ALKBH5通过m6A去甲基化下调MCM7表达而抑制结直肠癌增殖和转移OA

AIM:To investigate the effect of alkylation repair homologue 5(ALKBH5)on the proliferation and metastasis of colorectal cancer(CRC)through the downregulation of minichromosome maintenance protein 7(MCM7)expression by N6-methyladenosine(m6A)demethylation.METHODS:The expression levels of ALKBH5 in CRC tissues and adjacent tissues were detected by RT-qPCR,Western blot and immunohistochemical staining,and survival analysis was performed.Differences in ALKBH5 expression between CRC cell lines(HCT116,HT29,SW480 and SW620)and the normal human colonic cell line NCM460 were compared.Lentiviral transfection was used to construct stable overex-pression and knockdown cell lines.Cell counting experiments,cell colony formation experiments and cell cycle detection experiments were performed to determine the effects of ALKBH5 on the proliferation and metastasis of CRC cells,and changes in m6A levels were detected.Subcutaneous tumour xenograft and liver metastasis tumour models in nude mice were used to detect the in vivo effect of overexpressed ALKBH5.Bio-big data analysis,luciferase reporter gene assays,m6A immunoprecipitation PCR and Western blot were used to verify the mechanism by which ALKBH5 regulates MCM7.RESULTS:Compared with that in adjacent normal tissues and normal human colonic cell lines,the expression of ALK-BH5 was frequently downregulated in CRC tissues and the four CRC cell lines(HCT116,HT29,SW480 and SW620),and its low expression predicted poorer prognosis of CRC(P<0.05).The knockdown of ALKBH5 expression significantly promoted the proliferation and migration of HT29 and SW620 cells and increased the m6A content,whereas its overexpres-sion inhibited the growth and metastasis of CRC both in vitro and in vivo.ALKBH5 targeted and modified MCM7 through the m6A binding site.Knockdown of ALKBH5 significantly increased the expression of MCM7,whereas overexpression of ALKBH5 had the opposite effect.Furthermore,the expression level of MCM7 increased in CRC tissues,and overexpres-sion of MCM7 dramatically increased cyclin E1 expression.CONCLUSION:ALKBH5 inhibits the proliferation and me-tastasis of colorectal cancer by reducing the m6A level of MCM7 mRNA,thereby downregulating MCM7 expression.