Post-transcriptional modification has become an important regulator in many biological processes and has attracted increasing attention. N6-methyladenosine (m6A) is one of the common internal modifications in eukaryotic mRNA[1]. One of the methods widely used to study m6A modification at the transcriptional level is methylated RNA immunoprecipitation sequencing (MeRIP-seq), which involves the immunoprecipitation of m6A modified RNA fragment and peak detection by comparing to background gene coverage. In 2012, Meyer et al. [2] determined 7,676 human genes containing m6A in somatic cells using the MeRIP-seq method; they found that m6A was primarily localized to the conserved RRACH sequence (R = G or A;H = A, C or U), the sequence is enriched in the 3' untranslated regions(3′UTRs), and the stop codon region of the protein-coding mRNA. In mammals the amount of m6A modified adenine is between 0.1% and 0.4% [3, 4], so each mRNA has only 3–5 m6A methylation sites and regulates RNA stability, localization, splicing, transport, and translation at the post-transcriptional level[5].
m6A modification is mainly associated with three types of regulators. The first is m6A methyltransferases, which encode genes called “writers” and include METTL3, MetTL14, MetTL16, RBM15/15B, KIAA1429, ZC3H13 and WATP (Wilms Tumor 1 associated protein)[6]. Together, they form a complex that induces the writing of the m6A methylated group into RNA [7]. The second type of regulators is m6A demethylase, which encodes genes called “erasers” and includes FTO and ALKBH5 [8]. This protein can remove the m6A methylation group in RNA, thus affecting the biological process of tumor. The last group of regulators can bind to the m6A methylation site and can read information to play a role; the encoding genes of this group are called “readers”, some of which identified so far are YTHDF1, YTHDF2, YTHDF3,YTHDC1, YTHDC2, HNRNPC, HNRNPG, HNRNPA2B1 and IGF2BPs[9]. Readers are proteins that bind to the m6A sites on RNA in the nucleus or cytoplasm to perform a specific biological function. At present, the research on the molecular biological mechanism of m6A modification of tumor is still in the initial stage, and further exploration is needed to provide targets for tumor therapy.
Breakthroughs in the discovery and research of m6A writers, erasers and readers, as well as the development of high-throughput sequencing data analysis, have helped illuminate the biological functions and potential mechanisms of m6A. A growing number of studies have shown that m6A RNA methylation plays an important role in tumor genesis and development. The effect of m6A modification on tumors is mainly reflected in the regulation of these tumor-related genes. Abnormal m6A modifications are closely related to tumor development [10, 11]. Therefore, it is particularly important to further study the regulatory mechanism of m6A modification in the process of tumor development. Glioblastoma is the most deadly primary brain tumor. Studies have shown that m6A modification is associated with the growth and self-renewal of glioblastoma stem cells (GSCs)[12]. Knockdown of METTL3 or METTL14, key components of the RNA methyltransferase complex, promotes GSCs growth, self-renewal, and tumorigenesis, whereas upregulation of METTL3 or inhibition of FTO has the reverse effects[12]. Another study also reported the important role of METTL3-mediated m6A modification in GSC maintenance and in glial cell dedifferentiation[13]. This study found that METTL3 expression is upregulated in GSCs and is attenuated during differentiation. The role of METTL3 has also been reported in acute myeloid leukemia(AML). Studies have shown that METTL3 is expressed more abundantly in AML cells than in normal hematopoietic cells. And METTL3-mediated elevation of m6A in AML plays an important role in maintaining pluripotency and in inhibiting cell differentiation[14, 15]. In other tumors, such as gastric cancer, upregulation of METTL3 promotes the stability of ZMYM1, thereby enhancing EMT process in vitro and metastasis in vivo[16].
In the present study we obtained high-throughput sequencing data of MeRIP-seq for invasive malignant pleomorphic adenoma (IMPA), glioblastoma (GBM), acute myeloid leukemia(AML), and gastric cancer (GC) from GEO database, and we analyzed the level of m6A modification in different tumor samples. Combined with the corresponding RNA sequencing (RNA-seq) data of these tumor samples, we analyzed the differences of gene expression at the transcription level under different m6A modification levels. In addition, combined with the tumor clinical data from the The Cancer Genome Atlas (TCGA) RNA-seq database, we expected to identify the key genes that affect the survival of different tumor patients under the regulation of m6A modification, so as to provide guiding strategies for the clinical treatment of tumor patients. Based on this data analysis, we found that the enrichment degree of m6A modification was enhanced in IMPA and GBM but was decreased in METLL3 or YTHDF1 knockdown tumor cell lines, suggesting that m6A modification plays an important role in tumorigenesis. Among the genes with different levels of m6A modification that were common to different tumor types, the differential expression of DUSP7 is significantly correlated with the survival of patients with AML. DUSP7 may be involved in tumor genesis and development through the MAPK signaling pathway, and this gene has rarely been reported in AML, thus providing a new idea for subsequent clinical studies on AML.