The mammalian heart is classically described as arising from two cardiac lineages, first and second. Yet, recent lineage tracing of Mesp1 mesoderm and live imaging show that cardiac progenitors are already partly biased toward specific heart regions during gastrulation, despite extensive cell mixing. We propose that cardiac fate is probabilistic, analogous to stochastic fate strategies in the retina. Epiblast cells carry probability distributions over fates that resolve during gastrulation into specific regional allocations. Reanalyzing a retrospective clonal dataset, we find that the clones do not single out one fixed lineage tree. Instead, the data permit a family of 361 distinct restriction topologies, none dominant, and only 40 of these are binary. Among the binary trees, the best-supported recover both where progenitors sit along the proximal-distal axis of the streak and the order in which they leave it, as recent prospective lineage tracing shows. Rather than replacing the first/second lineage concept, this framework shifts the focus from fixed lineage identity to fate probabilities. Testing this idea will require prospective live imaging from epiblast through heart tube formation.
Tumor evolution, from premalignant lesions to metastasis, is increasingly recognized as shaped by continuous interplay between tumor cells metabolism and their microenvironment. During tumor initiation, major oncogenic pathways drive early metabolic reprogramming of lipid, amino acid, and energy pathways to promote cell competition and clonal expansion. These metabolic changes reciprocally shape the tumor microenvironment (TME) through metabolite fluxes, extracellular matrix remodeling, and immune reprogramming, generating adaptive niches that sustain tumor progression and metastasis. Cancer cell metabolic adaptability becomes even more crucial to survive dissemination and adapt to a new, distant microenvironment. Here, we discuss these dynamic interplays and highlight the p53 pathway as an integrative hub linking oncogenic signaling, metabolic rewiring, and tumor microenvironmental adaptation throughout carcinogenesis. We will also outline how emerging technologies may redefine the TME-p53-metabolism interplay uncovering therapeutically exploitable metabolic vulnerabilities.
Single-stranded DNA (ssDNA) is an emerging nucleic acid tool with the potential to become a more efficient genetic delivery vector than traditional plasmids in prokaryotes. While conventional double-stranded DNA (dsDNA) vectors are easily recognized and attacked by bacterial defense systems, ssDNA's structural differences make it resistant to corresponding enzymatic recognition. Moreover, the presence of unpaired bases in ssDNA enables efficient binding with other molecules, allowing for regulated gene expression and targeted delivery of other nucleic acids or molecules. In this study, we investigated the natural forms of ssDNA and summarized its synthesis and introduction methods. By analyzing existing applications of ssDNA in eukaryotes, we demonstrated the strategies and advantages of applying it to genetic manipulation in prokaryotes.
Euarthropods are characterized by segmented, sclerotized, and articulated bodies and appendages, expressed through arthrodization and arthropodization. Both are regarded as defining features of Euarthropoda and have been proposed to evolve in a specific sequence, during which arthropodization preceded arthrodization, and initiated in frontal appendages and was subsequently co-opted into other appendages. Here, we investigate their evolution across Ecdysozoa using morphological comparison, ancestral-state reconstruction (ASR), and semi-quantitative methods. Morphological comparisons and ASR reveal repeated gains, reductions, losses, and regional decoupling of segmentation, sclerotization, and articulation in both body and appendicular systems. Semi-quantitative indices indicate variable degrees of arthrodization and arthropodization across ecdysozoan lineages and do not show a progressive increase from the basal panarthropods and the stem-group euarthropods to crown-group euarthropods. Together, these results support a modular, convergent evolution of arthrodization and arthropodization in different lineages, probably through repeated modification, reduction, or redeployment of the ancestral developmental toolkit.
In humans, the FLOWER (FWE or CACFD1) gene encodes a set of small, alternatively spliced transmembrane protein isoforms (hFWE1-4). The canonical human isoform, hFWE4, and its various orthologues, which we collectively refer to as FWE for simplicity, assumes a four transmembrane pass structure with cytosolic N- and C-termini. This topology exposes YXXΦ motifs enabling interaction with trafficking machinery and positions highly-conserved cationic residues for interaction with membrane phospholipids. While identified initially as a mediator of synaptic vesicle endocytosis in presynaptic neurons, FWE is now recognized as a critical mediator of endo-lysosome related organelle (ELRO) trafficking across diverse cell types including cytotoxic T-lymphocytes and epidermal keratinocytes. A unifying feature of FWE function appears to be elevation of cytosolic Ca2+ levels. However, the underlying molecular mechanisms remain elusive. This review synthesizes current literature on FWE-mediated membrane trafficking, highlighting the evolutionary conservation of its coding sequence and tertiary structure. We propose new structure-compatible hypotheses for FWE-dependent cytosolic Ca2+ elevation and subsequent ELRO trafficking, highlighting experimental strategies that may aid testing of these hypotheses. Finally, we examine the distinct localizations and potential functions of non-canonical FWE isoforms.
