (ASCP) preserve the normal spatially unmixed identity of LUM-B cells and BAS cells that is regulated by ΔNp63. Next to ΔNp63, conserved drug targets were identified for both populations.

Conclusion

This study offers a refined pancreatic tumor classification based on the native ductal architecture, with better preservation of the LUM-B and BAS cell population identity in ASCP than in PDAC, including the conserved drug targets.
The Single-Cell Landscape of Peripheral and Tumor-infiltrating Immune Cells in HPV- HNSCC
Authors: Galvani Rômulo Gonçalves Agostinho , Rojas Adolfo , Matuck Bruno F. , Rupp Brittany T. , Kumar Nikhil , Huynh Khoa , de Biagi Junior Carlos Alberto Oliveira , Liu Jinze , Sheth Siddharth , Maria Krol Jelte Martinus , Maracaja-Coutinho Vinicius , Byrd Kevin Matthew , Severino Patricia
Published: January 2025

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. HPV-negative HNSCC, which arises in the upper airway mucosa, is particularly aggressive, with nearly half of patients succumbing to the disease within five years and limited response to immune checkpoint inhibitors compared to other cancers. There is a need to further explore the complex immune landscape in HPV-negative HNSCC to identify potential therapeutic targets. Here, we integrated two single-cell RNA sequencing datasets from 29 samples and nearly 300,000 immune cells to investigate immune cell dynamics across tumor progression and lymph node metastasis. Notable shifts toward adaptative immune cell populations were observed in the 14 distinct HNSCC-associated peripheral blood mononuclear (PBMCs) and 21 tumor-infiltrating immune cells (TICs) considering disease stages. All PBMCs and TICs revealed unique molecular signatures correlating with lymph node involvement; however, broadly, TICs increased ligand expression among effector cytokines, growth factors, and interferon-related genes. Pathway analysis comparing PBMCs and TICs further confirmed active cell signaling among Monocyte-Macrophage, Dendritic cell, Natural Killer (NK), and T cell populations. Receptor-ligand analysis revealed significant communication patterns shifts among TICs, between CD8+ T cells and NK cells, showing heightened immunosuppressive signaling that correlated with disease progression. In locally invasive HPV-negative HNSCC samples, highly multiplexed immunofluorescence assays highlighted peri-tumoral clustering of exhausted CD8+ T and NK cells, alongside their exclusion from intra-tumoral niches. These findings emphasize cytotoxic immune cells as valuable biomarkers and therapeutic targets, shedding light on the mechanisms by which the HNSCC sustainably evades immune responses.
Age-Associated Decline in Autophagy Pathways in Retinal Pigment Epithelium and Protective Effects of Topical Trehalose in Light-induced Outer Retinal Degeneration in Mice
Authors: Cox Katherine , Shi Gongyu , Read Neve , Ou Kepeng , Liu Zijia , Wu Jiahui , Cendanawati Suci , Powell Jenna Le Brun , Nicholson Lindsay B. , Dick Andrew D. , Liu Jian
Published: January 2025

Age is a primary risk factor for chronic conditions, including age-related macular degeneration (AMD). Impairments in autophagy processes are implicated in AMD progression, but the extent of autophagy’s contribution and its therapeutic potential remain ambiguous. This study investigated age-associated transcriptomic changes in autophagy pathways in the retinal pigment epithelium (RPE) and evaluated the protective effects of topical trehalose, an autophagy-enhancing small molecule, against light-induced outer retinal degeneration in mice. Transcriptomic analysis of human RPE/choroid and mouse RPE revealed consistent downregulation of autophagy pathways with age, alongside variable changes as AMD severity progressed. Given the age- and AMD-associated perturbation of autophagy pathways, we examined trehalose treatmentin vitro, which enhanced autophagic flux and restored mitochondrial respiratory function in primary murine RPE cells exposed to oxidative stress.In vivo, topical trehalose improved autophagy-lysosome activity in mouse RPE, demonstrated by elevated LC3B turnover and SQSTM1/p62 degradation. Furthermore, trehalose eyedrops protected mice from light-induced damage to the RPE and photoreceptors, preserving outer nuclear layer thickness, RPE morphology, and junctional F-actin organization. Taken together, the data support that age-related decline and severe dysregulation in autophagy contributed to AMD progression. By restoring autophagic flux, topical trehalose demonstrates therapeutic potential to address early autophagy-related pathological changes in AMD.
Investigating antiviral pathways in Atlantic salmon cells through interferon receptor knockouts via CRISPR-Cas9
Authors: Reza Mohammad Ali Noman , Harvey Thomas Nelson , Ahmed Abdelrahim Gamil Amr , Evensen Øystein , Gillard Gareth Benjamin , Sandvik Guro Katrine
Published: January 2025

