Construction of the recombinant expression lentivirus vector carrying HIV-1 Vif gene and effect of its protein expression on the lytic cycle replication of KSHV

Objective:To construct the recombinant lentivirus containing HIV-1 Vif gene and detect the effect of Vif protein expression on the lytic cycle replication of KSHV. Methods:The fragment of Vif gene from expression vector pCI-neo-Vif was cloned into the lentivirus vector pHAGE-CMV-MCS-IzsGreen,then the recombinant plasmid pHAGE-Vif,packaging vector psPAX2 and envelope vector pMD2.G were cotransfected into the 293T cells. Culture media were harvested and filtered through a 0.45 μm filter to remove the cells. The viral titer was checked by observing the expression of green fluorescent protein(GFP). After infected with the recombinant lentivirus,the mRNA transcription and protein expression of Vif in 293T cells were detected by RT-PCR and Western blot,respectively. Then,BCBL-1 cells were infected by the recombinant lentivirus with the MOI of 1.0,and the protein expression of Vif,vIL-6 and Rta were detected by Western blot. Results:The recombinant lentivirus vector carrying Vif was constructed successfully with the viral titer of 4×107 efu / ml. BCBL-1 cells could be efficiently infected by the recombinant lentivirus with the MOI of 1.0,and Vif protein was readily expressed in these cells. Moreover,the expression of Rta and vIL-6 proteins of KSHV were significantly downregulated by Vif protein. Conclusion:Recombinant lentivirus with high titer and efficient infection of BCBL-1 cells could be obtained quickly and simply by using the lentivirus vectors system,and Vif expression could be observed in BCBL-1 cells infected with lentivirus-Vif. Furthermore,the Vif protein may inhibit the lytic cycle replication of KSHV. The results of this study lay the foundation for further studying on the molecular mechanisms and signaling pathways involved.