Cancer-Associated Fibroblasts Hinder Lung Squamous Cell Carcinoma Oxidative Stress-Induced Apoptosis via METTL3 Mediated m(6)A Methylation of COL10A1.

Cancer-associated fibroblasts (CAFs) within the tumor microenvironment are key players in tumorigenesis and tumor development. Nevertheless, the regulatory mechanisms of CAFs on lung squamous cell carcinoma- (LUSC-) associated remain poorly elucidated. The microarray dataset GSE22874, containing 30 specimens of primary culture of normal fibroblasts (NFs) and 8 specimens of cancer-associated fibroblasts (CAFs) samples derived from LUSC, was retrieved from the Gene Expression Omnibus (GEO) database and then calculated by using the R language (limma package) to identify differentially expressed genes (DEGs). CAF-conditioned medium (CAF-CM) was collected and used to culture LUSC cells, followed by assessment of cell proliferation, apoptosis, and oxidative stress levels by using CCK-8, annexin V-FITC/PI double staining and ELISA assays. Subsequently, COL10A1 was knocked down in CAFs to assess the role of COL10A1 in CAF regulation of LUSC behavior. Bioinformatics online analysis and MeRIP were applied to predict and test the m6A modification of COL10A1 mRNA and the regulatory relationship with METTL3. Rescue experiments were next performed to explore the effects of METTL3 and COL10A1 in CAFs on LUSC cell proliferation, apoptosis, and oxidative stress. LUSC tumor cells with or without (COL10A1-silenced) CAFs were subcutaneously inoculated in nude mice to evaluate the effect of COL10A1 in CAFs on LUSC tumor growth. Elevated expression of COL10A1 was found in LUSC-derived CAFs by GSE22874 dataset analysis. We discovered that COL10A1 and METTL3 was expressed in both LUSC cells and matched CAFs, while COL10A1 expression was prominently higher in CAFs than in LUSC cells. CAF-CM memorably encouraged LUSC cell proliferation and suppressed apoptosis-induced oxidative stress, which was reversed by interfering with COL10A1 expression in CAFs, suggesting that COL10A1 might be secreted by CAFs into the culture medium to exert its effects inside LUSC cells. Global m6A modification was decreased in METTL3 knocked down CAFs. M6A modification, expression levels, and stability of COL10A1 mRNA were impaired upon METTL3 knockdown in CAFs. Overexpression of COL10A1 in CAFs partially reversed the effect of METTL3 knockdown on the malignant behavior of LUSC cells. In vivo studies confirmed that CAFs accelerated LUSC tumor growth, and this effect was counteracted by COL10A1 silencing. COL10A1 secreted by CAFs could facilitate LUSC cell proliferation and repress apoptosis-induced oxidative stress, and the mechanism was due to elevated expression mediated by METTL3 promoting its mRNA m6A modification, thereby accelerating tumor growth.

Li Y ，Li X ，Deng M ，Ye C ，Peng Y ，Lu Y ... -  《-》

Metformin attenuates multiple myeloma cell proliferation and encourages apoptosis by suppressing METTL3-mediated m6A methylation of THRAP3, RBM25, and USP4.

Chen CJ ，Huang JY ，Huang JQ ，Deng JY ，Shangguan XH ，Chen AZ ，Chen LT ，Wu WH ... -  《-》

Cancer-associated fibroblasts-derived exosomal METTL3 promotes the proliferation, invasion, stemness and glutaminolysis in non-small cell lung cancer cells by eliciting SLC7A5 m6A modification.

Cancer-associated fibroblasts (CAFs) can promote the crosstalk between cancer cells and tumor microenvironment by exosomes. METTL3-mediated N6-methyladenine (m6A) modification has been proved to promote the progression of non-small cell lung cancer (NSCLC). Here, we focused on the impacts of CAFs-derived exosomes and METTL3-mediated m6A modification on NSCLC progression. Functional analyses were conducted using Cell Counting Kit-8, EdU, colony formation, sphere formation and transwell assays, respectively. Glutamine metabolism was evaluated by detecting glutamate consumption, and the production of intercellular glutamate and α-ketoglutarate (α-KG). qRT-PCR and western blotting analyses were utilized to measure the levels of genes and proteins. Exosomes were isolated by kits. The methylated RNA immunoprecipitation assay detected the m6A modification profile of Amino acid transporter LAT1 (SLC7A5) mRNA. The NSCLC mouse model was established to conduct in vivo experiments. We found that CAFs promoted the proliferation, invasion, stemness and glutaminolysis in NSCLC cells. METTL3 was enriched in CAFs and was packaged into exosomes. After knockdown of METTL3 in CAF exosomes, it was found the oncogenic effects of CAFs on NSCLC cells were suppressed. CAFs elevated m6A levels in NSCLC cells. Mechanistically, exosomal METTL3-induced m6A modification in SLC7A5 mRNA and stabilized its expression in NSCLC cells. Moreover, SLC7A5 overexpression abolished the inhibitory effects of exosomal METTL3-decreased CAFs on NSCLC cells. In addition, METTL3 inhibition in CAF exosomes impeded NSCLC growth in vivo. In all, CAFs-derived exosomal METTL3 promoted the proliferation, invasion, stemness and glutaminolysis in NSCLC cells by inducing SLC7A5 m6A modification.

