Up-regulating methylation of CAP1 promoter can promote apoptosis and inhibit migration and invasion of lung adenocarcinoma cells.

backgroundLung adenocarcinoma (LUAD) is the most common histological subtype of lung cancer, which poses a significant threat to human health. Adenylate cyclase-associated protein 1 (CAP1) is an important protein closely linked to cancer initiation and progression.

methodTarget Gene fragments were amplified by PCR, and the products of 2 fragments were ligated to construct pdCas9-Dnmt3a-BSD plasmid. Stable cell lines with methylation of CAP1 promoter up-regulated were then established through transfection and screening. Cell proliferation was assessed using colony formation and proliferation assays, while apoptosis was assessed by flow cytometry. Wound healing, transwell migration, and invasion assays were conducted to evaluate cell migration and invasion. Western blot and PCR assays were used to study the expression of molecules involved in apoptosis, migration, and invasion.

resultCAP1 protein expression was higher in early-stage LUAD tissues than in adjacent normal tissues, and elevated in A549, H1299, and PC9 cells as compared to Beas-2B control cells. In addition, CAP1 promoter was abnormally hypo-methylated in LUAD cells and tissues. Up-regulating CAP1 promoter methylation through the CRISPR-dCas9-Dnmt3a system which reduced CAP1 expression can induce apoptosis via the Bax/Bcl-2/Caspase-3 pathway, and inhibited migration and invasion. We also found that the methylation of the CAP1 promoter was regulated by Dnmt3a, Tet1, and/or Tet2.

conclusionUp-regulating CAP1 promoter methylation promotes apoptosis and inhibits migration and invasion of LUAD cells. This suggests that methylating the CAP1 promoter could be a potential therapeutic approach for early-stage LUAD.