Homologous recombination (HR) is a DNA double-strand break repair pathway that preserves genome integrity by restoring genetic information lost through programmed or spontaneous DNA damage. As a template-directed process, HR identifies homologous DNA sequences to accurately repair broken chromosomes while minimizing inappropriate recombination events. Central to this process are the RecA-family recombinases. Most eukaryotes use Rad51 during mitosis and meiosis and Dmc1 during meiosis to locate and pair homologous DNA sequences. To ensure high-fidelity repair, eukaryotes have evolved regulatory protein networks that control recombinase filament assembly, organization, and strand exchange. Here, we review recent advances in understanding how recombinase filament length, architecture, and dynamics influence the fidelity and outcome of homologous recombination. We discuss their distinct roles in mitotic and meiotic recombination and propose how evolution has shaped filament properties to regulate interactions between donor and recipient DNA templates and promote accurate genome maintenance.
Recent work has expanded understanding of extracellular vesicle (EV) biology by identifying midbody remnants (MBRs) as large, translationally competent vesicles released during mitosis. MBRs contain ribosomes, mitochondria, translation factors, and selected mRNAs and small RNAs concentrated within a condensate‑like ribonucleoprotein core and can support protein synthesis after extracellular release. These features distinguish MBRs from more extensively studied exosomes and microvesicles yet also place them within a broader continuum of large EVs with organelle‑rich, cell‑like properties. This review summarizes current knowledge of MBR biogenesis, molecular organization, and translation competency; contrasts MBRs with canonical EVs and other large EVs; and discusses possible roles in development, tissue homeostasis, disease, and brain function. Particular emphasis is placed on outstanding mechanistic and physiological questions, including how MBR translation is regulated, which recipient cells interact with endogenous MBRs, and whether translation‑competent EVs offer practical advantages over existing EV platforms for therapeutic or biotechnological applications.
Prostanoids are involved in various physiological/pathophysiological functions, such as inflammation. Since persistent inflammation may cause chronic symptoms, their activation is very short and needs to be properly terminated. Due to structural similarities, some prostanoids and their metabolites may have ability to activate noncognate prostanoid receptors as biased agonists and/or blockers. However, the physiological roles of these biased activities remain unclear. This review mainly focused on prostaglandin (PG) E2 and PGD2 as well as their metabolites, and hypothesized that the physiological roles of their biased activities may be novel nongenomic immediate feedback systems strictly confined to local areas as autacoids. These nongenomic feedback systems completely differ from the genomic feedback systems used by endocrine hormones. Therefore, the biased activities of prostanoids may be newly defined and/or re-acknowledged as unique, rapid, and fine-tuned translation/transcription-free feedback systems which may be responsible for the roles of prostanoids in maintaining homeostasis as rapid-acting local-mediators, autacoids.
Regulatory T cells (Tregs) are central to immune tolerance, yet antigen-specific cell therapies lag behind cytotoxic CAR-T approaches. Most CAR-Treg programs still use single-chain variable fragments (scFvs) derived from monoclonal antibodies optimized for effector function, a bias that may promote CAR clustering, tonic signaling, and lineage instability. Here, we propose a phage-first binder discovery framework for CAR-Tregs and state testable hypotheses linking phage-display selection pressures to scFv biophysical properties and Treg fate. We hypothesize that multi-parameter selection and counter-selection can enrich scFvs with moderate affinity, low polyspecificity, and improved framework stability, thereby lowering tonic signaling in defined backbone architectures and preserving FOXP3/TSDR stability under inflammatory stress. We outline a falsifiable two-phase roadmap-bench triage followed by mechanistic and preclinical validation to determine how scFv properties, expression level, and intracellular signaling domains define an optimal tonic window for durable immune regulation.
