Methionine represses the autophagy of gastric cancer stem cells via promoting the methylation and phosphorylation of RAB37

This study focused on the role of methionine (MET) in the autophagy of gastric cancer stem cells (GCSCs) and aims to elaborate its regulatory mechanism. In the present study, the GCSCs were isolated from human gastric cancer cell lines using an anti-CD44 antibody, and then cultured in MET⁺ homocysteine (HCY)⁻ or MET⁻HCY⁺ medium. In MET⁺HCY^–treated GCSCs, autophagy was suppressed, the methylation and phosphorylation of RAB37 were elevated, and miR-200b expression was down-regulated. Lentiviral vector (LV-) carrying methionine-γ lyase (an enzyme that could specifically lyse MET; Metase) promoted autophagy, reduced the methylation and phosphorylation of RAB37, and up-regulated miR-200b expression in MET⁺HCY^–-treated GCSCs. Then, we found that miR-200b suppressed the expression of protein kinase C α (PKCα), a protein that could inactivate RAB37 through promoting its phosphorylation. LV-Metase down-regulated RAB37 phosphorylation via miR-200b/PKCα, thus promoting the RAB37-mediated autophagy and suppressing cell viability in MET⁺HCY^–treated GCSCs. Finally, the <i>in vivo</i> study proved that LV-Metase treatment inhibited tumor growth through up-regulating RAB37 expression. In conclusion, MET suppressed RAB37 expression via enhancing its methylation and suppressed RAB37 activity via miR-200b/PKCα axis, thus repressing RAB37-mediated autophagy in GCSCs. The supplementation of Metase lysed MET, thus inducing the autophagy of GCSCs and inhibiting tumor growth.

本研究聚焦于甲硫氨酸（methionine, MET）在胃癌干细胞（gastric cancer stem cells, GCSCs）自噬中的作用，旨在阐明其调控机制。本研究中，研究人员采用抗CD44抗体从人胃癌细胞系中分离得到GCSCs，随后将其培养于MET+同型半胱氨酸（homocysteine, HCY）阴性（HCY-）或MET-HCY+的培养基中。在经MET+HCY-处理的GCSCs中，细胞自噬受到抑制，RAB37的甲基化与磷酸化水平升高，而miR-200b的表达下调。携带甲硫氨酸γ裂解酶（methionine-γ lyase，一种可特异性裂解MET的酶，Metase）的慢病毒载体（lentiviral vector, LV-），可在MET+HCY-处理的GCSCs中促进细胞自噬，降低RAB37的甲基化与磷酸化水平，并上调miR-200b的表达。后续研究发现，miR-200b可抑制蛋白激酶Cα（protein kinase C α, PKCα）的表达，而PKCα可通过促进RAB37的磷酸化使其失活。LV-Metase通过miR-200b/PKCα信号轴下调RAB37的磷酸化水平，进而在MET+HCY-处理的GCSCs中促进RAB37介导的细胞自噬并抑制细胞活力。体内实验证实，LV-Metase治疗可通过上调RAB37的表达抑制肿瘤生长。综上，MET可通过增强RAB37的甲基化抑制其表达，并通过miR-200b/PKCα信号轴抑制RAB37的活性，从而阻遏GCSCs中RAB37介导的细胞自噬。补充Metase可裂解MET，进而诱导GCSCs的细胞自噬并抑制肿瘤生长。