In Atlantic salmon (Salmo salar), infectious salmon anemia virus (ISAV) and infectious pancreatic necrosis virus (IPNV) evade host immune response through complex antagonistic mechanisms. Type I interferons (IFNs) play a pivotal role in antiviral defense by signaling through heterodimeric receptors to activate the JAK-STAT pathway and drives the expression of interferon-stimulated genes (ISGs). In this study, CRISPR-Cas9 was used to knock out (KO) interferon receptor genes (crfb1a,crfb5a,il10rb,ifngr2a) and a combined group of candidate receptors (crfb1a,crfb5a,il10rb,ifngr2a,il10r2) to investigate their roles and their impact on downstream signaling cascades with RNA sequencing. Recombinant IFNa was used to induce an antiviral state before challenging cells with ISAV and IPNV.

The knockouts significantly disrupt downstream antiviral signaling, with two knockouts,crfb1aandcrfb5a, showing pronounced effects. During ISAV infection, thecrfb1aKO group exhibited a marked reduction in the expression of critical signaling genes such asstat1b, stat2, stat6,andirf3during ISAV infection, whileirf7was upregulated during IPNV infection. Thecrfb5aKO group exhibited reducedstat2expression in ISAV infection and upregulatedirf7during IPNV infection. Despite these disruptions, ISGs such asMxandisg15maintained their expression levels across all knockout groups, suggesting potential alternative signaling pathways. Pathway analysis further revealed upregulation of cellular processes like actin regulation and phagosome activity, which may compensate for impaired immune signaling. These findings highlight the distinct roles of IFN receptor genes in mediating antiviral responses and underscore the complexity of IFN signaling in Atlantic salmon.
Admixture and environmental fluctuations shape the evolutionary history of a predator radiation in East Africa’s Lake Tanganyika
Authors: Rick Jessica A. , Junker Julian , Lewanski Alexander L. , Swope Brittany , McGlue Michael M. , Sweke Emmanuel A. , Kimirei Ismael A. , Seehausen Ole , Wagner Catherine E.
Published: January 2025

Top predators have oversized impacts on food webs and ecosystem dynamics, and introducing a novel predator to a naive environment can have dramatic consequences for endemic biodiversity. Lake Tanganyika is unique among African lakes in the diversity of the pelagic top predators in the genusLates, where four species are endemic to the lake. Using a combination of reduced-representation and whole genome resequencing data, and pairing these with phylogenetic and demographic modeling approaches, we find thatLatescolonization of Lake Tanganyika was much more recent (∼1–2 Mya) than other major and diverse clades within the lake. Demographic modeling suggests that diversification amongLatesspecies within the lake occurred during a time period of dramatic changes in lake levels driven by glacial-interglacial cycles, supporting a role of these fluctuations as a “species pump” for lacustrine taxa. We further find that these lake level fluctuations likely contributed to multiple bouts of admixture amongLatesspecies during the mid- to late-Pleistocene (∼90–500 Kya). Together, our findings suggest a dynamic and environmentally linked evolutionary history of theLatesradiation with the potential for dramatic ecosystem consequences for the taxa already present in Lake Tanganyika prior toLatescolonization and diversification.

Significance Statement

When introduced to novel ecosystems, top predators can cause major alterations to biodiversity and food webs. Species introductions to novel habitats can also provide invading taxa with ecological opportunities that facilitate evolutionary diversification. Here, we find evidence that the radiation of endemic top predators in East Africa’s Lake Tanganyika originated surprisingly recently, and that these species have experienced periods of hybridization with a widespread riverine relative throughout their history. These findings have major implications for the history of the lake and suggest that the introduction of Nile perch into Lake Victoria, which caused dramatic ecosystem and food web changes, may be a contemporary analog to the historical events in Lake Tanganyika.
Bridging Earth and Space: A Flexible and Resilient Federated Learning Framework Deployed on the International Space Station
Authors: Casaletto James A. , Foley Patrick , Fernandez Mark , Sanders Lauren M. , Scott Ryan T. , Ranjan Shubha , Jain Shashi , Haynes Nate , Boerma Marjan , Costes Sylvain V. , Mackintosh Graham
Published: January 2025

