Intratumor bacteria represent an understudied yet influential component of the cancer ecosystem, critically impinging cancer progression. In PyMT breast tumors, we find intracellular bacteria, when residing in cancer cell cytosol, promote metastasis by triggering cytosolic double-stranded DNA (dsDNA) accumulation, which in turn activates the tumor intrinsic cGAS-STING-interleukin (IL)-17B pathway and redirects neutrophils toward a protumor phenotype that inhibits cytotoxic T cells. By contrast, the same strain of bacteria, when present extracellularly, induces antitumor neutrophil activity without engaging the STING pathway. Physiologically, eliminating intracellular bacteria, or therapeutically introducing extracellular bacteria components, abrogates immunosuppression and prevents postsurgical metastatic recurrence in preclinical models. Clinically, the bacteria invasion signature we have developed is associated with poor prognosis in patients with breast cancer. In summary, the spatial interplay between bacteria and host cells in metastatic niches can shape divergent tumor immunity, highlighting bacterial-host engagement as a crucial determinant of cancer immune regulation and a potential therapeutic target.

As a highly conserved post-translational modification, lysine crotonylation (abbreviated as Kcr) regulates diverse biological activities. To explore its role in plant osmotic stress resistance, we performed 4D-FastDIA-based crotonylome profiling on nuclear proteins of Populus davidiana x P. bolleana under osmotic stress. We identified 10,527 crotonylated peptides corresponding to 11,246 Kcr sites on 3606 proteins. Among these, 3020 sites on 1646 proteins were osmotic stress-responsive, with 958 upregulated and 688 downregulated. Among these modifications, H2A, H2B, H3, and H4 all exhibit crotonylation. KEGG analysis revealed 124 differentially modified proteins enriched in five pathways, including glyoxylate and dicarboxylate metabolism, suggesting their potential role in osmotic adaptation. Furthermore, 29 transcription factors (TFs) exhibited osmoticresponsive Kcr modifications. We studied 5 TFs, finding that their osmotic stress resistance significantly decreased upon removal of crotonylation, underscoring its regulatory importance. This study provides the evidence that non-histone crotonylation modulates plant osmotic resistance. In osmotic adaptation of woody plants, a vast network of non-histone Kcr modifications exists alongside traditional histone modifications. While histone Kcr modifications likely act as top-level regulators of stress responses, the extensive non-histone Kcr modifications may fulfill specific functional roles. These findings offer new insights into the mechanisms of plant stress responses.

This research examines how the basic leucine zipper (bZIP) transcription factor (TF) influences drought stress responses in tree species, emphasizing its related regulatory pathways, and thus offering a theoretical framework for understanding drought response mechanisms regulated by the bZIP TF family. Specifically, we characterized the functional role of the S subfamily bZIP gene, PtrbZIP12, from Populus trichocarpa, by developing transgenic poplars that either overexpressed or knocked down PtrbZIP12. The findings indicated that PtrbZIP12 markedly improved drought tolerance in transgenic plants by facilitating reactive oxygen species scavenging, enhancing proline biosynthesis, and reducing plasma membrane peroxidation and cell death. To pinpoint PtrbZIP12's downstream targets, RNA sequencing was performed, followed by chromatin immunoprecipitation-PCR (ChIP-PCR), yeast one-hybrid, and dual-luciferase assays. These analyses confirmed that PtrbZIP12 binds directly to the promoters of PtrDHN (dehydrin) and PtrPOD (peroxidase), leading to the activation of their expression. Transgenic poplars overexpressing (OE) PtrDHN or PtrPOD were subsequently generated, and similar to PtrbZIP12, their OE conferred enhanced drought tolerance. Moreover, coexpression of PtrbZIP12 with PtrbZIP3 further elevated PtrDHN transcript levels, resulting in improved drought resilience in the PtrbZIP12 transgenic lines. Moreover, phosphorylation was identified as a key factor in boosting PtrbZIP12-mediated transcriptional regulation of PtrPOD and PtrDHN, underscoring the significance of posttranslational modification in plant drought stress responses.

