The Replication Of T Virus RNA Induces Apoptosis And The Related Pathway In Hela Cells

Objective:Evolution occurs through natural selection. By the pressure of natural evolution and human exploration, it’s possible that some alien viruses would have chance to invade human beings. If their "fresh" host had no defense, these alien viruses might cause series pandemic outbreak, such as HIV, SARS and Avian Influenza Virus, which originated from the Chimpanzee, the common palm civet and even the bird. What would happen, when human beings came across with plant virus, collapse of defense? In order to model this alien virus invasion, we chose distant T virus, and human epithelial carcinoma cell (Hela cells) as its "fresh host", liposome transfection directly imposed T virus RNA into cells.Methods:We got morphological, mRNA, protein evidences for replication, the methods of RT-RCR, Real time PCR, Northern blotting and Western blotting were used to detect the quantity of T virus plus/negative strands RNA^protein and virion. Meanwhile, the same methods applied in detecting the generation expression of T virus RNA. In addition, IF-RNA-FISH was used to analysis the location of T virus plus/negative strands RNA and protein. Live cell image showed the transport of T virus RNA. FITC-Annexin V/PI staining to verify whether the replication of T virus RNA induced cell apoptosis. Besides, live cell image and electronic microscope presented the correlation among T virus RNA/protein、Endoplasmic reticulum stress and autophagy.Results:We report here when artificially import to human epithelial Hela cells, T virus even without specific receptor to human cells can replicate and produce "virion" in Hela cells. It also can express in Hela cell generation and even have great biological activity from Hela cell to plant and animal. In addition to, T virus plus/negative strands locate from cytoplasm to nucleolus; T virus protein (CP) locate on endoplasmic reticulum (ER); T virus RNA transport from ER to nucleolus. FITC-Annexin V/PI staining by Flow Cytometry prove the import of T virus RNA induced cell apoptosis. Besides, verify that T virus RNA trigger autophagy in Hela cells, which was evidenced by the appearance of autophagic vacuoles, conversion of LC3-I to LC3-II; the occurrence of ER stress is supported by the evidence of over-expression of GRP78on ER. With the electronic microscope evidences, we also suggest the formation of autophagosomes is closely related to the expanded ER membrane; T virus protein locate on ER and autophagosome membrane, even accumulate on autophagosome membrane with the time extended. Moreover, we prove T virus RNA transported to fuse with autophagosome on ER. All these evidences demonstrate that the biological behavior of T virus RNA are really associated to ER stress and autophagy, which serve as intarcelluar innate immunity to inhibit or be used by the replication of T virus RNA. At last, we preclude ERS triggered autophagy, inducing cell apoptosis.Significance;In this paper we firstly proved the replication of T virus RNA in Hela cells, we also display a novel focus in the intracellular innate immunity prevent virus, and disclose the possibility for the alien virus invading and influencing even without specific receptors. This work support new potential strategy in virus prevention for human beings.