Methyltransferase like 3 promotes colorectal cancer proliferation by stabilizing CCNE1 mRNA in an m6A-dependent manner.
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ABSTRACT:
SUBMITTER: Zhu W
PROVIDER: S-EPMC7131945 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Altered expression levels of the proteins that regulate N6-methyladenosine (m6A) RNA methylation, including methyltransferase-like 14 (METTL14), are associated with cancer development. Based on our analysis of m6A methylation regulators using The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets, we focused on the regulatory role of METTL14 in ovarian cancer. We performed bioinformatics and survival analyses with these datasets and also used METTL14-overexpressing SKOV-3 ovarian cancer cells for in vitro studies. Trophinin associated protein (TROAP) siRNA and treatment with or without actinomycin D was used in the cells for qRT-PCR, western blot, cDNA microarray, cell viability, colony formation, luciferase gene reporter, methylated RNA immunoprecipitation (MeRIP)-qPCR, total RNA methylation, and RNA stability assays. Additionally, ovarian cancer and normal tissue samples were analyzed by immunohistochemistry, qRT-PCR, and western blot assays. The TCGA and GEO data confirmed copy number variations (CNVs) of these m6A RNA methylation regulators in ovarian cancer tissues. Furthermore, reduced METTL14 expression was associated with alterations in CNVs as well as poor patient survival in ovarian cancer. Moreover, the METTL14 and m6A RNA methylation levels were both significantly reduced in ovarian cancer tissues than in normal tissues. Restoration of METTL14 expression suppresses ovarian cancer cell proliferation by inhibition of TROAP expression. Further in vivo and in vitro experiments confirmed that METTL14 is a negative regulator of ovarian cancer cell proliferation via TROAP expression and that m6A RNA methylation regulates TROAP mRNA stability. In conclusion, METTL14 overexpression decreased ovarian cancer proliferation by inhibition of TROAP expression via an m6A RNA methylation-dependent mechanism.
| S-EPMC8894193 | biostudies-literature
Project description:Rationale: N6-methyladenosine (m6A) mRNA methylation is the most abundant chemical posttranscriptional modification in mRNA and is involved in the regulation of a number of biological processes. Insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) has recently been reported as having the capacity to recognize m6A sites in mRNA and plays a role in regulating mRNA metabolization. However, it is unclear which genes IGF2BP1 targets to identify m6A sites and what are their respective functions in endometrial cancer (EC). Methods: Quantitative PCR, western blot and immunohistochemistry were used to measure IGF2BP1 expression in EC cell lines and tissues. Xenograft experiments were performed to examine the in vivo role of IGF2BP1 in EC cell growth. RNA-binding protein immunoprecipitation sequencing, methylated RNA-binding protein immunoprecipitation sequencing and RNA-sequencing were also conducted to identify potential IGF2BP1 targets involved in EC regulation. Co-immunoprecipitation and mass spectrometry were used to identify IGF2BP1-interacting proteins. Results: IGF2BP1 expression increased in EC, and high expression of this protein correlated with poor prognosis. IGF2BP1 overexpression/knockdown can promote (and inhibit) cell proliferation and regulate the tumor cell cycle and cancer progression, both in vivo and in vitro. Mechanistically, IGF2BP1 can recognize m6A sites in the 3' untranslated region (3'UTR) of Paternally Expressed Gene 10 (PEG10) mRNA and recruits polyadenylate-binding protein 1 (PABPC1) to enhance PEG10 mRNA stability, which consequently promotes PEG10 protein expression. Additionally, it would appear that a large number of PEG10 proteins bind p16 and p18 gene promoter sequences, thereby repressing expression and accelerating the cell cycle. Conclusion: This investigation found that IGF2BP1 has a crucial role in the m6A-dependent regulatory mechanism for endometrial cancer. This study provides new insights into our understanding of disease progression and provides another potential route for understanding biological functions.
