TRIM25泛素化IGF2BP3后抑制食管癌EC109细胞的活力

目的:探讨含三重基序蛋白25(tripartite motif-containing protein 25, TRIM25)和胰岛素样生长因子2 mRNA结合蛋白3(insulin-like growth factor 2 mRNA-binding protein 3,IGF2BP3)与人食管癌EC109细胞活力的关系。方法:将EC109细胞分为过表达TRIM25组、过表达IGF2BP3组、敲减TRIM25组和敲减IGF2BP3表达且过表达TRIM25组,用MTT法和CCK-8法测定EC109细胞的活力;检测过表达TRIM25对IGF2BP3稳定性的影响;用免疫共沉淀技术检测TRIM25和IGF2BP3之间的相互作用;对EC109细胞转染过表达TRIM25载体或空载体,用免疫沉淀技术检测IGF2BP3泛素化程度的差异。结果:过表达TRIM25时,EC109细胞的活力受到抑制;过表达IGF2BP3时,EC109细胞的活力受到促进;敲减IGF2BP3表达的同时过表达TRIM25,EC109细胞的活力与单纯敲减IGF2BP3表达组比较无明显变化;过表达TRIM25时IGF2BP3的表达量减少...

TRIM25 ubiquitinates IGF2BP3 and inhibits viability of esophageal carcinoma EC109 cells

AIM:To investigate the relationship among tripartite motif-containing protein 25(TRIM25),insulin-like growth factor 2 mRNA-binding protein 3(IGF2 BP3)and viability of human esophageal carcinoma EC109 cells.METHODS:The EC109 cells were divided into TRIM25 over-expression group,IGF2 BP3 over-expression group,TRIM25 knock-down group,and IGF2 BP3 knock-down and TRIM25 over-expression group.The viability of EC109 cells was mea-sured by MTT assay and CCK-8 assay.The stability of IGF2 BP3 was detected by Western blot.The interaction between TRIM25 and IGF2 BP3 was evaluated by immunoprecipitation,and the TRIM25 vector or empty vector was transfected to detect the ubiquitination of IGF2 BP3 by immunoprecipitation.RESULTS:Over-expression of TRIM25 inhibited,but over-expression of IGF2 BP3 promoted the viability of EC109 cells.However,the viability of the cells with knock-down of IGF2 BP3 and over-expression of TRIM25 was lower than that of the cells with knock-down of IGF2 BP3 only.Over-expression of TRIM25 resulted in reducing the expression level of IGF2 BP3,which was recovered if the cells were treated with MG132,a proteasome inhibitor.When TRIM25 expression was knocked down,the viability of EC109 cells was significantly promoted on days 2,3 and 4.The interaction between TRIM25 and IGF2 BP3 was confirmed.At the same time,over-expression of TRIM25 increased the ubiquitination degree of IGF2 BP3 in the EC109 cells.CONCLUSION:TRIM25 ubi-quitinates IGF2 BP3,resulting in degradation of IGF2 BP3 by proteasomes,thereby inhibiting the viability of esophageal carcinoma cells.