Plant basic Leucine Zipper (bZIP) proteins are crucial in growth, development, and responses to various abiotic stresses. Extensive research has demonstrated that bZIP transcription factors play a crucial role in mediating plant responses to drought stress. However, to systematically analyze the functions of its family genes, the specific functions of these factors in the drought response of poplar require further investigation. In this study a "Shanxin" poplar bZIP gene, PdabZIP69, was identified which is significantly upregulated under drought stress. Transgenic lines over-expressed PdabZIP69 (OE) were generated through Agrobacterium-mediated transformation. Drought stress assays revealed that PdabZIP69-OE plants exhibit enhanced drought tolerance and reduced leaf water loss compared to wild-type (WT). Physiological analyses revealed that the OE plants exhibited significantly reduced levels of hydrogen peroxide (H2O2) and malondialdehyde (MDA), elevated proline content, as well as enhanced activities of superoxide dismutase (SOD) and peroxidase (POD) when compared to the WT. These results indicate that the augmented reactive oxygen species (ROS) scavenging capacity in PdabZIP69-OE plants confers greater drought tolerance. Y1H, ChIP-PCR and Dual-luciferase assay revealed that PdabZIP69 can bind to the promoter of the PdbbHLH1 gene, a positive regulator of drought tolerance in "Shanxin" poplar, and activate its expression. Collectively, PdabZIP69 enhances drought tolerance in poplar by regulating the antioxidant system and the expression of the PdbbHLH1 gene, providing genetic resources for the systematic study of the regulation of drought resistance of bZIP family members.

Novel SBCP chromatin precipitation bypasses nuclear isolation to recover abundant intact chromatinproteins and enhance ChIP performance in diverse plants. Chromatin isolation remains a major technical bottleneck in plant molecular biology, often compromising studies on genome regulation. We present SBCP (Saline-Based Chromatin Precipitation), a method that directly isolates native chromatin by exploiting its differential solubility in salt solutions, thereby eliminating the need for prior nuclear purification. Compared with the traditional workflow that relies on nuclear isolation, SBCP achieves substantially higher yields (3.4-fold in poplar, 2.4-fold in Arabidopsis) and superior genomic DNA purity, while reducing chloroplast contamination by 63–73

Background Liquid-liquid phase separation (LLPS) has emerged as an important regulatory mechanism in cancer biology and has been implicated in hepatocellular carcinoma (HCC) progression. LLPS has also been linked to ferroptosis, a regulated cell death process with tumor-suppressive potential. However, how LLPS-related regulators modulate ferroptosis in HCC remain unclear. Here, we report the precise mechanism of TRIM21 in regulating the susceptibility of HCC to ferroptosis. Methods Bioinformatics analysis was employed to evaluate TRIM21 expression in HCC and identify ferroptosis suppressors potentially associated with TRIM21. In vitro experiments were then conducted to assess the impact of TRIM21 on cell malignant phenotypes. Finally, immunofluorescence and FRAP experiments were carried out to investigate the phase separation phenomenon of TRIM21. Results TRIM21 was found to be upregulated in HCC and associated with prognosis. Bioinformatic screening identified EZH2 as a candidate ferroptosis suppressor associated with TRIM21. TRIM21 knockdown enhanced ferroptosis sensitivity in HCC cells and suppressed malignant phenotypes. Mechanistically, TRIM21 underwent LLPS that depended on its coiled-coil domain (a predicted IDR region). Co-immunoprecipitation assays and immunofluorescence further supported a molecular association between TRIM21 and EZH2. Disruption of LLPS by TRIM21 mutant effectively abolished its cytoplasmic colocalization with EZH2, thereby attenuating the invasive capacity of HCC cells. Conclusion This study demonstrates that TRIM21 inhibits ferroptosis through LLPS, thereby promoting HCC cell survival. These findings reveal a previously unrecognized link between TRIM21 LLPS, ferroptosis, and HCC progression, providing a rational basis for targeting LLPS mechanisms to treat HCC.

