关键词: β-catenin 蛋白, 突变, 肝癌, 增殖 

Abstract: Objective To investigate the effect of β-catenin H36P mutation on the proliferation of hepatocellular carcinoma cells. Methods The CTNNB1genes (WT-CTNNB1, G34R-CTNNB1, H36P-CTNNB1) were amplified by PCR amplification. It was cloned into the p-CMV5 plasmid vector with a double digestion method, and the corresponding plasmids were transiently transferred into the Huh7 and HepG2 cells. The protein expression levels were then detected by Western blotting. Cells transfected with the target genes were treated with CHX in a time gradient, the degradation time of the H36P-β-catenin, WT-β-catenin and G34R-β-catenin were compared and their stabilities were analyzed. The nuclear translocation ability of H36P-β-catenin was examined after separation of the nuclear and cytoplasmic fractions. Finally, the effect of H36P-β-catenin on the proliferation of hepatocellular carcinoma cells was examined by CCK-8 method. Results The protein expression levels of β-catenin mutants (G34R and H36P) were not significantly different from that of the wild-type β-catenin. In HepG2 cell line, WT-β-catenin was almost completely degraded by CHX in 8 hours, while the mutants only degraded 75 % and 60 % in the same time period. Similarly, in the Huh7 cell line, WT-β-catenin was almost completely degraded in 6 hours, while the mutants were only degraded about 40 % in the same time period. The differences in the above results were statistically significant (P< 0. 05). Compared with the WT-β-catenin, mutant β-catenin promoted the proliferation of HepG2 cell. The nuclear translocation ability of H36P mutant β-catenin was increased compared with those of WT and G34R mutant β-catenin in the hepatocellular carcinoma cells. Conclusion β-catenin H36P mutant protein blocks the protein ubiquitination degradation process, promotes the proliferation of HepG2 and Huh7 hepatocellular carcinoma cells, and has an increased nuclear translocation.

Key words: β-catenin, mutation, hepatocellular carcinoma cells, proliferation