Artificial intelligence (AI) strategies are revolutionizing genomics by extracting complex patterns that traditional statistical pipelines are likely to miss. This mini-review aims to provide a concise overview of how AI is transforming major genomic technologies including variant calling, gene expression analysis, single-cell transcriptomics, CRISPR-Cas9 optimization, and multi-omics integration. In genome sequencing, machine learning variant callers greatly improve the accuracy and the rate at which single nucleotide and structural variants are called. In bulk RNA-Seq, AI augmented quantification, denoising, and differential expression modules complement the highly established STAR-featureCounts-DESeq2 pipeline, revealing subtle signals in big data sets. In single cell transcriptomics, deep learning approaches enhance batch correction, automate cell type annotation, and track developmental trajectories, hence clarifying cellular heterogeneity. AI-assisted guide RNA design, outcome prediction, and nuclease engineering enable more efficient CRISPR-Cas9 editing, reducing experimental cycles, and off-target effects. Finally, integrated platforms that combine genomic, transcriptomic, epigenomic, proteomic, and metabolomic layers provide an integrative view of cellular regulation and disease mechanisms. The review also covers current limitations, sparsity of data, model bias, privacy, and the need for standardized benchmarks and offers future directions in the form of interpretable models, collaborative learning, and open science practices. Together, these developments render AI an indispensable partner to unravel genomic complexity and accelerate precision medicine applications.
We review recent multi‐omics analyses of the coral heat stress response to explore the generality of the Oxidative Theory of Coral Bleaching (OTCB), which posits that algal symbiont release is the final act of defense by the coral host to survive alga‐derived oxidative stress. The OTCB is particularly relevant given that ocean warming, which is accelerating under climate change, has proven devastating for corals, leading to the bleaching phenotype and widespread reef loss. Multi‐omics results, in combination with other data, such as genome‐wide association studies, support the idea that coral bleaching is a multifactorial response that reflects a wide array of causes and effects and is population‐specific under most conditions, with coral ploidy and genotype being critical to bleaching sensitivity. This perspective leverages the location, algal and prokaryotic microbiome, and host genotype‐specific aspects of coral resilience to promote a new “personal genomics” approach to coral conservation, analogous to that used in human health.
Morphogenesis, the sculpting of tissues into functional architectures, requires precise orchestration of cell shape changes and rearrangements. In the Drosophila salivary gland (SG), recent studies reveal that Crumbs (Crb) and non-muscle Myosin II (MyoII) act as collaborative antagonists to drive SG internalization. Crb, a conserved type I transmembrane protein best known as a master regulator of apical-basal polarity, stabilizes attachments by forming homophilic extracellular bridges between neighboring cells and restricting the distribution of junctional MyoII. Conversely, MyoII is a highly conserved motor protein that drives apical constriction and junctional shrinkage by generating contractile forces on apicomedial and cortical actin and promoting apical membrane removal. During SG invagination, these opposing activities are exquisitely tuned, with Crb-mediated stabilization balancing MyoII-mediated destabilization to facilitate apical constriction for internalization and neighbor exchange for tube elongation. The conserved interplay between Crb and MyoII goes beyond the shaping of organs, serving as a critical mechanism for cell extrusion and EMT-like processes in animal development.
The One Health (OH) framework has gained prominence in recent years, promoting an approach integrating the health of humans, animals, and the environment. However, its implementation as it pertains to the microbial world has largely prioritized zoonotic diseases, antimicrobial resistance, as well as pandemic preparedness and management, narrowing its ecological scope and confining microorganisms to the realm of pathogens. We explore the historical development of OH to identify key gaps and limitations and argue for the integration of microorganisms as ecologically constitutive, critical to sustain life processes and ecosystem functioning. We provide actionable recommendations, beginning with a shift in how microorganisms are conceptualized, extending to the development of microbial metrics and culminating in proposals for microbial governance to address current shortcomings and inform future policymaking. Collectively, these measures aim to strengthen the OH framework in confronting global challenges, including biodiversity loss and climate change, while aligning with its broader commitments to health and sustainable development.
Cryopreservation-enabled workflows decouple tissue acquisition from organoid generation, allowing archived biopsies and surgical explants to be revisited as renewable experimental models. This shift expands access to patient-specific material, reduces the logistical and batch variability inherent in fresh-tissue pipelines, and enables retrospective and longitudinal studies anchored in real-world clinical cohorts. Proof-of-concept studies show that liver organoids can be derived from cryopreserved human tissues that retain viability and disease-relevant phenotypes, but performance remains sensitive to the source material, culture lines, and protocol details. The field remains fragmented, lacking broadly adopted standard operating procedures, shared post-thaw quality benchmarks, and interoperable data infrastructures linking organoid biobanks to clinical metadata. In this work, we argue that cryopreservation-enabled organoid biobanking should be treated as foundational infrastructure for precision hepatology, and we outline a pragmatic roadmap for coordinated implementation over the next decade.
