m6 A-mediated regulation of PBX1-GCH1 axis promotes gastric cancer proliferation and metastasis by elevating tetrahydrobiopterin levels

Background: Methyltransferase 3 (METTL3)-mediated N6-methyladenosine (m⁶ A) RNA modification has been demonstrated to be a potential factor in promoting gastric Cancer (GC). METTL3 regulates a series of signaling pathways by modifying various mRNAs. This study aimed to identify novel METTL3-mediated signaling pathways and explored possible targets for use in the clinical setting of gastric Cancer.

Methods: To investigate the proliferation and metastatic capacity of GC cell lines with METTL3 knockdown, a xenograft, lung metastasis, and popliteal lymph node metastasis model was used. The m⁶ A-modified RNA immunoprecipitation (Me-RIP) sequence was utilized to explore the target mRNAs of METTL3. Cell counting kit 8 and transwell assays were performed to investigate the promoting function of pre-B cell leukemia homeobox 1 (PBX1) and GTP cyclohydrolase 1 (GCH1). Western blotting and chromatin immunoprecipitation were employed to confirm the involvement of the METTL3-PBX1-GCH1 axis. ELISA and liquid chromatography-mass spectrometry were used to explore the biological function of tetrahydrobiopterin (BH₄ ).

Results: Knockdown of METTL3 suppressed xenograft tumor growth and lung/lymph node metastasis in vivo. Mechanistically, we found that METTL3 combined with and stabilized PBX1 mRNAs. Chromatin immunoprecipitation (ChIP) and further experiments suggested that PBX1 acted as a transcription factor inducing GCH1 expression. Moreover, the METTL3-PBX1-GCH1 axis increased BH₄ levels in GC cells, thereby promoting tumor progression.

Conclusions: This study suggested that METTL3 enzymes promote tumor growth and lung/lymph node metastasis via METTL3-PBX1-GCH1 axis increasing BH4 levels in GC.