The public and commercial space industries are planning longer duration and more distant space missions, including the establishment of a habitable lunar base and crewed missions to Mars. To support Earth-independent scientific and medical operations, such missions can leverage artificial intelligence and machine learning models to assist with crew healthcare, spacecraft maintenance, and other critical tasks. However, transferring large volumes of data between Earth and space for model development consumes valuable bandwidth, is vulnerable to communication disruptions, and may compromise crew safety and data privacy. Federated learning enables model training while keeping datain situand only transferring model parameters. In this work, we present a flexible, resilient federated learning framework that provides the secure transmission of model updates between Earth and the International Space Station. On March 15, 2024, this framework pioneered the deployment of federated learning in a spaceflight setting, training classifier models between Earth and the ISS using both real biomedical research data and synthetically generated data.
Transcriptomic signature reveals sensitivity to tubulin inhibitors in colon cancer
Authors: Liñares-Blanco Jose , Chicote Irene , Ros Javier , Alcántara Anna M , Martínez-Quintanilla Jordi , Elez Elena , G Palmer Héctor , Fernandez-Lozano Carlos , Seoane Jose A
Published: January 2025

Colon cancer is the second most common cause of cancer death worldwide. Despite advances in the development of new molecular strategies for stratifying patients with colon cancer, many of these patients do not respond adequately to the standard of care. While previous studies have focused on the development of prognostic gene expression signatures, the exploration of predictive signatures to inform treatment decisions remains incomplete.

In this study, we leveraged public gene expression datasets to design and experimentally validate a 37-gene expression signature for prognosis in colon cancer patients. We obtained a C-index of 0.732 (0.610-0.853) in four independent studies. Specifically, we discovered that the signature is associated with the mitotic phase of the cell cycle. Furthermore, the signature identified a population of colon cancer patients sensitive to tubulin inhibitor drugs. In particular, we validatedin vitroandin vivothe efficacy of paclitaxel, a commonly used tubulin inhibitor in breast cancer treatment, in patient-derived preclinical models.

These results highlight the importance of incorporating gene expression signatures to identify new therapeutic options for colon cancer treatment. Furthermore, the identification of alternative treatment options with potentially improved efficacy holds promise for the development of new clinical trials, and reshapes the biomarker-based treatment strategy for second line and refractory colon cancer patients.
Fire shifts the structure and dynamics of woody plant communities in a mediterranean coastal dune ecosystem
Authors: Mira André F. , Medina Nagore G. , Chozas Sergio , Divieso Raquel , Castro-Rivadeneyra Juan , Díaz-Barradas Mari Cruz , Hortal Joaquín
Published: January 2025

Coastal dune ecosystems are dynamic and fragile environments shaped by complex interactions between vegetation and its characteristic abiotic factors. This study investigates the effects of the wildfire that affected the Doñana Nacional Park (SW Spain) in 2017 on the composition and structure of woody plant communities in two adjacent Mediterranean coastal dune areas: one affected by fire (Cuesta Maneli) and one unburned (Laguna del Jaral). Using a combination of several analyses (such as non-metric multidimensional scaling (NMDS); Mantel tests; and co-occurrence analyses), we examined how fire influences species distribution, abundance, and their interactions along a coastal-inland gradient. Our results reveal that fire homogenizes woody plant community composition across the coastal-inland gradient, reducing spatial differentiation and increasing overall species abundance in the burned area. Soil characteristics influenced species’ composition significantly only in the unburned site, suggesting that fire disrupted abiotic filtering processes. Co-occurrence patterns indicate more neutral interactions in the burned area, consistent with reduced competition and increased resource availability post-disturbance. In contrast, the unburned site displayed structured communities with stronger negative co-occurrence at the smaller scales and spatial clustering of species. Spatial pattern analyses showed that positive co-occurrence was more prevalent in the burned area, particularly at smaller spatial scales, indicating potential facilitation effects during early post-fire recovery. These findings highlight how fire reshapes community structure and dynamics after succession. Understanding these processes is critical for effective conservation and management of Mediterranean coastal dune ecosystems facing increasing anthropogenic pressures and climate change-related disturbances.
Modeling of ATP Transport in an Axon: Effects of Spontaneous Neuron Firing and Mitochondrial Transfer via Tunneling Nanotubes
Authors: Kuznetsov Andrey V.
Published: January 2025

