Sepsis is systemic inflammation with high mortality, accompanied by multi-organ failure including acute respiratory distress syndrome. Extracellular vesicles (EVs) encapsulating bioactive cargoes, mediate cell-cell communication to exert systemic regulation. However, whether and how host lung tissue responds quickly to plasma bacterial infection through EVs in sepsis is poorly understood. Here, we identify that peripheral blood macrophages secrete more exosomal G6PD protein to induce lung injury by rewiring purine metabolism during sepsis. Guanine accumulation reduces H3K27 trimethylation and subsequently induces Nos2, Ccl6 and Il6 expression by suppressing de novo EZH2 synthesis. Macrophage-specific Rab27a or G6pd knockout mice had low exosomal G6PD protein in serum, and failed to exert lung injury upon bacterial infection. Beyond, targeting macrophage-derived G6PD by G6PD inhibitors or engineered EVs delivering si-G6pd relieves lung injury in septic mice. In summary, our findings reveal that circulating G6PD promotes lung injury by rewiring purine metabolism and remodeling the epigenetic profile in sepsis, shedding light on the critical role of EVs as a pro-inflammatory signal and targeting G6PD for future sepsis diagnosis and treatment.
Tryptophan catabolism by the gut microbiota is increasingly recognized as a hub connecting environmental challenges to cardiovascular health. By systematically profiling tryptophan metabolites in humans who had moved from low altitude to an altitude of 4300 m and in mice exposed to hypobaric hypoxia, we found that circulating and fecal levels of indole-3-propionic acid (IPA), a gut microbiota-derived tryptophan metabolite, are reduced in both settings, and these reductions correlate with the severity of cardiac injury. IPA supplementation in mice ameliorated hypobaric hypoxia-induced cardiac dysfunction. Mechanistically, IPA directly bound to hypoxia-inducible factor-1α (HIF-1α) at residue P564, competitively inhibiting its interaction with von Hippel-Lindau protein and preventing proteasomal degradation, thereby stabilizing HIF-1α. HIF-1α activation shifted cardiac metabolic substrate utilization from fatty acids to glucose and reduced oxygen consumption. Cardiomyocyte-specific HIF-1α knockout completely abolished IPA's cardioprotective effects. Moreover, gut microbiota depletion abrogated the protective effect of tryptophan against hypobaric hypoxia exposure. Finally, IPA protected against myocardial infarction in a HIF-1α-dependent manner. These results suggest that IPA is an endogenous HIF-1α agonist to protect the heart against hypoxia/ischemia, representing a promising therapeutic candidate for hypoxic/ischemic diseases.
Hypervirulent Acinetobacter baumannii (hvAB) is a leading cause of refractory bacterial pneumonia. However, cellular and molecular mechanisms underlying hvAB-induced host protection and pulmonary immunopathology remain poorly understood. Here, we performed longitudinal single-cell RNA sequencing on lung tissues in a mouse model of hvAB pneumonia and delineated a time-dependent innate immune landscape of the infected lung. Functional experiments demonstrated that neutrophil (Neu) was dispensable for hvAB clearance but exacerbated acute lung injury through Neu extracellular traps formation (NETosis). Lung-resident CD11b- alveolar macrophage (AM) was indispensable for host defense and for restraining early Neu infiltration. Upon hvAB infection, CD11b- AM underwent phenotypic switching to a hyper-activated CD11b+ pro-inflammatory state, characterized by markedly decreased phagocytic activity and robustly enhanced IL-1β secretion. Mechanistically, IL-17A acted as a key pathogenic mediator by driving excessive Neu recruitment, aberrant NETosis, and progressive AM loss. IL-17A was predominantly produced by group 3 innate lymphoid cell (ILC3) in an IL-1β-dependent manner. Collectively, our study defined a critical pathogenic AM-ILC3-Neu axis and highlighted a paradoxical dual role of AM in driving host defense and pulmonary immunopathology through dynamic phenotypic switching. We suggested that targeting AM phenotypic plasticity represented a promising therapeutic strategy to alleviate excessive immunopathology while preserving protective host defense during hvAB pneumonia.