Fan Y ，Yu Y 《Human Cell》

Cancer-associated fibroblasts promote migration and invasion of non-small cell lung cancer cells via METTL3-mediated RAC3 m(6)A modification.

Cancer progression depends on the communication between tumor cells and tumor microenvironment. Cancer-associated fibroblasts (CAFs) are a major component of stromal cells. CAFs promote cancer metastasis; however, it has not been evaluated whether N6-methyladenosine (m6A) modification is responsible for CAFs' role in metastasis. In the present study, we found that CAFs promoted migration and invasion of non-small cell lung cancer (NSCLC) cells by elevating m6A modification in NSCLC cells. Methyltransferase-like 3 (METTL3) in NSCLC cells mediated CAFs' effect on m6A modification, and was regulated by CAFs-secreted vascular endothelial growth factor A (VEGFA). METTL3 knockdown in NSCLC cells dramatically inhibited cell migration and invasion, and suppressed tumor growth in vivo. Database analysis revealed that METTL3 was associated with poor prognosis of lung cancer. The mechanism study showed that METTL3 increased m6A level of RAC3 mRNA, resulting in increased stability and translation of RAC3 mRNA. RAC3 was responsible for the CAFs' promoting effect on cell migration via the AKT/NF-κB pathway. This study established a CAF-METTL3-RAC3 m6A modification-dependent regulation system in NSCLC metastasis, suggesting potential candidates for metastasis treatment.

Chen M ，Zhang Q ，Zheng S ，Guo X ，Cao L ，Ren Y ，Qian Y ，Wang M ，Wu X ，Xu K ... -  《International Journal of Biological Sciences》

METTL3 in cancer-associated fibroblasts-derived exosomes promotes the proliferation and metastasis and suppresses ferroptosis in colorectal cancer by eliciting ACSL3 m6A modification.

Cancer-associated fibroblasts (CAFs) have been reported that can affect cancer cell proliferation, metastasis, ferroptosis, and immune escape. METTL3-mediated N6-methyladenine (m6A) modification is involved in the tumorigenesis of colorectal cancer (CRC). Herein, we investigated whether METTL3-dependent m6A in CAFs-derived exosomes (exo) affected CRC progression. qRT-PCR and western blotting analyses detected levels of mRNAs and proteins. Cell proliferation and metastasis were evaluated using MTT, colony formation, transwell, and wound healing assays, respectively. Cell ferroptosis was assessed by detecting cell viability and the levels of Fe+, reactive oxygen species, and glutathione after erastin treatment. Exosomes were isolated from CAFs by ultracentrifugation. The m6A modification profile was determined by methylated RNA immunoprecipitation assay and the interaction between METTL3 and ACSL3 (acyl-CoA synthetase 3) was verified using dual-luciferase reporter assay. Animal models were established for in vivo analysis. CAFs promoted CRC cell proliferation and metastasis, and suppressed cell ferroptosis. METTL3 was enriched in CAFs and was packaged into exosomes. The m6A modification and METTL3 expression were increased in CRC samples. Knockdown of METTL3 in CAFs-exo suppressed CRC cell proliferation and metastasis, and induced cell ferroptosis. Mechanistically, METTL3 induced ACSL3 m6A modification and stabilized its expression. The anticancer effects mediated by METTL3-silenced CAFs-exo could be rescued by ACSL3 overexpression. Moreover, in vivo assay also showed that CAFs-exo with decreased METTL3 could hinder CRC growth and metastasis in mice models. CAFs promoted the proliferation and metastasis, and restrained the ferroptosis in CRC by exosomal METTL3-elicited ACSL3 m6A modification.

Ren H ，Wang M ，Ma X ，An L ，Guo Y ，Ma H ... -  《Biology Direct》