While electrical activity in neurons has been extensively studied, the transport and distribution of adenosine triphosphate (ATP), the primary cellular energy carrier, remain less understood, particularly in relation to metabolic processes in axons. ATP is primarily generated in mitochondria and consumed at synapses, the primary sites of energy demand. Even in healthy axons, approximately half of synaptic boutons lack stationary mitochondria, raising questions about ATP transport between boutons with and without mitochondrial ATP production. This study addresses two key questions: the role of spontaneous neuronal firing in maintaining ATP levels during periods of low energy demand and the ability of a single bouton with a donated mitochondrion to supply ATP to neighboring boutons lacking mitochondria. Using computational simulations, the study examines ATP transport under various firing patterns and mitochondrial distributions, incorporating factors such as quiescent periods, duty cycles, and ATP diffusivity. Spontaneous neuronal firing stabilizes ATP levels during periods of low energy demand, preventing reactive oxygen species (ROS) release from mitochondria. Simulations reveal that in neurons damaged by neurodegeneration, a single bouton containing a donated mitochondrion can support ATP levels in multiple empty boutons. However, as the number of empty boutons increases, ATP concentration declines, potentially falling below the critical threshold required for synaptic transmission.

Nomenclature

a0kinetic constant characterizing the rate of ATP consumption in a bouton

Accross-sectional area of the axon

CATP concentration per unit length of the axon

C0typical value of ATP concentration per unit length of the axon

Cminminimum ATP concentration required to sustain synaptic transmission

dutyduty cycle

DATP diffusivity in the cytosol

ffrequency at which neuron fires during the active phase

iactivenumber of action potentials that occur during the active phase

itotaltotal number of action potentials that propagate down the axon during the active phase plus the number of action potentials that were missed during the quiescent phase

Ldistance between boutons

mATP production rate per unit length of a mitochondrion

ATP production rate per unit mass of tissue

Nemptynumber of boutons in the CV that lack stationary mitochondria

ttime

xposition along the axon

xmhalf-length of a mitochondrion

Greek symbols

γpercentage of tissue volume occupied by mitochondria

δhalf the width of an axonal varicosity

ϖmass of an individual mitochondrion

Λhomeostaticportion of energy expended on homeostatic maintenance of a bouton
Eusociality evolved at a much higher rate in haplodiploid insects
Authors: Suresh Sachin , Linksvayer Timothy A.
Published: January 2025

As a corollary to inclusive fitness theory, W.D. Hamilton’s famous haplodiploid hypothesis proposed that eusociality evolves more readily in haplodiploids because of an inflated genetic relatedness between sisters. For the past sixty years, theoreticians have developed an array of models that further clarify how eusociality might evolve more readily in haplodiploids. Despite this sustained theoretical interest, the predicted evolutionary association between haplodiploidy and eusociality was only recently empirically tested when a family-level analysis in insects found mixed support. Here, we use a large species-level insect phylogeny with all available sociality and ploidy trait data to test whether there is an evolutionary association between haplodiploidy and eusociality. We show that the transition rate from solitary to eusocial life was 15-100 times higher in haplodiploid than diploid lineages, depending on the definitions of eusociality and haplodiploidy. Our results strongly support the prediction of the haplodiploid hypothesis and related models that the evolutionary transition from solitary to eusocial life is facilitated by haplodiploidy.

Significance statement

Sixty years ago, Hamilton hypothesized that eusociality evolves more readily in haplodiploid populations due to inflated relatedness among sisters. This haplodiploid hypothesis is often presented as a textbook example of how inclusive fitness theory can explain the phylogenetic distribution of eusociality across the tree of life. Theoreticians soon noted flaws in Hamilton’s original formulation but have developed more models, still predicting that haplodiploidy facilitates the evolution of eusociality. Despite these theoretical predictions, most researchers have long doubted any phylogenetic association between haplodiploidy and eusociality. We tested this association using a large species-level insect phylogeny with all the available trait data. Our results show that eusociality evolved at a much higher rate in haplodiploids, conclusively supporting the predicted association.
Temperature vaulting: A method for screening of slow- and tight-binding inhibitors that selectively target kinases in their non-native state
Authors: Suzuki Sora , Umezawa Koji , Furuie Gaku , Kikuchi Masaki , Nakamura Daichi G. M. , Fukahori Nanae , Kimura Ninako , Yamakawa Masato , Niwa Takashi , Umehara Takashi , Hosoya Takamitsu , Kii Isao
Published: January 2025