For decades, eukaryotic circadian timing has been framed mainly through nuclear transcription-translation feedback loops (TTFLs). Here, we synthesize evidence supporting a broader organelle-centered model in which cellular time emerges from dynamic coupling between TTFL clocks, post-translational feedback loop (PTFL) oscillators, and entrained rhythmic modules across mitochondria, endoplasmic reticulum, lysosomes, peroxisomes, Golgi apparatus, plasma membrane, and cytoskeleton. Metabolic flux, redox cycling, proteostasis, ion handling, membrane excitability, trafficking, and mechanotransduction act as temporal currencies that either sustain selected transcription-independent rhythms or transmit phase information within a TTFL-coordinated network. In this layered architecture, the TTFL remains a central integrator that stabilizes inter-organelle phase relationships, aligns intracellular rhythms with environmental Zeitgebers, and links biochemical state to epigenetic and RNA-based regulation. We propose that circadian dysfunction reflects progressive intracellular desynchronization rather than isolated clock-gene failure, opening diagnostic and therapeutic opportunities aimed at restoring subcellular temporal coherence.
The interactions between human viruses and human stem cells may differ from those with differentiated cells. These differences may arise through heterogeneous intercellular mechanisms and responses to specific infectious agents, resulting in different phenotypes that affect human pathophysiology. Understanding and exploiting such differences could have clinical and translational potential. Here, we discuss the various mechanistic interactions between stem cells and viruses related to entry mechanisms, replication dynamics, and immunomodulation. In doing so, we critically assess proposed models and hypotheses about how viruses manipulate stem cell biology, while also providing new paradigms for stem cell biology and therapeutic interventions. We highlight recent discoveries on the dual role of viruses in oncogenesis and oncolysis. In parallel, we explore similarities between stem cells and complex viruses-such as giant viruses and jumbo phages-to propose novel perspectives on viral adaptability and pathogenesis. We examine both established mechanisms and emerging viral phenomena to encourage further research and debate on the clinical implications of viral interactions with stem cells.
Fungal DNA is systematically under-detected in shotgun metagenomics, likely due in part to physical barriers like melanized cell walls and complex DNA conformations. Additionally, Oxford Nanopore Technologies sequencing with native fungal DNA often results in rapid pore clogging and unusual translocation dynamics, possibly due to intrinsic, yet undescribed, structural complexities. Exploring these signals could reveal novel fungal genome architectures, enhance sequencing accuracy, and drive advances in fungal biology.
Student navigation through a PhD Program is marked by intellectual challenges, emotional fluctuations, and personal growth. Students develop excellence by absorbing and adjusting, recovering, and improving in response to challenges. The ability to bounce back is termed elasticity, an engineering principle where systems are designed to withstand stress, recover from disruptions, and maintain functionality. Students can proceed from elasticity (resilience) to then develop antifragility, the ability to become better from an experience. It occurred to the first author (a PhD graduate student in biomedical sciences) that developing antifragility is conceptually easier to understand by applying basic engineering concepts. We examine five thematic categories that connect engineering concepts with practical realities of graduate study: (1) load and stress management; (2) redundancy and backup systems; (3) setback modes and recovery; (4) adaptive capacity and flexibility; and (5) sustainability and long-term performance. By understanding how to gain antifragility, one can navigate demands more effectively.
Nuclear entry of HIV-1 viral cores triggers the formation of phase-separated membraneless organelles called HIV-1-MLOs. Numerous unknowns remain regarding the persistence, in vivo formation, and function of these recently discovered condensates. New evidence shows that HIV-1-MLOs can persist stably for several weeks in infected cells and that their presence precedes the viral genome expression. These new results are accompanied by the first in vivo observation of HIV-1-MLOs, which have only been described in vitro until now. Additional evidence demonstrates that HIV-1-MLOs are the main hub of nuclear reverse transcription for a wild-type virus and shield viral genomes from innate immunity. Intriguingly, empty viral cores-likely remnants following the completion of reverse transcription-have been observed, challenging the classical view of uncoating as the complete disassembly of the viral core shell.
Isogenic lines, understood here as lines approaching genome-wide homozygosity while remaining fertile and stable across successive generations, have transformed genetics and toxicology in vertebrate models. In zebrafish (Danio rerio), however, more than four decades of gynogenesis, clonal line production and inbreeding have not yielded stable, broadly usable fully homozygous resources. We argue that, in practice, this reflects biological constraints rather than merely technical limitations. Historical, genomic and developmental evidence points to three major barriers: polygenic and environmentally sensitive sex determination, strong dependence on maternal factors during early development, and a structurally dynamic genome shaped by duplication, copy-number variation and substrain divergence. Together, these features make extreme homozygosity biologically fragile, with recurrent sex-ratio bias, infertility and line collapse, and they support experimental strategies that treat genetic background as a variable to be reported, tested and exploited rather than idealized away.
