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AIM: To explore the effects of the nucleoside analogues β-L-D4A and β-LPA on hepatitis B virus (HBV) promoters. METHODS: Four HBV promoters were amplified by polymerase chain reaction (PCR) and subcloned into the expression vector pEGFP-1. The four recombinants controlled by HBV promoters were confirmed by restriction analysis and sequencing. Human hepatoma HepG2 cells transfected with the recombinant plasmids were treated with various concentrations of β-L-D4A and β-LPA. Then, enhanced green fluorescent protein (EGFP)-positive cells were detected by fluorescence microscopy and using a fluorescence activated cell sorter (FACS). RESULTS: Four HBV promoters were separately obtained and successfully cloned into pEGFP-1. Expression of EGFP under the control of the surface promoter (Sp) and the X promoter (Xp) was inhibited by β-L-D4A in a dose-dependent manner, while expression of EGFP under the control of the core promoter (Cp) and Xp was inhibited by β-LPA in a dose-dependent manner. CONCLUSION: The two novel nucleoside analogues investigated here can inhibit the activities of HBV promoters in a dose-dependent manner. These findings may explain the mechanisms of action by which these two novel compounds inhibit HBV DNA replication.
AIM: To explore the effects of the nucleoside analogues β-L-D4A and β-LPA on hepatitis B virus (HBV) promoters. METHODS: Four HBV promoters were amplified by polymerase chain reaction (PCR) and subcloned into the expression vector pEGFP- 1. The four recombinants controlled by HBV promoters were confirmed by restriction analysis and sequencing. Human hepatoma HepG2 cells transfected with the recombinant plasmids were treated with various concentrations of β-L-D4A and beta-LPA. Then, enhanced green fluorescent protein (EGFP ) -positive cells were detected by fluorescence microscopy and using a fluorescence activated cell sorter (FACS). RESULTS: Four HBV promoters were separately obtained and successfully cloned into pEGFP-1. Expression of EGFP under the control of the surface promoter (Sp) and the X promoter (Xp) was inhibited by β-L-D4A in a dose-dependent manner, while expression of EGFP under the control of the core promoter (Cp) and Xp was inhibited by β-LPA in a dose-dependent manne r. CONCLUSION: The two novel nucleoside analogues investigated here can inhibit the activities of HBV promoters in a dose-dependent manner. These findings may explain the mechanisms of action by which two novel compounds inhibit HBV DNA replication.