A polypeptide folds into its protein tertiary structure in the native state through a folding intermediate in the non-native state. The transition between these states is thermodynamically driven. A folding intermediate of dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) autophosphorylates intramolecularly, whereas DYRK1A in the native state no longer catalyzes this reaction. The alteration in substrate specificity suggests a conformational transition of DYRK1A during its folding process. Consistent with this hypothesis, we identified FINDY (1), which inhibits the intramolecular autophosphorylation but not the intermolecular phosphorylation, suggesting that the folding intermediate possesses an alternative inhibitor-binding site. Meanwhile, it remains an issue that the methods for approaching the alternative binding site require an intricate assay tailored to the individual target. Here we show a method, designated as “temperature vaulting,” for screening the non-native-state-targeted inhibitors of DYRK1A. Transient heating of recombinant DYRK1A protein drove the reversible transition between the native state and the non-native state targeted by FINDY (1). At physiological temperature, FINDY (1) slowly bound to the DYRK1A protein. These results indicate that transient heating accelerates the slow-binding process by assisting the protein to overcome the high-energy barrier leading to the target non-native state. The energy barrier also slowed down the dissociation process, resulting in tight binding between DYRK1A and FINDY (1). Furthermore, this study suggests that the dissociation rate underlies the inhibition selectivity of FINDY (1) between DYRK1A and its family kinase DYRK1B. This method enables the identification of slow- and tight-binding inhibitors that have been missed in conventional assays.

HIGHLIGHTS

A temperature vaulting method facilitated the screening of a slow- and tight-binding inhibitor targeting DYRK1A in the non-native state.

A structure–activity relationship study of FINDY (1) demonstrated that a slight alteration in chemical structure impacts the inhibition kinetics.

A molecular dynamics simulation of DYRK1A revealed binding-ready conformations for FINDY (1).

The dissociation rate may underlie the inhibition selectivity of FINDY (1) between DYRK1A and DYRK1B in cellular systems.
Gene family evolution reveals dietary adaptations in butterflies and moths
Authors: Weng Yi-Ming , Martinez Jose I. , Markee Amanda , Plotkin David , Sondhi Yash , Mongue Andrew J. , Frandsen Paul B. , Kawahara Akito Y.
Published: January 2025

Butterflies and moths (Lepidoptera) are a hyperdiverse lineage of nearly 160,000 described species and their evolutionary success is postulated to be tightly correlated with the radiation of their primary host — angiosperms. Previous studies found that a significant number of emergent gene families are specific to Lepidoptera, with many genes linked to odorant receptors and peptidases, suggesting that such genetic innovations may be linked to their diversification. Here, we use genomic resources to identify lineage-specific gene families in four nested lepidopteran clades (e.g. Lepidoptera, Glossata, Ditrysia, and Apoditrysia). Among nearly a hundred gene families specific to each group, a handful of gene families have specific and interpretable functions. We found that many digestion-related gene families emerged early in the evolution of Lepidoptera, followed by genes associated with detoxification and immunity. This result aligns with the evolutionary transition in ancient Lepidoptera diets to terrestrial plants, highlighting the emergence of detoxification mechanisms in the megadiverse Ditrysia as a critical adaptation driven by the proliferation of plant chemical defenses. We also found gene families originating from horizontal gene transfer (HGT) events, likely introduced from bacteria and fungi to the common ancestors of Lepidoptera and Ditrysia, respectively. These HGT-derived genes likely played a pivotal role in supporting the dietary transition from algae, diatoms, and aquatic plant debris to fungi and primitive terrestrial plants, ultimately enabling the adaptation to the most dominant angiosperm species.
IL-9 as a naturally orthogonal cytokine with optimal JAK/STAT signaling for engineered T cell therapy
Authors: Jiang Hua , Limsuwannarot Sam , Kulhanek Kayla R. , Pal Aastha , Rysavy Lea W. , Su Leon , Labiad Ossama , Testa Stefano , Ogana Heather , Waghray Deepa , Tao Pingdong , Jude Kevin M. , Seet Christopher S. , Crooks Gay M. , Moding Everett J. , Christopher Garcia K. , Kalbasi Anusha
Published: January 2025

