Researchers published a study in the journal Cell Division on November 3, 2025, detailing how the DNA methyltransferase DNMT3B promotes the progression of non-small cell lung cancer (NSCLC). The study identifies a specific molecular mechanism in which DNMT3B suppresses the expression of the transcription factor TAL1 through DNA methylation. This suppression allows for the transcriptional activation of pyruvate kinase M2 (PKM2), a key enzyme in glycolysis, thereby enhancing tumor cell proliferation and survival. The findings suggest that the DNMT3B-TAL1-PKM2 axis represents a critical regulatory pathway in the metabolic reprogramming of lung cancer cells.

The investigation began with bioinformatics analysis of multiple gene expression datasets, including GSE18842, GSE19188, and GSE159857, to identify differentially expressed transcription factors in NSCLC. Among the screened factors, TAL1 was identified as a hub gene that was significantly downregulated in lung cancer tissues compared to normal controls. Further analysis using the UALCAN database confirmed that TAL1 expression was low in both lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC) subtypes. The researchers also observed a negative correlation between TAL1 and PKM2 expression levels in these cancer datasets, prompting an investigation into their functional relationship.

To validate these computational findings, the study utilized human NSCLC cell lines, specifically A549 and H460, alongside the normal lung epithelial cell line BEAS-2B as a control. Quantitative reverse transcription PCR (RT-qPCR) and Western blot analysis confirmed that TAL1 protein and mRNA levels were markedly lower in the cancer cell lines than in the normal control cells. Conversely, PKM2 levels were found to be elevated in the NSCLC cells. These results established a baseline for the subsequent functional assays, indicating that the loss of TAL1 and the gain of PKM2 are characteristic features of the NSCLC cellular phenotype.

Functional experiments demonstrated that the overexpression of TAL1 in A549 and H460 cells significantly inhibited malignant phenotypes. The MTT and EdU assays showed that TAL1 overexpression reduced the proliferative capacity of the cancer cells by decreasing the number of cells in the DNA replication phase. Additionally, wound healing and Transwell assays revealed that TAL1 expression substantially decreased the migration and invasion abilities of the cells. Western blot analysis further indicated that TAL1 overexpression reduced the levels of the proliferation marker Ki67 and the mesenchymal marker N-cadherin, while restoring the epithelial marker E-cadherin, suggesting an inhibition of the epithelial-mesenchymal transition (EMT).

Mechanistic studies revealed that TAL1 acts as a transcriptional repressor of PKM2. Chromatin immunoprecipitation (ChIP) assays showed that TAL1 binds directly to the promoter region of the PKM2 gene. Dual-luciferase reporter assays confirmed that the presence of TAL1 decreases the transcriptional activity of the PKM2 promoter. When PKM2 was overexpressed in the presence of TAL1, the inhibitory effects of TAL1 on cell proliferation, migration, and glycolysis were reversed. This indicates that TAL1 suppresses NSCLC progression primarily by blocking the transcription of PKM2, a critical step in the glycolytic pathway.

The study then investigated the upstream regulation of TAL1, focusing on DNA methylation. Database analysis showed that the TAL1 promoter is hypermethylated in lung cancer tissues, and this methylation level was negatively correlated with TAL1 expression. The researchers identified DNMT3B as a key enzyme associated with this process. Pearson’s correlation analysis in LUAD and LUSC datasets revealed a positive correlation between DNMT3B expression and TAL1 promoter methylation. In cell line experiments, DNMT3B was found to be highly expressed in NSCLC cells relative to normal lung epithelial cells, supporting its potential role in the epigenetic silencing of TAL1.

Knockdown experiments using small hairpin RNAs (shRNAs) demonstrated that reducing DNMT3B expression led to a significant increase in TAL1 levels. Quantitative methylation-specific PCR (qMSP) showed that DNMT3B knockdown lowered the DNA methylation level at the TAL1 promoter. ChIP assays further confirmed that DNMT3B binds to the TAL1 promoter, and its depletion reduced this enrichment. These findings establish that DNMT3B directly mediates the methylation of the TAL1 promoter, thereby repressing its transcription in NSCLC cells.

The metabolic consequences of the DNMT3B-TAL1-PKM2 axis were assessed through cellular metabolism assays. Overexpression of DNMT3B in NSCLC cells increased lactate production, a hallmark of aerobic glycolysis. This effect was accompanied by upregulation of glycolysis-related proteins, including glucose transporter 1 (GLUT1), lactate dehydrogenase A (LDHA), and pyruvate dehydrogenase kinase 1 (PDK1). When TAL1 was co-overexpressed with DNMT3B, the increased lactate levels and glycolytic protein expression were reversed. This suggests that DNMT3B promotes glycolysis by silencing TAL1, which in turn removes the transcriptional brake on PKM2.