| S-EPMC7738899 | biostudies-literature
Project description:Glioma is one of the leading causes of tumor-related deaths worldwide, but its potential mechanism remains unclear. This study aimed to explore the biological role and potential mechanism of argininosuccinate synthase 1 (ASS1) in glioma. The relative expression levels of ASS1 in glioma specimens and cell lines were calculated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blotting. The biological functions of ASS1 were demonstrated using the 5-ethynyl-2'-deoxyuridine (EdU) assay, transwell assay, and in vivo experiments. In addition, methylated RNA immunoprecipitation (MeRIP), RNA immunoprecipitation (RIP), and luciferase reporter assays were performed to explore the molecular mechanism of ASS1 in glioma. ASS1 expression levels were found to be downregulated in glioma specimens and cell lines. Functionally, we confirmed that ASS1 inhibited glioma cell proliferation, migration, invasion, and growth both. Furthermore, we found that ASS1 was a target of N(6)-adenosine-methyltransferase-14 (METTL14)-mediated N6-methyladenosine (m6A) modification. Overexpression of METTL14 markedly elevated ASS1 mRNA m6A modification and suppressed ASS1 mRNA expression. We also revealed that METTL14-mediated ASS1 mRNA degradation relied on the YTH m6A RNA-binding protein 2 (YTHDF2)-dependent pathway. We confirmed that decreased ASS1 expression promoted the cell proliferation, migration, and invasion in glioma, and that the METTL14/ASS1/YTHDF2 regulatory axis may be an effective therapeutic target for glioma.
| S-EPMC8805915 | biostudies-literature
Project description:Background:The modification and regulation of N6-methyladenosine (m6A) at mRNA level can affect the development and progression in various tumors. ALKBH5, as an m6A demethylase, plays different roles in tumors by regulating the m6A modification of mRNA. However, its role in renal cell carcinoma (RCC) remains unclear. Methods:First, levels of ALKBH5 in RCC tissues and cell lines were verified by qRT-PCR and western blot. We analyzed the relationship between ALKBH5 and the clinicopathological characteristics of RCC patients and the influence of ALKBH5 on the prognosis of patients. Then we generated ALBKH5-overexpression, ALBKH5-knockdown stable RCC cell lines and their control cell lines. Through cell proliferation assay, colony formation assay, cell invasion and tumor migration assay, cell cycle assay and xenograft studies, we studied the ALKBH5 roles in RCC cell lines. AURKB was predicted to be its potential target based on TCGA database analysis and verified by western blot. The role of AURKB in RCC was verified by TCGA database and Kaplan-Meier analysis with TMA immunohistochemical analysis. Finally, the specific molecular mechanism of ALKBH5 targeting AURKB was explored by dual-luciferase reporter assay, RNA immunoprecipitation (RIP), m6A dot-blot assay, m6A RNA Immunoprecipitation (MeRIP) assay, and mRNA stability assay. Results:We found that ALKBH5 was highly expressed in both RCC tumor tissues and cell lines. Clinicopathological analysis showed that high ALKBH5 expression was associated with larger tumor volume (P=0.017) and higher TNM staging (P=0.006), and worse prognosis (log rank: P=0.0199). The cellular functional assays showed that stably overexpression ALKBH5 could promote the cell proliferation, colony formation, cell migration and cell invasion of renal cell carcinoma cells in vitro and promote tumor growth in vivo. In contrast, ALKBH5 knocked down inhibited cell proliferation, colony formation, migration and invasion of renal cell carcinoma cells in vitro. Based on TCGA database analysis, AURKB was predicted highly expressed in RCC and a potential target of ALKBH5. Both database prediction and TMA immunohistochemical analysis supported that AURKB could affect the prognosis of RCC patients (P values of 5.5e-08 and 0.0004, respectively) and was regulated by ALKBH5 expression level. Subsequent mechanism experiments showed that ALKBH5 regulated the expression of AURKB by regulating the stability of AURKB mRNA in the m6A-dependent manner, and finally promoted cell proliferation. Furthermore, we found that hypoxia-induced HIF could up-regulate both expressions of AURKB and ALKBH5. Conclusions:Our findings suggest that ALKBH5 may play a carcinogenic role in renal cell carcinoma by stabilizing AURKB mRNA in a m6A-dependent manner. These data suggest that ALKBH5 may play a key role in RCC and targeting the ALKBH5 signaling pathway may be a promising strategy for the treatment of RCC.
