Title: Unravelling an important DNA knot – analysis of HBV’s promoter G4-quadruplex
Background: Hepatitis B virus (HBV) chronically persists due to covalently-closed circular DNA (cccDNA). Recently, a G4-quadruplex (G4Q) in HBV's pre-core promoter, that anchors the transcriptional protein Sp1, was discovered and could be an approach to targeting this chronic form.
Purpose: Identify key nucleic acids responsible for HBV's G4Q-Sp1 interaction using a luciferase assay.
Methods: Using a luciferase plasmid under HBV’s preC promoter control, mutations are introduced in G4Q-accessible regions. Plasmids are transfected into HepG2 cells and luciferase activity is measured.
Results: Eleven different G4Q mutations were successfully created. A subset of mutations in the loop regions significantly reduced activity compared to the wild type, indicating its importance in the G4Q function.
Conclusion: The loop regions of HBV’s G4Q appear essential for Sp1 interaction and function. This will enhance our insight into the G4Q-Sp1 interaction potentially guide cccDNA-targeting therapies.