The invasion of tobacco mosaic virus RNA induces endoplasmic reticulum stress-related autophagy in HeLa cells

Li, Li; Wang, Li; Xiao, Ruijing; Zhu, Guoguo; Li, Yan; Liu, Changxuan; Yang, Ru; Tang, Zhiqing; Li, Jie; Huang, Wei; Chen, Lang; Zheng, Xiaoling; He, Yuling; Tan, Jinquan

Because there is no receptor for TMV on human cellular membranes, TMV was not able to enter HeLa cells; when TMV was co-cultured with HeLa cells, CP was not detected in HeLa cells. To import intact TMV particles or TMV-RNA into HeLa cells, we used two methodologies: electroporation or liposome transfection, respectively. Following the electroporation procedure [27] , intact TMV particles were transformed into HeLa cells by electric shock with different voltages (0, 140, 280 and 360 V). Western blotting results showed that there was a little more CP expression in HeLa cells treated with 280 V than with the other voltages, but not high enough in CP expression ( Figure 1A-a) . This result suggests that some cells may have been fatally damaged by the strong electric currents. Thus we concluded that electroporation was not the proper method for TMV transfection. Recent reports have demonstrated that the liposome transfection method is more effective and less toxic to cells than other transfection methods [28] . Thus we used this method for TMV-RNA transfection. HeLa cells were incubated with a mixture of TMV-RNA and liposomes at ratios of 1:1 and 1:2 for 6 h. Western blot analysis showed that CP was produced more abundantly at the ratio of 1:1 than 1:2 ( Figure 1A -b), indicating that TMV-RNA can be effectively transfected into HeLa cells by using an equal proportion of liposomes. Moreover, the morphology of the cells did not change during the process of incubation. Thus we found an effective way to import TMV-RNA into HeLa cells, and this technique was applied exclusively in the following experiments.