Arming T cells with a synthetically orthogonal IL-9 receptor (o9R) permits facile engraftment and potent anti-tumor functions. We considered whether the paucity of natural IL-9R expression could be exploited for T cell immunotherapy given that, in mice, high doses of IL-9 were well-tolerated without discernible immune modulation. Compared to o9R, T cells engineered with IL-9R exhibit superior tissue infiltration, stemness, and anti-tumor activity. These qualities are consistent with a stronger JAK/STAT signal, which in addition to STAT1/3/5, unexpectedly includes STAT4 (canonically associated with IL-12 but not common γ-chain cytokines). IL-9R T cells are exquisitely sensitive to perturbations of proximal signaling, including structure-guided attenuation, amplification, and rebalancing of JAK/STAT signals. Biased IL-9R mutants uncover STAT1 as a rheostat between proliferative stem-like and terminally differentiated effector states. In summary, we identify native IL-9/IL-9R as a natural cytokine-receptor pair with near-orthogonal qualities and an optimal JAK/STAT signaling profile for engineered T cell therapy.
Synthetic inhibition of the SUMO pathway by targeting the SAE1 component via TAK-981 compound impairs tumor growth and chemosensitizers embryonal and alveolar rhabdomyosarcoma cell lines
Authors: Codenotti Silvia , Lauschke Volker M. , Casella Emma V. , Andersson Daniel C. , Fanzani Alessandro , Gastaldello Stefano
Published: January 2025

Background

Rhabdomyosarcoma (RMS) is a highly aggressive pediatric soft tissue sarcoma with limited therapeutic options, particularly for cases resistant to conventional treatments. The SUMOylation pathway, which plays a key role in regulating cell cycle, apoptosis, and transcription, has emerged as a potential therapeutic target in RMS. Elevated levels of SUMO1 and SUMO2/3 conjugates in RMS cell lines, compared to normal human skeletal muscle cells, underscore the association between hyper-SUMOylation and aggressive cancer phenotypes. Understanding these molecular underpinnings is critical for the development of innovative and effective treatments.

Methods

The investigation encompassed transcriptomic and protein analyses to profile SUMOylation pathway components across alveolar and embryonal RMS subtypes, aiming to identify heterogeneity that could guide personalized therapy approaches. TAK-981’s interactions with chemotherapeutic agents, were evaluated for synergistic effects. Additionally, its impact on radiosensitivity and key signaling pathways, such as AKT and ERK phosphorylation, was assessed to elucidate its mechanism of action.

Results

Transcriptomic and proteomic analyses revealed distinct expression profiles of SUMOylation pathway components across RMS subtypes, highlighting heterogeneity that could guide personalized therapeutic strategies. Notably, SAE1 protein was overexpressed in RMS tissues and cells, positioning it as a potential biomarker for this cancer. Its activity was effectively counteracted by TAK-981, a SUMO inhibitor that demonstrated significant therapeutic potential by suppressing RMS cell proliferation and migration and enhancing the cytotoxic effects of chemotherapeutic agents, actinomycin D and doxorubicin. However, TAK-981 did not increase radiosensitivity, suggesting its selective action through chemical inhibition mechanisms. Mechanistically, TAK-981 reduced phosphorylation of key signaling proteins, including AKT and ERK, critical for RMS cell survival.

Conclusion

The findings of this study establish TAK-981 as a promising therapeutic agent for RMS. The results also provide foundational insights into the role of SUMOylation associated to the new biomarker SAE1 in RMS and its subtypes, paving the way for the development of personalized treatment strategies that leverage SUMO pathway inhibition.
Tumor suppressor genotype influences the extent and mode of immunosurveillance in lung cancer
Authors: Adler Keren M. , Xu Haiqing , Gladstein Amy C. , Irizarry-Negron Valerie M. , Robertson Maggie R. , Doerig Katherine R. , Petrov Dmitri A. , Winslow Monte M. , Feldser David M.
Published: January 2025

The impact of cancer driving mutations in regulating immunosurveillance throughout tumor development remains poorly understood. To better understand the contribution of tumor genotype to immunosurveillance, we generated and validated lentiviral vectors that create an epi-allelic series of increasingly immunogenic neoantigens. This vector system is compatible with autochthonous Cre-regulated cancer models, CRISPR/Cas9-mediated somatic genome editing, and tumor barcoding. Here, we show that in the context of KRAS-driven lung cancer and strong neoantigen expression, tumor suppressor genotype dictates the degree of immune cell recruitment, positive selection of tumors with neoantigen silencing, and tumor outgrowth. By quantifying the impact of 11 commonly inactivated tumor suppressor genes on tumor growth across neoantigenic contexts, we show that the growth promoting effects of tumor suppressor gene inactivation correlate with increasing sensitivity to immunosurveillance. Importantly, specific genotypes dramatically increase or decrease sensitivity to immunosurveillance independently of their growth promoting effects. We propose a model of immunoediting in which tumor suppressor gene inactivation works in tandem with neoantigen expression to shape tumor immunosurveillance and immunoediting such that the same neoantigens uniquely modulate tumor immunoediting depending on the genetic context.