In vivo experiments using a xenograft mouse model were conducted to validate the findings in a physiological context. A549 cells were injected subcutaneously into BALB/c nude mice, and tumor growth was monitored over 15 days. Mice with overexpressed DNMT3B developed significantly larger and heavier tumors compared to the control group. Immunohistochemistry (IHC) of the tumor tissues showed that DNMT3B overexpression increased the levels of GLUT1, LDHA, PDK1, and Ki67, while decreasing the levels of cleaved caspase-3, an apoptosis marker. These results confirm that DNMT3B promotes tumor growth and suppresses apoptosis in vivo.

Further in vivo experiments tested the necessity of PKM2 in this process. When PKM2 was knocked down in the presence of DNMT3B overexpression, tumor weight and volume were significantly reduced compared to the DNMT3B-only group. This indicates that the pro-tumorigenic effects of DNMT3B are dependent on the activity of PKM2. The study also showed that overexpression of TAL1 inhibited tumor growth in vivo, and this inhibition was reversed by the co-overexpression of PKM2. These reciprocal experiments highlight the critical balance between the repressive and activating signals in the DNMT3B-TAL1-PKM2 pathway.

The authors note that while TAL1 is known as an oncogenic factor in T-cell acute lymphoblastic leukemia, its role in NSCLC appears to be tumor-suppressive. The study provides the first evidence that DNMT3B promotes glycolysis in NSCLC through the methylation of TAL1. By linking epigenetic regulation to metabolic reprogramming, the research offers a new perspective on the molecular drivers of lung cancer progression.

The findings suggest that targeting the DNMT3B-TAL1-PKM2 axis could be a potential therapeutic strategy for NSCLC. Inhibition of DNMT3B or restoration of TAL1 expression might help to suppress glycolysis and reduce tumor growth. However, the authors emphasize that clinical trials are necessary to validate these findings in human patients.

The work was published in the journal Cell Division, Volume 20, article number 25, in 2025. The authors utilized a combination of bioinformatics and experimental validation to dissect the molecular interactions between DNMT3B, TAL1, and PKM2 in the context of non-small cell lung cancer.

In the bioinformatics phase, the researchers integrated data from three specific Gene Expression Omnibus datasets: GSE18842, GSE19188, and GSE159857. They screened for differentially expressed genes using an adjusted p-value threshold of less than 0.05 and a Log2 FoldChange magnitude of at least 1. This analysis identified 14 upregulated and 37 downregulated transcription factors. Using the String database and Cytoscape software, the team applied the BottleNeck algorithm to identify hub genes, selecting FOS, TAL1, and FOXM1 as the top three scoring candidates. TAL1 was specifically noted for its under-expression in the NSCLC datasets, with Log2 FoldChange values of -1.981 in GSE18842, -1.941 in GSE19188, and -1.361 in GSE159857. This computational approach provided the initial rationale for investigating TAL1’s functional role in tumor progression.

The experimental validation involved several NSCLC cell lines, including H1299, HCC827, A549, H460, and PC-9, alongside the normal human lung epithelial cell line BEAS-2B. The researchers employed lentiviral vectors to manipulate gene expression, including constructs for overexpressing TAL1, PKM2, and DNMT3B, as well as constructs for knocking down DNMT3B and PKM2. These vectors were constructed by VectorBuilder and had titers exceeding 10^9 TU/mL. The cells were infected with these lentiviruses and subsequently screened using puromycin to ensure stable expression or knockdown of the target genes.

Statistical analysis was performed using Prism 10.4.2 software, with data presented as mean ± standard error of the mean. The researchers used unpaired t-tests, one-way, or two-way ANOVA for comparisons, with the Tukey test for post hoc analyses. Cellular experiments were repeated three times to ensure reproducibility, while animal experiments involved five mice in each group. The significance level was set at p < 0.05. The animal experiments were approved by the Ethics Committee of Jiangxi Cancer Hospital, ensuring compliance with ethical standards for research involving laboratory animals. The mice used were four-week-old BALB/c nude mice, which were housed under pathogen-free conditions.

The study highlights the importance of understanding the metabolic reprogramming in cancer cells, particularly the shift towards aerobic glycolysis. This metabolic shift facilitates tumor survival and contributes to the acidification of the tumor microenvironment, which can lead to drug resistance. By targeting glycolytic metabolism, the researchers suggest a potential therapeutic strategy for NSCLC.

Subject of Research: DNMT3B-mediated methylation of TAL1 and its role in PKM2-driven glycolysis in non-small cell lung cancer

Article Title: DNMT3B blocks TAL1-mediated PKM2 transcriptional repression to promote non-small cell lung cancer progression through inducing glycolysis

Article References: Guo, Y., Xu, B., Mao, W., Zhao, Z., Li, Z., Qiu, Z., & Guo, C. (2025). DNMT3B blocks TAL1-mediated PKM2 transcriptional repression to promote non-small cell lung cancer progression through inducing glycolysis. Cell Division, 20(1), Article 25. https://doi.org/10.1186/s13008-025-00168-8

Image Credits: AI Generated

DOI: 10.1186/s13008-025-00168-8

Keywords: Non-small cell lung cancer, DNMT3B, TAL1, PKM2, Glycolysis, DNA methylation, Transcriptional repression, Metabolic reprogramming, Cell Division, Epigenetics, blocks, TAL1-mediated