| S-EPMC7290639 | biostudies-literature
Project description:BackgroundLymph node metastasis is the main cause of poor prognosis of head and neck squamous carcinoma (HNSCC) patients. N6-methyladenosine (m6A) RNA modification is an emerging epigenetic regulatory mechanism for gene expression, and as a novel m6A reader protein, IGF2BP2 has been implicated in tumor progression and metastasis. However, not much is currently known about the functional roles of IGF2BP2 in HNSCC, and whether IGF2BP2 regulates lymphatic metastasis through m6A modification in HNSCC remains to be determined.MethodsThe expression and overall survival (OS) probability of m6A-related regulators in HNSCC were analyzed with The Cancer Genome Atlas (TCGA) dataset and GEPIA website tool, respectively. The expression levels of IGF2BP2 were measured in HNSCC tissues and normal adjacent tissues. To study the effects of IGF2BP2 on HNSCC cell metastasis in vitro and in vivo, gain- and loss- of function methods were employed. RIP, MeRIP, luciferase reporter and mRNA stability assays were performed to explore the epigenetic mechanism of IGF2BP2 in HNSCC.ResultsWe investigated 20 m6A-related regulators in HNSCC and discovered that only the overexpression of IGF2BP2 was associated with a poor OS probability and an independent prognostic factor for HNSCC patients. Additionally, we demonstrated that IGF2BP2 was overexpressed in HNSCC tissues, and significantly correlated to lymphatic metastasis and poor prognosis. Functional studies have shown that IGF2BP2 promotes both HNSCC cell migration as well as invasion via the epithelial-mesenchymal transition (EMT) process in vitro, and IGF2BP2 knockdown significantly inhibited lymphatic metastasis and lymphangiogenesis in vivo. Mechanistic investigations revealed that Slug, a key EMT-related transcriptional factor, is the direct target of IGF2BP2, and essential for IGF2BP2-regulated EMT and metastasis in HNSCC. Furthermore, we demonstrated that IGF2BP2 recognizes and binds the m6A site in the coding sequence (CDS) region of Slug and promotes its mRNA stability.ConclusionsCollectively, our study uncovers the oncogenic role and potential mechanism of IGF2BP2, which serves as a m6A reader, in controlling lymphatic metastasis and EMT in HNSCC, suggesting that IGF2BP2 may act as a therapeutic target and prognostic biomarker for HNSCC patients with metastasis.
| S-EPMC8722037 | biostudies-literature
Project description:Insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3), an RNA-binding protein, is associated with tumorigenesis and progression. However, the exact molecular mechanisms of IGF2BP3 in colorectal cancer (CRC) oncogenesis, progression, and drug resistance remain unclear. This study found that IGF2BP3 was upregulated in CRC tissues. Clinically, the elevated IGF2BP3 level is predictive of a poor prognosis. Functionally, IGF2BP3 enhances CRC tumorigenesis and progression both in vitro and in vivo. Mechanistically, IGF2BP3 promotes epidermal growth factor receptor (EGFR) mRNA stability and translation and further activates the EGFR pathway by serving as a reader in an N6-methyladenosine (m6A)-dependent manner by cooperating with METTL14. Furthermore, IGF2BP3 increases the drug resistance of CRC cells to the EGFR-targeted antibody cetuximab. Taken together, our results demonstrated that IGF2BP3 was a functional and clinical oncogene of CRC. Targeting IGF2BP3 and m6A modification may therefore offer rational therapeutic targets for patients with CRC.
| S-EPMC10474290 | biostudies-literature
Project description:Growing evidence indicates N6-methyladenosine (m6A) has biological function in oncogenesis. METTL3, the catalytic component, is the most important part of methyltransferase complex and plays a crucial role in cancers. However, the biological function of circRNAs derived from METTL3 in breast cancer and the underlying molecular mechanism remains unclear. Herein, we report circMETTL3, which has not been explored in breast cancer, and it is markedly upregulated in breast cancer. Moreover, we uncovered that circMETTL3 could facilitate cell proliferation, migration and invasion in breast cancer. Mechanism investigation showed that circMETTL3 might act as a competing endogenous RNA (ceRNA) of miR-31-5p and upregulate its target cyclin-dependent kinases (CDK1). Moreover, m6A modification of circMETTL3 might affect its expression. Taken together, our results elucidate that circMETTL3 promotes breast cancer progression through circMETTL3/miR-31-5p/CDK1 axis. Moreover, METTL3, the host gene of circMETTL3, may regulate circMETTL3 expression in an m6A-dependent manner, while circMETTL3 has no effect on METTL3 expression, providing a new relationship between the circRNA and the corresponding host gene. Thus, it may serve as a new therapeutic target for breast cancer.