Abstract Neurons face a fundamental proteostasis challenge: synapses and axons located far from the soma must rapidly remodel their proteome during activity, stress, and development. While local protein synthesis has long been recognized as essential for meeting these demands, classical models largely focused on ribonucleoprotein granules as autonomous carriers of translationally silent mRNAs, treating membranous organelles as parallel logistics or metabolic systems. Recent work overturns this view, revealing that endosomes, lysosomes, axonal endoplasmic reticulum, mitochondria, and their contact sites actively function as mobile translation platforms. In this review, we propose an RBP-centered framework in which phase-separated condensates physically tether specific mRNA cohorts to organelle surfaces, coupling mRNA transport, translational control, and organelle dynamics into a unified network. By organizing recent discoveries into functional modules—long-range transport, localized translation, and stress buffering—this neuron-focused framework identifies organelle-anchored translation factories as a unifying principle of synaptic proteostasis and a broadly applicable design paradigm for highly polarized cells. Keywords RNA-binding proteins, ribonucleoprotein granules, neuronal local translation, organelle-anchored translation
CRISPR-Cas systems provide adaptive immunity in prokaryotes, yet how multiple CRISPR-Cas subtypes coexist and coordinate within a single genome remains unclear. Comparative genomic analysis revealed that nearly one-third of type I-A CRISPR-Cas3 systems are adjacent to a type I-B system, often sharing a single CRISPR array. Using Thermococcus siculi RG-20 (Tsi) as a model, we show that purified TsiCas6a and TsiCas6b independently recognize and cleave the shared pre-crRNA, producing mature crRNAs with comparable efficiency. Plasmid interference assays further demonstrated that crRNAs produced by either Cas6a or Cas6b enzyme could guide both type I-A and type I-B interference complexes. This interchangeability shows that crRNAs generated by either Cas6a or Cas6b can be loaded into, and function with, both type I-A and type I-B interference complexes. Structural modelling revealed distinct but complementary recognition strategies for Cas6a and Cas6b, and mutational analysis of their RNA-binding residues impaired pre-crRNA cleavage and abolished interference activity. Together, these results uncover a shared-array logic in which Cas6a/Cas6b-compatible processing routes a single pre-crRNA to multiple type I effectors-providing a potential mechanism for subtype co-existence, a plausible explanation for array-less (or "orphan") interference modules, and an evolutionary bet-hedging strategy that prevents Acrs from shutting down immunity wholesale.
Lipid droplets (LDs) are intracellular organelles that dynamically regulate lipid and energy homeostasis, mediate hormone production, produce inflammation signaling, while also participating in numerous biological processes and activities. Dysregulation of LD homeostasis is linked to various diseases, such as lipodystrophy, obesity, type 2 diabetes (T2D), cardiovascular diseases (CVD), metabolic dysfunction-associated steatotic liver disease (MASLD), neuronal diseases, and among others. The core of LDs consists of neutral lipids, including triacylglycerol (TAG), cholesteryl esters (CE), and retinyl esters (RE), which are encircled by a monolayer of phospholipid membrane decorated with a subset of LD proteins, both resident as well as dynamic, that vary in different cells, tissues, organs, and organisms. Over the past two decades, together with technological advances, significant achievements have been made in understanding LD biology, including their lifecycle: such as biogenesis, growth/expansion, fusion, and degradation, as well as their diversity and heterogeneity, under both physiological and pathological conditions. In this review, we summarize the current knowledge and methodologies of LD biology in animal cells, and also provide guiding questions, hopefully bringing new directions for future study of LDs and for potential therapeutic treatments for LD-related human diseases.