One Sentence Summary

Here we uncover an under-appreciated role for tumor suppressor gene inactivation in shaping immunoediting upon neoantigen expression.
Bioinformatics Analysis of Non-Coding UTR Variants inSTAT1Reveals Disruption of miRNA Binding, mRNA Stability, and Oncogenic Potential
Authors: Kamal Ebtihal , Ahmed Mehad
Published: January 2025

Single nucleotide polymorphisms (SNPs) are associated with a wide range of disorders, including diverse cancer types. In the context of cancer, alterations within non-coding regions, specifically untranslated regions (UTRs), have proven substantially important. SNPs localized in the 3 prime UTRs of theSTAT1gene were assessed for their association with miRNAs using PolymiRTS, miRNASNP, and MicroSNIper. In addition, the 5 UTR SNPs were analyzed by SNP INFO for changes in the transcription factors binding sites. The significant SNPs were further analysed by Cscape to predict their oncogenic probability. The secondary structures of the wild type and the mutant mRNA were analysed by RNAfold.

Out of 605 SNPs analyzed, 14 UTR SNPs (six in the 3⍰ UTR region and eight in the 5⍰ UTR region) were identified with functional annotation ≤2a by RegulomeDB. The associations of 8 SNPs with miRNAs common between the 3 databases (PolymiRTS, miRNASNP, and MicroSNIper) were found.

Moreover we identified six SNPs (rs1197872838, rs3088307, rs45470392, rs1413522785, rs531009254, and rs190508584) destabilize the mRNA structure resulting in substantial change in free energy (ΔG). The SNP rs45470392 in 5⍰ UTR was predicted to alter the transcription factor binding sites. rs188557905, rs1220766131, rs1413522785, rs1168, and rs999207177 were predicted to be oncogenic SNPs in this study using bioinformatics tools.

Our findings highlight the impact of 3⍰ and 5⍰ UTR SNPs on miRNA, transcription, and translation ofSTAT1. These analyses suggest that these SNPs can have substantial functional importance in theSTAT1gene. Future experimental validation could establish their potential role in the diagnosis and therapeutics of various diseases, including cancer.
Zebrafishadamtsl4knockout recapitulates key features of humanADAMTSL4-related diseases: a gene involved in extracellular matrix organization, cell junctions and development
Authors: Tevar Angel , Aroca-Aguilar Jose-Daniel , Atiénzar-Aroca Raquel , Ramírez Ana I. , Fernández-Albarral José A. , Escribano Julio
Published: January 2025

Loss-of-function mutations inADAMTSL4,a gene encoding an extracellular matrix-associated protein with incompletely understood biological roles, are linked to autosomal recessive disorders predominantly characterized by lens dislocation. Pupil ectopia, increased intraocular pressure, retinal detachment, cataracts, and skeletal abnormalities are also observed in some patients. To investigateADAMTSL4biology and related diseases we established a zebrafish knockout line using CRISPR/Cas9 genome editing. The generated zebrafish model harboured the c.234-351del mutation inadamstsl4, reducing its mRNA levels by 75% in 6 days post fertilization (dpf) larvae, and predicting to produce an inactive protein (p.(Gln78Hisfs*127)). Forty percent of F3 knockout larvae (6 dpf) displayed lethal phenotypes characterized by multiple ocular and non-ocular developmental defects, including pericardial, perivitelline and periocular edema, absence of swim bladder, craniofacial malformations and microphthalmia. The remaining 60% larvae survived and displayed only reduced pupil area, indicating incomplete penetrance of the lethality. Histology revealed extracellular matrix (ECM) and intercellular junctions abnormalities within the cornea, iris, lens, and retinal pigment epithelium (RPE). Adult knockout zebrafish (6 months) presented phenotypes resembling ectopia lentis et pupillae and craniosynostosis, with optical defects in the lens and impaired visual function. ECM and cell junction disorganization in the cornea, lens and RPE were also present in these animals. Transcriptomic analysis revealed disrupted expression of genes involved in development, ECM and cell junctions among other biological processes. These findings show that adamtsl4recapitulates key features of humanADAMTSL4-related disorders and that this gene is essential for normal ECM structure, cell junctions and embryonic development.

