Analysis of the function of paramyxovaus V protein

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1. I identified a host protein, AIPI/Alix, involved in apoptosis and efficient budding of several enveloped viruses as an interacting partner of hPIV2 NP and V proteins. To investigate possible involvement of AIPI in hPIV2 virus growth, I established cell lines in which were depleted the intracellular AIP 1 by using short hairpin RNA induced by adding Dox. The wt hPIV2 growth in the established cells was inhibited by 10- to 100-fold, but the growth rates of rPIV2s that have mutant V proteins of inability to bind with AIP I were similar. These results have shown that the association between AIPI/Alix and V proteins is important for virus growth.2. I investigated the role of the V protein in hPIV2 replication, using a mini-genome system free of vaccinia virus. I found that the C-terminal of the V protein was essential for the inhibition and for interaction with the L protein, but not for interaction with the NP protein. These data suggest that the inhibitory effect of the hPIV2 V protein is the result of L protein binding, and not that of NP.3. I found that a homogeneous set of NP, P and L proteins of PIV2 or PIV5 was able to drive reporter gene expression from the hPIV2 mini-genome system, but all heterogeneous combinations were inactive.4. By using a yeast two-hybrid-based screening assay with hPIV2 V as bait and a target cDNA library, I got some positive clones. I would like to investigate the interaction between these clones and V.