Although ERF transcription factors (TFs) play critical roles in abiotic stress tolerance, the molecular mechanisms underlying this role remain incompletely understood. This study identified BpERF1A in birch (Betula platyphylla) as a drought-responsive TF through co-expression regulatory network analysis. Expression of BpERF1A was stalwartly induced by drought stress, and drought treatment markedly boosted its promoter activity. Functional analyses demonstrated that overexpression of BpERF1A markedly improved drought tolerance compared with wild-type (WT) birch, whereas its repression increased drought sensitivity. Overexpression lines also exhibited higher antioxidant enzyme activities and proline content relative to WT. Chromatin immunoprecipitation-polymerase chain reaction, yeast one-hybrid and dual-luciferase (dual-LUC) assays confirmed that BpERF1A directly binds to G-box elements in the promoters of BpDHN (dehydrin) and BpAOS (allene oxide synthase), activating their transcription. Furthermore, overexpression of BpDHN/BpAOS enhanced drought tolerance and promoted reactive oxygen species (ROS) scavenging in transgenic plants. Protein-protein interaction analysis using bimolecular fluorescence complementation revealed that BpERF1A interacts with BpRAV1 to form a heterodimer, which further enhances BpERF1A binding to the BpDHN promoter and its transcriptional activation. Collectively, these findings establish a central role for the BpERF1A regulatory module in drought acclimation and highlight its synergistic interaction with BpRAV1, providing an effective strategy to enhance drought tolerance through improved ROS scavenging capacity.

Background:Circular RNAs (circRNAs) play a pivotal role in the development and advancement of various cancer types. However, the involvement of circ-PAN3 in hepatocellular carcinoma (HCC) is not well understood. To shed light on this, we conducted a comprehensive study through biochemistry, cell biology, molecular biology, and bioinformatics techniques to investigate the role of circ-PAN3 and its associated pathway in the progression of HCC. Methods:Cell Counting Kit-8 (CCK-8) assay and colony formation assay were utilized to evaluate cell proliferation; Quantitative real-time PCR (RT-qPCR) and Western blot were adopted for assessing mRNA and protein expression; Annexin V/propidium iodide (PI) staining was applied to detect cellular apoptosis; CircInteractome and Targetscan databases were searched to predict potential targets of circRNA and miRNA; Luciferase reporter assay and RNA pull-down assay were performed to examine the interaction of RNA molecules. Conclusions:Our findings revealed a significant increase in circ-PAN3 expression in HCC clinical specimens, which correlated with a poor survival rate in HCC patients. Knockdown of circ-PAN3 resulted in impaired cell proliferation, reduced cell survival, and inhibited tumorigenesis of HCC in vivo. Further analysis demonstrated that circ-PAN3 could serve as a sponge for miR-153, leading to a decrease in its expression level. This in turn upregulated cyclin D1 and ultimately promoted the proliferation of HCC cells. Additionally, overexpression of cyclin D1 mitigated the inhibitory effect on HCC proliferation induced by circ-PAN3 knockdown. Our study highlights the presence of a novel circ-PAN3/miR-153/cyclin D1 regulatory axis that plays a crucial role in the progression of HCC.

The systematic identification and functional characterization of noncanonical translation products, such as novel peptides, will facilitate the understanding of the human genome and provide new insights into cell biology. Here, we constructed a high-coverage peptide sequencing reference library with 11,668,944 open reading frames and employed an ultrafiltration tandem mass spectrometry assay to identify novel peptides. Through these methods, we discovered 8945 previously unannotated peptides from normal gastric tissues, gastric cancer tissues and cell lines, nearly half of which were derived from noncoding RNAs. Moreover, our CRISPR screening revealed that 1161 peptides are involved in tumor cell proliferation. The presence and physiological function of a subset of these peptides, selected based on screening scores, amino acid length, and various indicators, were verified through Flag-knockin and multiple other methods. To further characterize the potential regulatory mechanisms involved, we constructed a framework based on artificial intelligence structure prediction and peptide‒protein interaction network analysis for the top 100 candidates and revealed that these cancer-related peptides have diverse subcellular locations and participate in organelle-specific processes. Further investigation verified the interacting partners of pep1-nc-OLMALINC, pep5-nc-TRHDE-AS1, pep-nc-ZNF436-AS1 and pep2-nc-AC027045.3, and the functions of these peptides in mitochondrial complex assembly, energy metabolism, and cholesterol metabolism, respectively. We showed that pep5-nc-TRHDE-AS1 and pep2-nc-AC027045.3 had substantial impacts on tumor growth in xenograft models. Furthermore, the dysregulation of these four peptides is closely correlated with clinical prognosis. Taken together, our study provides a comprehensive characterization of the noncanonical proteome, and highlights critical roles of these previously unannotated peptides in cancer biology.

