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Methyltransferase-like 3 (METTL3) is the key subunit of methyltransferase complex responsible for catalyzing N6-methyladenosine (m6A) modification on mRNA, which is the most prevalent post-transcriptional modification in eukaryotes. In this study, we utilized online databases to analyze the association between METTL3 expression and various aspects of tumorigenesis, including gene methylation, immunity, and prognosis. Our investigation revealed that METTL3 serves as a prognostic marker and therapeutic target for liver hepatocellular carcinoma (LIHC). Through experimental studies, we observed frequent upregulation of METTL3 in LIHC tumor tissue and cells. Subsequent inhibition of METTL3 using a novel small molecule inhibitor, STM2457, significantly impeded tumor growth in LIHC cell lines, spheroids, and xenograft tumor model. Further, transcriptome and m6A sequencing of xenograft bodies unveiled that inhibition of METTL3-m6A altered genes enriched in SMAD and MAPK signaling pathways that are critical for tumorigenesis. These findings suggest that targeting METTL3 represents a promising therapeutic strategy for LIHC. [ABSTRACT FROM AUTHOR]

Forsythia suspensa is a deciduous shrub that belongs to the family Myrtaceae, and its dried fruits are used as medicine. F. suspensa contains several secondary metabolites, which exert pharmacological effects. One of the main active components is forsythin, which exhibits free radical scavenging, antioxidant, anti-inflammatory, and anti-cancer effects. Mitogen-activated protein kinase (MAPKs) can increase the activity of WRKY family transcription factors in a phosphorylated manner, thereby increasing the content of secondary metabolites. However, the mechanism of interaction between MAPKs and WRKYs in F. suspensa remains unclear. In this study, we cloned the genes of FsWRKY4 and FsMAPK3, and performed a bioinformatics analysis. The expression patterns of FsWRKY4 and FsMAPK3 were analyzed in the different developmental stages of leaf and fruit from F. suspensa using real-time fluorescence quantitative PCR (qRT-PCR). Subcellular localization analysis of FsWRKY4 and FsMAPK3 proteins was performed using a laser scanning confocal microscope. The existence of interactions between FsWRKY4 and FsMPAK3 in vitro was verified by yeast two-hybridization. Results showed that the cDNA of FsWRKY4 (GenBank number: OR566682) and FsMAPK3 (GenBank number: OR566683) were 1587 and 522 bp, respectively. The expression of FsWRKY4 was higher in the leaves than in fruits, and the expression of FsMAPK3 was higher in fruits but lower in leaves. The subcellular localization results indicated that FsWRKY4 was localized in the nucleus and FsMAPK3 in the cytoplasm and nucleus. The prey vector pGADT7-FsWRKY4 and bait vector pGBKT7-FsMAPK3 were constructed and co-transferred into Y2H Glod yeast receptor cells. The results indicated that FsWRKY4 and FsMAPK3 proteins interact with each other in vitro. The preliminary study may provide a basis for more precise elucidation of the synthesis of secondary metabolites in F. suspensa. [ABSTRACT FROM AUTHOR]

Background: Gegen Qinlian decoction (GQD) is a classic prescription for treating ulcerative colitis (UC) in traditional Chinese medicine. However, the therapeutic mechanism has not been fully clarified. Purpose: In the present study, we aimed to evaluate the role of ferroptosis-mediated IEC death in UC treated mice with GQD by using DSS-induced a colitis mouse model and RSL3-induced ferroptosis in intestinal organoids. Methods: The effects of GQD on DSS-treated colitis were examined via daily body weight, DAI, colon length, HE staining, PAS staining, ZO-1 and Occludin immunohistochemical staining. Ferroptosis was determined by analysis of iron load, MDA, GSH, mitochondrial morphology, and expression of ferroptosis-associated proteins (GPX4, SLC7A11 and ACSL4). Results: In vivo, GQD administration reduced body weight loss and DAI scores, increased colon length, and improved intestinal histological characteristics and epithelial barrier dysfunction. GQD administration obviously improved the levels of ferroptosis markers (iron load, MDA, GSH, and mitochondrial morphology) and the expression of ferroptosis-associated proteins (GPX4, SLC7A11 and ACSL4). Consistent with in vivo results, GQD administration partially reversed the levels of mtROS, Fe2+ and MDA in intestinal organoids induced by RSL3, and notably improved morphological destruction, histological damage and epithelial barrier dysfunction in organoids. Conclusions: In this study, we demonstrated that ferroptosis was triggered in DSS-induced experimental colitis and that GQD adiministration could protect against colonic damage and intestinal epithelial barrier dysfunction by inhibiting ferroptosis. [ABSTRACT FROM AUTHOR]

