Inhibitory effect of lncRNA SNHG16 Knockdown on G2/M Transition and Proliferation of Gallbladder Carcinoma Cells via miR-3138/CDC25B/CDK1 Axis Running title: SNHG16’s mechanisms in Gallbladder cancer

Abstract Background Gallbladder cancer (GBC) is the most common and aggressive malignant tumor in the biliary system, and its tumorigenesis and development are associated with abnormal expression of the long non-coding RNA (lncRNA). Although several studies have demonstrated that SNHG16 plays an important role in various tumors, its mechanism in GBC has not been systematically investigated. Methods This study is described from three aspects, including tissue, cell lines, and animal model. The qRT-PCR assay was applied to calculate the mRNA levels of SNHG16, miR-3138, and CDC25B. The link between SNHG16, miR-3138, and CDC25B expression was verified with the dual-luciferase reporter assay, RNA-binding protein immunoprecipitation assay (RIP), rescue assay, gene expression correlation analysis, etc. Results SNHG16 was highly expressed, and miR-3138 was lowly expressed in GBC tissues and cells compared to normal tissues and cells. Low expression of SNHG16 prevents the G2/M transition of the cell cycle by inhibiting the dephosphorylation of CDK1, inducing cell apoptosis, and suppressing cell proliferation in vitro and in vivo. LncRNA SNHG16 directly binds to miR-3138 and regulates miR-3138 expression in GBC-SD and NOZ cells. Cell proliferation and G2/M transition were inhibited by mimics of miR-3138. miR-3138 mediated CDC25B/CDK1 expression by binding directly to CDC25B mRNA. Importantly, the miR-3138 inhibitor could partially reverse the effect of SNHG16 knockdown on cell proliferation, G2/M transition, and cell apoptosis. SNHG16 knockdown reduced CDC25B expression in GBC-SD and NOZ cells. Conclusion As a competitive endogenous RNA, SNHG16 participates in the molecular regulatory network of the GBC cell cycle and proliferation through the mir-3138/CDC25B/CDK1 axis, enhancing the understanding of the pathogenesis of gallbladder cancer.