| S-EPMC8040468 | biostudies-literature
Project description:The aberrant expression of fat mass and obesity-associated protein (FTO) has been confirmed to be associated with a variety of cancers and participates in the regulation of multiple biological behaviours. FTO plays an oncogenic role in bladder cancer, but few studies have focused on how FTO promotes bladder cancer progression by regulating miRNA synthesis. Here, we confirmed that FTO expression was significantly increased in bladder cancer and was associated with a poor prognosis. FTO overexpression promoted bladder cancer cell proliferation, whereas FTO knockdown inhibited bladder cancer cell proliferation. We also demonstrated that FTO promoted bladder cancer cell proliferation via the FTO/miR-576/CDK6 pathways. Taken together, our work revealed that FTO plays a critical role in bladder cancer and could be a potential diagnostic or prognostic biomarker for this disease.
| S-EPMC8560827 | biostudies-literature
Project description:The oncofetal mRNA-binding protein IGF2BP1 and the transcriptional regulator SRF modulate gene expression in cancer. In cancer cells, we demonstrate that IGF2BP1 promotes the expression of SRF in a conserved and N6-methyladenosine (m6A)-dependent manner by impairing the miRNA-directed decay of the SRF mRNA. This results in enhanced SRF-dependent transcriptional activity and promotes tumor cell growth and invasion. At the post-transcriptional level, IGF2BP1 sustains the expression of various SRF-target genes. The majority of these SRF/IGF2BP1-enhanced genes, including PDLIM7 and FOXK1, show conserved upregulation with SRF and IGF2BP1 synthesis in cancer. PDLIM7 and FOXK1 promote tumor cell growth and were reported to enhance cell invasion. Consistently, 35 SRF/IGF2BP1-dependent genes showing conserved association with SRF and IGF2BP1 expression indicate a poor overall survival probability in ovarian, liver and lung cancer. In conclusion, these findings identify the SRF/IGF2BP1-, miRNome- and m6A-dependent control of gene expression as a conserved oncogenic driver network in cancer.
| S-EPMC6326824 | biostudies-literature
Project description:BackgroundIncreasing studies have shown that long noncoding RNAs (lncRNAs) are pivotal regulators participating in carcinogenic progression and tumor metastasis in colorectal cancer (CRC). Although lncRNA long intergenic noncoding RNA 460 (LINC00460) has been reported in CRC, the role and molecular mechanism of LINC00460 in CRC progression still requires exploration.MethodsThe expression levels of LINC00460 were analyzed by using a tissue microarray containing 498 CRC tissues and their corresponding non-tumor adjacent tissues. The correlations between the LINC00460 expression level and clinicopathological features were evaluated. The functional characterization of the role and molecular mechanism of LINC00460 in CRC was investigated through a series of in vitro and in vivo experiments.ResultsLINC00460 expression was increased in human CRC, and high LINC00460 expression was correlated with poor five-year overall survival and disease-free survival. LINC00460 overexpression sufficiently induced the epithelial-mesenchymal transition and promoted tumor cell proliferation, migration, and invasion in vitro and tumor growth and metastasis in vivo. In addition, LINC00460 enhanced the protein expression of high-mobility group AT-hook 1 (HMGA1) by directly interacting with IGF2BP2 and DHX9 to bind the 3' untranslated region (UTR) of HMGA1 mRNA and increased the stability of HMGA1 mRNA. In addition, the N6-methyladenosine (m6A) modification of HMGA1 mRNA by METTL3 enhanced HMGA1 expression in CRC. Finally, it suggested that HMGA1 was essential for LINC00460-induced cell proliferation, migration, and invasion.ConclusionsLINC00460 may be a novel oncogene of CRC through interacting with IGF2BP2 and DHX9 and bind to the m6A modified HMGA1 mRNA to enhance the HMGA1 mRNA stability. LINC00460 can serve as a promising predictive biomarker for the diagnosis and prognosis among patients with CRC.
| S-EPMC7851923 | biostudies-literature