African swine fever virus (ASFV) is a lethal pathogen that triggers uncontrolled cytokine storms and severe immunopathology. However, the viral factors responsible for systemic inflammation remain unclear. Here, we show that the ASFV-encoded proteins MGF_110-3L and MGF_110-4L are secreted via the conventional ER-Golgi pathway. Single-cell RNA sequencing of porcine PBMCs revealed that MGF_110-3L preferentially activates inflammatory responses in monocytic cells. Mechanistically, both proteins bind Toll-like receptor 2 (TLR2) and signal through TLR2/TLR1 and TLR2/TLR6 heterocomplexes, with the co-receptor CD14 enhancing ligand recognition and signal amplification. These interactions activate MyD88-dependent NF-κB signaling, leading to robust induction of proinflammatory cytokines. ASFV strains lacking either MGF_110-3L or MGF_110-4L cause attenuated cytokine responses in vitro and impaired inflammatory pathology in pigs. Together, these findings establish MGF_110-3L and MGF_110-4L as secreted virulence factors that subvert innate immune recognition and drive lethal inflammation, highlighting their potential as targets for antiviral and vaccine development.
Myocardial infarction results in the loss of a massive amount of cardiomyocytes (CMs), ultimately leading to heart failure (HF). Although current medical therapeu tics can alleviate the symptoms of myocardial infarction, they cannot eliminate the ischemia-induced CM loss (Hume et al., 2023). In adult mammals, the reparatory response to myocardial infarction is the formation of scar tissue, which compromises heart function (Ebrahimi, 2018). For decades, the adult heart has been regarded as a post-mitotic organ, with CMs typically characterized as terminally differentiated that are unable to proliferate (Hashmi and Ahmad, 2019). In contrast to adult mam mals, adult zebrafish possess the full capacity of car diac regeneration after ventricular resection (Poss et al., 2002). In the first 7 days after birth, the neonatal mouse heart can also fully regenerate. However, this regenera tive ability is quickly lost after birth for 7 days (Porrello et al., 2011). In the regenerating hearts, CMs undergo dedifferentiation and proliferation to supply new CMs (Jopling et al., 2010; Kikuchi et al., 2010). Elucidating the mechanisms underlying CM proliferation and cardiac regeneration is of paramount significance. A wealth of research works has demonstrated a close correlation between CM proliferation and metabolism. The induc tion of metabolic reprogramming in CMs to promote cardiac regeneration has emerged as a cutting-edge and highly active area of research works within the field of cardiac regeneration.
While the cell surface has long been studied in the context of its critical cell biology roles for mechanisms that relate to proteins, lipids, and glycans, only recently has RNA become an active player. Classically, surface-presented biopolymers such as proteins and lipids are glycosylated, facilitating their trafficking, folding, and biological activity in the extracellular space. The discovery that small non coding RNAs also serve as templates for N-glycosylation (glycuronans (Flynn et al. 2019; 2021)) and the later iden tification of a covalent linkage between these two biopoly mers (Xie et al. 2024) provided a framework to consider a broader suite of cell surface RNA biology.
Acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) in non-small cell lung cancer remains a pressing clinical challenge. Liquid-liquid phase separation has emerged as a new mechanism of drug resistance, yet its role in EGFR-TKI resistance in lung cancer is largely unexplored. Herein, we prioritized Sequestosome 1 (SQSTM1/p62) as a key condensate by integrating proteomics data from EGFR-TKI resistance cell lines and clinical biopsy specimens, in which the cytoplasmic p62 condensate formation positively correlated with EGFR-TKI resistance. Domain mapping demonstrated that the PB1 and UBA domains of p62 were critical for promoting phase separation and reducing sensitivity to EGFR-TKI treatment, whereas S403 phosphorylation promoted p62 condensation and EGFR-TKI resistance. Further xenograft studies validated that reduction of p62 condensation restores EGFR-TKI sensitivity. Kinase enrichment and interaction assays identified TBK1 as an upstream regulator of p62 S403 phosphorylation to promote p62 condensation after pharmacological inhibition of EGFR. Notably, among the five compounds identified from a drug library screen that both disrupted p62 condensate formation and inhibited the viability of resistant cells, the highly selective TBK1 inhibitor GSK8612 stood out. GSK8612 inhibited p62 S403 phosphorylation and consequently, at subcytotoxic doses, synergized with EGFR-TKIs to suppress the viability of resistant cells and tumor growth in the xenograft mouse model. These findings propose TBK1-p62 axis links p62 condensate homeostasis to EGFR-TKI resistance as an underlying mechanism of action and an emerging strategy to resensitize EGFR-TKI treatment in resistant lung cancer.
