Objective:To study the radiosensitization effect of homocysteine-inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1 ( HerpUD1 ) on human liver L-02 cells through silencing the expression of HerpUD1 gene by RNA interference technology. Methods: Based on the HerpUD1 gene sequence, shRNA expression vector interfering HerpUD1 gene was constructed and L-02 cells were stably transfected. The expressions of HerpUD1 mRNA and protein in cells after transfection were detected by real-time PCR and Western blotting. The transfected cells were radiated by 8 Gy X ray and the cell apoptosis was analyzed by Hoechst fluorescence assay. Results: Eukaryotic expression interference vector targeting HerpUD1 gene was successfully constructed and L-02 cell line stably transfected with shHerpUD1 was established. The shHerpUD1-1420 interference fragment had the highest interference efficiency in stably transfected L-02 cells. Compared with pGPU6-NC (empty vector) group, the expression level of HerpUD1 mRNA in shHerpUD1-1420 group was significantly decreased (\[0.15±005\] vs \[1.00±0.08\], P<0.05), and the expression level of HerpUD1 protein was also significantly reduced (\[0.19±0.01\] vs \[1.62±0.08\], P<0.01). The apoptotic rate was higher in L-02-shHerpUD1-1420 group than that in L-02-NC group (negative control group) 24 h after 8 Gy X ray irradiation (\[1023±3.37\]% vs \[6.89±1.49\]%, P<0.05), while there was no significant difference between L-02-NC and L-02-Neo group. Conclusion:Silencing the expression of HerpUD1 gene significantly increases the X ray radiation-induced apoptosis of L-02 cells, therefore playing an role in radiosensitization.