Heliothis Armigera Nucleopolyhdrovirus Infection Results The Host Cell Cycle Arrest With Alteration Of Mitotic Cyclins

Viruses modify their host cells in many ways to optimize their own replication. In turn, the host cell modifies its metabolism in response to stresses such as viral infection in attempt to protect itself. A lot of viruses can induce cell cycle arrest, for example paryoviruses, herpesviruses, retroviruses et al. Little is known of the cell-cycle-related interations between baculovirus and host cells. We choose HaSNPV and the host cell-Hz cell to investigate the effect of baculovirus infection on the insect cell cycle. All results as follows.1. Cell cycle phase distributions and durations of Hz cellsFlow cytometry indicated that unsynchronized Hz cell culture had 47-53% of the cells in GI phase, 20-27% in S phase and 33-20% in the Gi/M phase. We also find there are not cells with >4N DNA content in Hz cell culture. Hydroxyurea successfully arrested Hz cells in GI phase. After 24h of incubation with 60ug/ml hydroxyurea, 74% cells were arrested at GI phase. To estimate the duration of individual cell cycle were synchronized and released from the block. Flow cytometry indicated culture doubling time was estimated to be 40h and the duration of the GI, S, G2/M phases was estimated to be 10, 14 and 16h, respectively.2. Effect of HaSNPV on Hz cell cycleWhen unsynchronized Hz cells were affected by HaSNPV, Hz cells began to accumulate in G2/M phase as early as 8h postinfection. There are 44% cells in Ga/M phase by 8h. After 24h postinfection, a significant population of cells were in G2/M phase and remained there for the duration of infection. In all experiments, infection resulted in approximately 70% of cells arrest in G2/M phase. Flow cytometry also indicated 14.30% cells with more than 4N DNA content appearedafter 24h postinfection and accumulated to high levels after 72h postinfection. We noted that at times when most of the cells were arrested in Ga/M phase, the number of cells with >4N DNA continued to increase. Effect of HaSNPV infection on the cell-cycle phase distribution was examined in Hz cells synchronized in G1 phase, comparing with the mock-infection Hz cells. We also find that HaSNPV infection results synchronized Hz cells arrest at G2/M phase.3. HaSNPV infection affects cyclin of the host cellsWhen HaSNPV infection the unsynchronized Hz cells, expression levels of cyclinBi were significantly increased after 16h of infection, reach peaks 48h postinfection. When Hz cells were synchronized by hydroxyurea, HaSNPV infection also induces the accumulation of cyclinBi. We examined the subcellular location of cyclinBi in mock-infected cells and Hz cells after 24h of infection by Confocal scanning light microscope. In mock-infection cells, cyclinBi was detectable in both the cytoplasm and nucleus. But there is a restricted localization of cyclinBi to only the cytoplasm, but not the nucleus. The results suggested the possible that HaSNPV infection induces the Hz cell cycle arrest at the G2 phase. We also examined the expression levels of cyclinA after HaSNPV infection and found HaSNPV infection can't induce the alternation of the cyclinA levels.