Highlights

Zebrafishadamtsl4knockout recapitulates humanADAMTSL4-related phenotypes.

adamtsl4is involved in embryonic development.

adamtsl4participates in ECM organization and cell adhesion.
Ohno-miRNAs: intragenic miRNA pairs derived from whole-genome duplication
Authors: Agasso Leonardo , Molineris Ivan , Caselle Michele
Published: January 2025

Two rounds of whole-genome duplication (WGD) occurred about 500 million years ago and played a major role in the evolution of the vertebrate genomes. Human genes derived from WGD are called “ohnologs”. Ohnologs are involved in fundamental biological processes and significantly contributed to the complexity of the human gene regulatory network. Given the central role of miRNAs in gene regulation, we investigated the contribution of ohnolog miRNAs (Ohno-miRNAs) to the human gene regulatory network. We worked on the identification of miRNA pairs in the human genome derived from the two rounds of whole-genome duplication in early vertebrate lineage and their role in the structure of the human gene regulatory network. By focusing on intragenic miRNAs within ohnolog gene pairs, we identified Ohno-miRNAs as having higher retention rates and sequence similarity compared to miRNAs hosted on paralogue gene pairs from small-scale duplications (SSD). They also show a stronger tendency to regulate common target genes. Analyzing the role of Ohno-miRNAs in the human gene regulatory network, we showed that Ohno-miRNAs are statistically overrepresented in specific network motifs commonly associated with redundancy and complexity, highlighting their central role in this particular layer network.
Spacer Extension Reconciles Specificity and Activity in High-Fidelity Cas9 Genome Editing
Authors: Lu Zhike , Wei Rongwei , Zheng Rong , Shin Young-Cheul , Zhang Qingfeng , Li Jianbo , Wang Xiaoqi , Wei Yi , Liu Botao , Chen Yang , Zhang Heng , Chen Hui , Ma Lijia
Published: January 2025

Abstract

Engineering high-fidelity CRISPR enzymes often leads to reduced cleavage activity, creating a significant hurdle in balancing nuclease specificity and efficiency for clinical applications. Here, we demonstrate that extending the spacer to 21 or 22 nucleotides restores the impaired cleavage activity of SuperFi-Cas9, an engineered high-fidelity Cas9 variant with seven mutations at the PAM-distal region. Structural and mutational analyses reveal that the spacer extension strengthens additional interactions at the PAM-distal end, stabilizing the nuclease– sgRNA–DNA complex, which appears to be disturbed due to the seven mutations. This approach not only provides a high-fidelity Cas9 with uncompromised efficiency but also introduces a novel strategy to enhance CRISPR complex stability. Our findings offer a promising avenue for precise and efficient genome editing, crucial for advancing CRISPR technologies toward clinical translation.
Limit of detection of Raman spectroscopy using polystyrene particles from 25 to 1000 nm in aqueous suspensions
Authors: Mayorga Cindy , Athalye Shreya Milind , Boodaghidizaji Miad , Sarathy Neelesh , Hosseini Mahdi , Ardekani Arezoo , Verma Mohit S.
Published: January 2025

Raman spectroscopy is an analytical method capable of detecting various microorganisms and small particles. Here, we used 25-1000 nm polystyrene particles in aqueous suspensions, which are comparable in size to viral particles and viral aggregates, to determine the limit of detection of a confocal Raman microscope. We collected Raman spectra using a 785 nm wavelength laser with a power of 300 mW and a 10 s exposure time, with a 5X objective lens. We detected the most prominent peak of the polystyrene particles at 1001 cm-1, corresponding to the ring breathing mode. We established the minimum and maximum limit of detection (LODminand LODmax) using a partial least squares (PLS) model. The LOD of the smallest size of 50 nm was identified as 7.47 × 1012–7.64 × 1012particle/mL, and for the largest size of 1000 nm, 6.09 × 108–6.24 × 108particle/mL. We demonstrated that Raman spectroscopy was non-destructive under these conditions by comparing the particle size before and after collecting Raman spectra using dynamic light scattering. Due to their size similarity to viral particles and viral aggregates, this systematic characterization of polystyrene particles provides detailed information on their Raman spectral signatures in aqueous suspensions. These findings establish a foundation for using Raman spectroscopy for the detection of small particles in aqueous suspensions and highlight its potential as a tool for real-time monitoring in vaccine manufacturing.