Structural and functional analysis of dengue virus RNA

Area of Science:

• Virology
• Molecular Biology
• Genetics

Background:

• The 5' and 3' untranslated regions (UTRs) of RNA viruses contain critical sequences and structures influencing translation, RNA synthesis, and viral assembly.
• Dengue virus (DENV), a mosquito-borne flavivirus, utilizes complementary sequences (CS and UAR) at its genome ends for RNA-RNA interactions, essential for genome circularization and viral viability.

Purpose of the Study:

• To investigate the molecular mechanisms of RNA-RNA interactions in the DENV life cycle.
• To differentiate the roles of 5' and 3' UTR RNA elements in translation versus RNA synthesis.

Main Methods:

• Development of a DENV replicon system for transfection into mosquito and mammalian cells.
• Analysis of mutations within CS and UAR regions affecting base pairing.
• Systematic mutational analysis of UAR sequences.

Main Results:

• Mutations disrupting CS and UAR base pairing did not significantly impact input RNA translation.
• These mutations severely compromised or abolished viral RNA synthesis.
• Specific nucleotides within the UAR sequence were identified as crucial for efficient RNA synthesis, beyond their role in cyclization.

Conclusions:

• DENV 5'/3' UTR RNA-RNA interactions, mediated by CS and UAR, are essential for viral RNA synthesis, not translation initiation.
• The UAR region plays a dual role, facilitating genome cyclization and directly promoting efficient RNA synthesis through specific nucleotide sequences.