The dendritic cell (DC)-initiated and sustained cancer immunity cycle is indispensable for effective endogenous and therapeutically mobilized antitumour T cell responses1-8. This necessitates the continuous migration of antigen-carrying DCs from the tumour microenvironment (TME) to the tumour draining lymph nodes (tdLNs)7-13. Here, through longitudinal analysis of human and mouse tumours, we observed a progressive decrease in migratory conventional DCs (mig-cDCs) in the tdLNs during tumour progression. This decline compromised tumour-specific T cell priming and subsequent T cell supply to the TME. Using a genome-wide in vivo CRISPR screen, we identified phosphodiesterase 5 (PDE5) and its substrate cyclic guanosine monophosphate (cGMP) as key modulators of DC migration. Advanced tumours disrupted cGMP synthesis in DCs to decrease their motility, while PDE5 perturbation preserved the cGMP pool to restore DC migration. Mechanistically, cGMP enhanced myosin-II activity through Rho-associated factors, extending the paradigm of cGMP-regulated amoeboid migration from Dictyostelium to mammalian immune cells. Pharmacological inhibition of PDE5 using sildenafil restored mig-cDC homing to late-stage tdLNs and sustained antitumour immunity in a DC-dependent manner. Our findings bridge fundamental DC interstitial motility to antitumour immunity, revealing that its disruption in chaotic TME promotes immune evasion, and its enhancement offers a promising direction for DC-centric immunotherapy.

Osteoarthritis (OA) affects nearly 500 million people worldwide and is characterized by an irreversible loss of glycosaminoglycans (GAGs) at articular cartilage surfaces, which are essential in maintaining cartilage mechanical properties and chondrocyte phenotypes. Despite advances, preserving cartilage GAGs and controlling their turnover in living cells remain challenging. On the basis of the hypothesis that GAGs can interact with cationic molecules, we demonstrated a cost-effective strategy to increase human cartilage GAGs using a cationic polymer hexadimethrine bromide (HDMBr). HDMBr promoted chondrogenesis of mesenchymal stem cells by attracting pericellular GAGs and up-regulating vesicle formation, leading to increased matrix secretion. HDMBr also acted like a molecular assembler to promote the assembly of chondroitin sulfate (CS) into highly concentrated condensates during intracellular trafficking, resulting in more efficient GAG secretion. HDMBr was then evaluated as a potential therapeutic in two animal models. In a rabbit model of large cartilage defects, HDMBr promoted the intrinsic regeneration of GAG-rich hyaline-like cartilage and improved tissue integration. In a rat model of OA, low-dose HDMBr treatment increased cartilage thickness, supported cartilage matrix homeostasis, and supported cell-based therapy, reducing OA damage as compared with other tested clinical treatments. Overall, this study introduces a cost-effective GAG manipulation approach to cartilage repair and joint preservation, offering insights into the mechanisms of cell-material interactions.

Radiation therapy (RT) is a key treatment strategy for lung cancer, yet its efficacy is frequently compromised by radioresistance. The combination of RT with targeted therapies enhances treatment outcomes for non-small cell lung cancer (NSCLC). This study aims to investigate new mechanisms of metastasis after RT for NSCLC and improve the durability of the benefits of radiotherapy for lung cancer patients. This integrative study utilized human NSCLC tissue arrays, bulk RNA-sequencing, CUT Tag sequencing, and single-cell RNA-sequencing to identify gene alterations induced by RT. In vitro experiments and animal studies were used to investigate the role of Jumonji domain-containing 6 (JMJD6)/ETS homologous factor (EHF) axis in post-RT metastasis of NSCLC. RT triggered the upregulation of JMJD6 in NSCLC tissues. This upregulation led to the activation of EHF and the subsequent transcription of pluripotency factor genes through the demethylation of H4R3me2s. JMJD6/EHF axis plays a critical role in NSCLC cell metastasis, potentially through the TGF-β/SMAD and AKT/ERK signaling pathways. These findings suggest JMJD6 as a potential therapeutic target to combat post-RT metastasis in NSCLC.

