Targeting YTHDC1 Inhibited the Progression of Mantle Cell Lymphoma By Inhibiting the Expression of MYC Gene through m6A Modification

Objective

YTHDC1 (YT521-B homology domain-containing protein 1) is an in vivo nuclear RNA n⁶ -methyladenosine (m6A) reader with YTH domain. At present, YTHDC1 has been found to play an important role in the pathogenesis of various tumors, such as acute myeloid leukemia. However, there are no relevant studies on YTHDC1 in mantle cell lymphoma (MCL). Therefore, this study was to investigate the role and mechanisms of YTHDC1 gene in MCL.

Methods

The expression of YTHDC1 in MCL patients and normal human cells was compared in GEO database. JEKO1 cells with stable knockdown of YTHDC1 were constructed. MTT method, flow cytometry, western blot were used to investigate the effect of YTHDC1 knockdown on proliferation, apoptosis, cell cycle and proteins of apoptosis and G2M phase of cell cycle. MCL subcutaneous tumor model was used to explore the effect of knocking down YTHDC1 in vivo; HE staining and Ki-67 immunohistochemical staining were performed to value the tumor load. RNA-Seq analysis was performed to explore the possible mechanisms of YTHDC1 genes in MCL. MERIP-qPCR and YTHDC1-RIP-qPCR were used to investigate the mRNA level of the target gene of YTHDC1 modified by m6A in MCL. Additionally, qPCR was used to evaluate the mRNA stability of target genes, based on the principle of actinomycin D blocking mRNA synthesis.

Results

The expression of YTHDC1 in MCL was significantly increased compared to normal people. Knocking down YTHDC1 inhibited the proliferation of MCL cells, and promoted cell apoptosis, and leaded the cell cycle arrest at G2M stage, and decreased the expression of apoptosis-related proteins PARP, CASPASE 3 and G2M related proteins CDC25C and CYCLINB 1, while increased the expression of CLEVEAD CASPASE 3. In addition, the tumor growth was decreased and the tumor load was reduced after YTHDC1 was knocked down. There were 3333 differentially expressed genes in sh-YTHDC1 and control cells, including 2178 down-regulated genes and 1155 upregulated genes. The top down-regulated genes included MYC, etc. Enrichment analysis showed that YTHDC1 down-regulated gene may be located in cell membrane, down-regulated cancer pathway, ECM-receptor interaction, etc. Intersection analysis of RNA-Seq data, YTHDC1- iCLIP data and YTHDC1-miCLIP data revealed 174 differentially expressed genes with m6A modification that bind to YTHDC1 in MCL, including MYC. Furthermore, we demonstrated that YTHDC1 regulated the mRNA level of MYC target gene through m6A modification by regulating the stability of MYC mRNA.

Conclusion

The expression of YTHDC1 gene was higher in MCL patients than that in normal people and played a important role in the progression of MCL. Mechanically, YTHDC1 regulated the mRNA level of MYC target gene through m6A modification by regulating the stability of MYC mRNA.

No relevant conflicts of interest to declare.