Background: Melanoma is one of the most malignant skin carcinomas with high metastatic potential. Increasing evidence has demonstrated that β-tubulin 4A (TUBB4A) plays a key role in the development and progression of several types of human cancer. However, the potential function of TUBB4A in cutaneous melanoma remains to be determined. Methods: We first performed a differential expression analysis based on skin melanoma tissues and normal tissues from Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) datasets and then a survival analysis to identify prognostic-related key genes. We further conducted in vitro biochemical experiments to verify the functional roles of the key gene TUBB4A. Two small-molecule inhibitors of TUBB4A, Dihydroartemisinin (DHA) and Nocodazole, were used to validate the effect of TUBB4A on the apoptosis and cell cycle of melanoma cells. Results: We found that TUBB4A expression was positively correlated to the overall survival (OS) of cutaneous melanoma patients. The coexpressed genes with TUBB4A were enriched in melanoma-related pathways and functions. The experimental results showed that knockdown of TUBB4A inhibited the proliferation and migration of A375 and B16-F10 melanoma cells. Moreover, DHA and Nocodazole promoted the apoptosis of melanoma cells and blocked the melanoma tumor cell cycle in the G2/M stage. Conclusion: TUBB4A may be a prognostic biomarker and therapeutic target for melanoma. [ABSTRACT FROM AUTHOR]

The B-BOX (BBX) gene family is widely distributed in animals and plants and is involved in the regulation of their growth and development. In plants, BBX genes play important roles in hormone signaling, biotic and abiotic stress, light-regulated photomorphogenesis, flowering, shade response, and pigment accumulation. However, there has been no systematic analysis of the BBX family in Platanus × acerifolia. In this study, we identified 39 BBX genes from the P. × acerifolia genome, and used TBtools, MEGA, MEME, NCBI CCD, PLANTCARE and other tools for gene collinearity analysis, phylogenetic analysis, gene structure, conserved domain analysis, and promoter cis-element analysis, and used the qRT-PCR and transcriptome data for analyzing expression pattern of the PaBBX genes. Collinearity analysis indicated segmental duplication was the main driver of the BBX family in P. × acerifolia, and phylogenetic analysis showed that the PaBBX family was divided into five subfamilies: I, II, III, IV and V. Gene structure analysis showed that some PaBBX genes contained super-long introns that may regulate their own expression. Moreover, the promoter of PaBBX genes contained a significant number of cis-acting elements that are associated with plant growth and development, as well as hormone and stress responses. The qRT-PCR results and transcriptome data indicated that certain PaBBX genes exhibited tissue-specific and stage-specific expression patterns, suggesting that these genes may have distinct regulatory roles in P. × acerifolia growth and development. In addition, some PaBBX genes were regularly expressed during the annual growth of P. × acerifolia, corresponding to different stages of flower transition, dormancy, and bud break, indicating that these genes may be involved in the regulation of flowering and/or dormancy of P. × acerifolia. This article provided new ideas for the study of dormancy regulation and annual growth patterns in perennial deciduous plants. [ABSTRACT FROM AUTHOR]

The camphor tree (Cinnamomum camphora (L.) Presl.) is the representative species of subtropical evergreen broadleaved forests in eastern Asia and an important raw material for essential oil production worldwide. Although MYBs have been comprehensively characterized and their functions have been partially resolved in many plants, it has not been explored in C. camphora. In this study, 121 CcMYBs were identified on 12 chromosomes in the whole genome of C. camphora and found that CcMYBs were mainly expanded by segmental duplication. They were divided into 28 subgroups based on phylogenetic analysis and gene structural characteristics. In the promoter regions, numerous cis-acting elements were related to biological processes. Analysis of RNA sequencing data from seven tissues showed that CcMYBs exhibited different expression profiles, suggesting that they have various roles in camphor tree development. In addition, combined with the correlation analysis of structural genes in the flavonoid synthesis pathway, we identified CcMYBs from three subgroups that might be related to the flavonoid biosynthesis pathway. This study systematically analyzed CcMYBs in C. camphora, which will set the stage for subsequent research on the functions of CcMYBs during their lifetime and provide valuable insights for the genetic improvement of camphor trees. [ABSTRACT FROM AUTHOR]

Radiation-induced esophageal injury remains a limitation for the process of radiotherapy for lung and esophageal cancer patients. Esophageal epithelial cells are extremely sensitive to irradiation, nevertheless, factors involved in the radiosensitivity of esophageal epithelial cells are still unknown. Terminal uridyl transferase 4 (TUT4) could modify the sequence of miRNAs, which affect their regulation on miRNA targets and function. In this study, we used transcriptome sequencing technology to identify mRNAs that were differentially expressed before and after radiotherapy in esophageal epithelial cells. We further explored the mRNA expression profiles between wild-type and TUT4 knockout esophageal epithelial cells. Volcano and heatmap plots unsupervised hierarchical clustering analysis were performed to classify the samples. Enrichment analysis on Gene Ontology functional annotations and Kyoto Encyclopedia of Genes and Genomes pathways was performed. We annotated differential genes from metabolism, genetic information processing, environmental information processing, cellular processes, and organismal systems human diseases. The aberrantly expressed genes are significantly enriched in irradiation-related biological processes, such as DNA replication, ferroptosis, and cell cycle. Moreover, we explored the distribution of transcription factor family and its target genes in differential genes. These mRNAs might serve as therapeutic targets in TUT4-related radiation-induced esophageal injury. [ABSTRACT FROM AUTHOR]

