微小RNA-506-3p调控UHRF1抑制大肠癌细胞转移的机制

MiR-506-3p suppresses metastasis of colorectal cancer by targeting UHRF1

关键词:  大肠癌  转移  microRNA-506-3p  含PHD及指环结构域的类泛素蛋白1

Key words:  colorectal cancer  metastasis  microRNA-506-3p  ubiquitin-like containing PHD and ring finger domain 1

摘要:

目的 探讨微小RNA-506-3p(miR-506-3p)通过调控含PHD及指环结构域的类泛素蛋白1（UHRF1）的表达，进而影响大肠癌细胞转移能力的机制。方法 利用生物信息学的方法预测并筛选UHRF1上游可能发挥调控作用的miRNA-506-3p；在大肠癌细胞及组织中分别检测miR-506-3p及UHRF1的表达水平并分析其相关性；运用荧光素酶报告基因技术检测miR-506-3p与UHRF1 3′非翻译区（3′UTR）特异性结合的情况；荧光定量PCR（qRT-PCR）及Western blot检测大肠癌细胞转染miR-506-3p类似物（mimics）后对UHRF1表达水平的影响；构建UHRF1过表达的大肠癌稳转细胞株，分别转染miR-506-3p mimics及其对照序列（mimics control）后，利用Transwell实验检测处理前后大肠癌细胞转移能力的变化。结果 相对于正常大肠上皮细胞及癌旁组织而言，miR-506-3p在大肠癌细胞及组织中的表达均下调（均P<0.05），且与UHRF1的表达水平呈负相关（r=-0.456，P=0.044）；miR-506-3p通过其“种子序列”与UHRF1 3′UTR特异性结合（P<0.01），并减少了UHRF1的mRNA及蛋白表达（均P<0.01）；转染miR-506-3p mimics抑制了大肠癌LoVo细胞的转移能力（P<0.01），而在此基础上增加UHRF1的表达则逆转了miR-506-3p的抑制作用（P<0.01）。结论 miR-506-3p通过调控UHRF1抑制了大肠癌细胞的转移能力，在大肠癌中发挥抑癌基因的作用。

Abstract:

Objective To explore the effect of microRNA-506-3p(miR-506-3p) on metastasis of colorectal cancer (CRC) and its relation to UHRF1 (ubiquitin-like, containing PHD and ring finger domain 1). Methods Bioinformatics screening was performed to predict UHRF1 as a potential target of miR-506-3p. The expression of miR-506-3p and UHRF1 was detected by qRT-PCR in 20 pairs of CRC and adjacent non-tumoral tissue samples, CRC cell lines LoVo, SW480, HT29, Caco-2 and normal intestinal epithelial cell line NCM460. Luciferase reporter assays, qRT-PCR, Western blotting and Transwell assays were used to evaluate the effects of miR-506-3p on the regulation of UHRF1 in CRC cells. Results Bioinformatics analysis indicated that UHRF1 was a putative target of miR-506-3p. The expression of miR-506-3p decreased in CRC cell lines and in CRC tissues compared to adjacent non-cancerous tissues, and it was negatively correlated with UHRF1 expression. Luciferase reporter assay indicated that miR-506-3p directly bound to 3′-UTR of UHRF1. Over-expression of miR-506-3p decreased the expression of UHRF1. In vitro migration assay showed that synthetic miR-506-3p suppressed UHRF1 expression and reduced metastasis of CRC cells. Conclusion miR-506-3p suppresses metastasis of CRC through targeting UHRF1 and it may play an important role as a tumor suppressor in CRC.