Background:Inflammation plays a pivotal role in modulating the pathophysiological progression of myocardial injury and the subsequent repair and remodeling of the infarcted myocardium. Cathepsin B, a member of the cysteine protease family, has been recognized for its ability to initiate various signaling cascades essential to inflammatory processes. This study aims to investigate whether cathepsin B influences cardiomyocyte survival under inflammatory conditions. Methods:Mice were randomly divided into four groups (n = 6 per group) based on whether they received an intraperitoneal injection of Ca-074 Me (50 mg/kg) and whether myocardial ischemia/reperfusion (I/R) surgery was performed. Results:The cathepsin B-specific inhibitor Ca-074 Me significantly attenuated myocardial infarction caused by I/R in vivo and reduced hypoxia-induced cardiomyocyte apoptosis in vitro. Mechanistically, Ca-074 Me appeared to inhibit caspase-3 signaling, thereby mitigating cardiomyocyte apoptosis under chemical hypoxia induced by cobalt chloride or physical oxygen deprivation. Conclusion:Targeted inhibition of cathepsin B may represent an innovative strategy for the amelioration of myocardial injury.

Differentially expressed genes involved in tension wood in ‘Shanxin’ poplar have been identified, and gene expression regulatory network has been established, laying the foundation for research on the molecular mechanisms. Tension wood (TW), a specialized wood structure, forms in woody angiosperms under mechanical force or gravity. Studying the transcriptional regulation of genes during TW formation is valuable for molecular biology research and the genetic improvement of wood properties. In this study, TW xylem cell walls in ‘Shanxin’ poplar (Populus davidiana × P. alba var. pyramidalis) exhibited a relatively higher cellulose content and a distinct gelatinous layer (G-layer), as identified by safranin-fast green staining, compared to opposite wood (OW) and normal wood (NW). Transcriptome analysis of TW, OW, and NW yielded 52,697 unigenes. Differential expression analysis identified 2,321, 462, and 2,683 differentially expressed genes(DEGs)in TW_NW, OW_NW, and TW_OW comparisons, respectively. GO enrichment analysis of DEGs highlighted their involvement in secondary processes including cell wall organization, response to stimulus, hormone-mediated signaling pathway, cell wall, transcription regulator activity and oxidoreductase activity. KEGG enrichment analysis revealed significant pathways including phenylpropanoid biosynthesis, flavone and flavonol biosynthesis, plant hormone signal transduction, and starch and sucrose metabolism. Specifically, 13 out of 16 DEGs were noted in the glucose synthesis pathway within the starch and sucrose metabolism. A two-layer gene expression regulatory network (GRN) was constructed, involving 6 transcription factors (TFs) and 37 functional genes, totaling 181 interactions. Validation using ChIP-PCR and RT-qPCR for a total of 35 interactions involving PdaMYB52, PdaERF6, and PdaERF17 demonstrated a 91