Diabetic nephropathy (DN) is one of the primary complications of diabetes. Fisetin is a flavonoid polyphenol that is present in several vegetables and fruits. The present study investigated the mechanisms of fisetin in DN-induced podocyte injury both in vitro and in vivo. The results revealed that fisetin ameliorated high glucose (HG)-induced podocyte injury and streptozotocin (STZ)-induced DN in mice. CDKN1B mRNA expression in the glomeruli of patients with DN decreased based on the Nephroseq dataset, and fisetin reversed CDKN1B expression at mRNA and protein levels in a dose-dependent manner in podocytes and mice kidney tissues. Furthermore, fisetin suppressed the phosphorylation of P70S6K, a downstream target of CDKN1B , activated autophagosome formation, and inhibited Nod-like receptor protein 3 (NLRP3) inflammasomes. Interfering CDKN1B reduced the protective effects of fisetin against high glucose-induced podocyte injury. Molecular docking results revealed a potential interaction between fisetin and CDKN1B. In summary, the present study revealed that fisetin alleviated high glucose-induced podocyte injury and STZ-induced DN in mice by restoring autophagy-mediated CDKN1B/P70S6K pathway and inhibiting NLRP3 inflammasome. [ABSTRACT FROM AUTHOR]

The development of CRISPR-based barcoding methods creates an exciting opportunity to understand cellular phylogenies. We present a compact, tunable, high-capacity Cas12a barcoding system called dual acting inverted site array (DAISY). We combined high-throughput screening and machine learning to predict and optimize the 60-bp DAISY barcode sequences. After optimization, top-performing barcodes had ∼10-fold increased capacity relative to the best random-screened designs and performed reliably across diverse cell types. DAISY barcode arrays generated ∼12 bits of entropy and ∼66,000 unique barcodes. Thus, DAISY barcodes—at a fraction of the size of Cas9 barcodes—achieved high-capacity barcoding. We coupled DAISY barcoding with single-cell RNA-seq to recover lineages and gene expression profiles from ∼47,000 human melanoma cells. A single DAISY barcode recovered up to ∼700 lineages from one parental cell. This analysis revealed heritable single-cell gene expression and potential epigenetic modulation of memory gene transcription. Overall, Cas12a DAISY barcoding is an efficient tool for investigating cell-state dynamics. [Display omitted] • DAISY is a CRISPR-Cas12a-based evolvable barcoding technology • Optimization of DAISY barcodes using an iterative machine-learning pipeline • Lentiviral delivery enables scalable, tunable barcoding in multiple cancer lines • Integration with scRNA-seq reveals cell dynamics and transcriptional memory Hughes et al. present a single-cell barcoding technology that uses an iterative machine-learning pipeline to optimize the diversity of editing outcomes generated by Cas12a for lineage tracking. Integration with scRNA-seq revealed features of transcriptional memory and nominated EZH2 as a putative epigenetic regulator. [ABSTRACT FROM AUTHOR]

PLAU is known as a selected serine protease converting plasminogen to plasmin. The role of PLAU in the development of pan-cancer, especially bladder urothelial carcinoma (BLCA) remains unclear. A variety of online tools and cancer databases, including TCGA, GETx, HPA database, GSCALite, UALCAN, ESTIMATE, CIBERSORT, ssGSEA algorithms and SangerBox website, were applied to investigate the associations between PLAU expression and prognosis, genetic alterations, pathway activation, and tumor immunity in pan-cancer. Through cBioPortal and STITCH platforms, the oncogenic role of PLAU and related targeting medicines in BLCA were also explored. We verified the expression of PLAU in pan-cancer cells and its function in bladder cancer cell lines using wet-lab experiments. PLAU expression levels were significantly higher in most cancer tissues. PLAU had a certain accuracy in the diagnosis of various types of cancers (90 % AUC > 0.700). In BLCA, PLAU has abundant methylated sites and showed statistical differences in clinical features. PLAU was involved in tumor immune infiltration, and especially positively correlated with neutrophil infiltration. High-expressed PLAU indicated poorer prognosis in the BLCA patients receiving Atezolizumab. A high mRNA and protein expression levels of PLAU were observed in pan-cancer cell lines, especially BLCA cells. Knockdown of PLAU inhibited the invasive, proliferative, and aggressive phenotypes of bladder cancer cells. Immunohistochemical staining validated PLAU's higher expression in BLCA tissues than in adjacent non-cancerous tissues. And overexpression of PLAU was associated with more advanced TNM stage, and high infiltrating depth. Our study revealed that PLAU can serve as a potential therapeutic target and prognostic marker for various malignancies, especially BLCA. • Integrated analyses on multi-omics characteristics of PLAU on tumor microenvironment, immunotherapy, and prognosis. • A high expression level of Plasminogen Activator was related to a short Overall Survival in most cancer types. • PLAU expression is positively correlated with neutrophils infiltration in cancers. • High-expressed PLAU indicated poorer prognosis in the bladder cancer patients receiving Atezolizumab. [ABSTRACT FROM AUTHOR]