Cardiovascular disease remains the leading cause of death worldwide, with atherosclerosis (AS) serving as a critical underlying pathological process and major risk factor. Regular physical exercise is widely recognized as an effective strategy to reduce the risks and severity of AS, yet the precise molecular mechanisms through which exercise exerts its protective effects are still not fully understood. MicroRNAs (miRNAs), key regulators of gene expression, play integral roles in the progression of AS by influencing vascular function, lipid metabolism, and inflammation. The exercise-induced improvement of AS is a complex process, with miRNAs playing essential roles not only within cells and tissues but also circulating stably in the bloodstream as novel signaling molecules. These circulating miRNAs mediate communication between organs and tissues, acting as potential biomarkers that could provide deeper, systemic insights into the metabolic benefits of exercise. In this review, we explore recent advancements in our understanding of how exercise affects both intracellular and circulating miRNAs. We emphasize how exercise-regulated miRNAs contribute to endothelial function, promote lipid metabolism across various metabolic organs, and reduce monocyte-mediated systemic inflammation, while also addressing their role in alleviating frailty. Circulating miRNAs, which dynamically reflect tissue-specific responses to exercise, hold great promise as diagnostic and prognostic biomarkers for AS. Moreover, we discuss the challenges and future directions in this field, aiming to uncover how exercise-induced miRNA modulation could offer innovative therapeutic strategies for the prevention and treatment of AS.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mainly infects the tissues of the human respiratory, nervous, digestive and urinary systems. Here, we conducted an integrated study of the viral–host protein‒protein interactions between 29 wildtype SARS-CoV-2 proteins and 15 SARS-CoV-2 variant S proteins and human lung-, bronchus-, neuron-, liver- and kidney-derived cells, identified 745,163 viral–host protein‒protein interactions and established phylointeractomics of SARS-CoV-2 variants across susceptible human organs. We highlighted interacting proteins involved in the function evolution of viral proteins under different cellular contexts, and identified several as potential drug targets for treating related complications and sequalae. The results reveal how various viral proteins contribute to the infection, replication and immune evasion by interacting with general and cell type-specific host proteins.

The basic helix-loop-helix (bHLH) family of transcription factors (TFs) plays a critical role in regulating plant resistance to various abiotic stresses, including drought, low temperature, high salinity, and iron deficiency, as well as in overseeing key processes in plant growth and development. Despite the well-documented functions of bHLH TFs in many plant species, their roles in Populus davidiana x P. bolleana under abiotic stress conditions remain largely unexplored. This study identified a bHLH gene, designated as PdbbHLH1, whose expression is markedly upregulated in response to PEG6000 treatment. To investigate its function, the research generated transgenic Populus plants overexpressing (OE) PdbbHLH1 through Agrobacterium tumefaciens-mediated transformation. Stress experiments demonstrated that OE PdbbHLH1 significantly enhanced the drought resistance of these transgenic plants, as evidenced by increased fresh weight and chlorophyll content compared to wild-type (WT) plants under drought conditions. Physiological analyses further revealed that PdbbHLH1-OE plants exhibited lower levels of hydrogen peroxide (H2O2) and malondialdehyde (MDA), indicating reduced cellular damage and a more robust reactive oxygen species (ROS) scavenging capability than WT plants. Furthermore, the expression of drought resistance-related genes, such as PdbAAO, PdbGST, and PdbPOD, was significantly upregulated in OE plants post-drought stress. Notably, PdbbHLH1 was shown to directly bind to the promoters of PdbPOD1 and PdbPOD4, activating their expression and thereby enhancing the drought resistance of transgenic Populus davidiana x P. bolleana. Collectively, these results suggest that PdbbHLH1 is a key regulator of drought tolerance in Populus, modulating this response through the enhancement of antioxidant enzyme activities at the physiological level.

B cell acute lymphoblastic leukemia (B-ALL) is the most prevalent type of cancer in young children and is associated with high levels of reactive oxygen species (ROS). The antioxidant N-acetylcysteine (NAC) was tested for its ability to alter disease progression in a mouse model of B-ALL. Mb1-CreΔPB mice have deletions in genes encoding PU.1 and Spi-B in B cells and develop B-ALL at 100% incidence. Treatment of Mb1-CreΔPB mice with NAC in drinking water significantly reduced the frequency of CD19 + pre-B ALL cells infiltrating the thymus at 11 weeks of age. However, treatment with NAC did not reduce leukemia progression or increase survival by median 16 weeks of age. NAC significantly altered gene expression in leukemias in treated mice. Mice treated with NAC had increased frequencies of activating mutations in genes encoding Janus Kinases 1 and 3. In particular, frequencies of Jak3 R653H mutations were increased in mice treated with NAC compared to control drinking water. NAC opposed oxidization of PTEN protein ROS in cultured leukemia cells. These results show that NAC alters leukemia progression in this mouse model, ultimately selecting for leukemias with high Jak3 R653H mutation frequencies.