Sequence-Resolved Imaging of mRNA G-Quadruplexes for Profiling the Translational Suppression in Oncogene Expression

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• G-quadruplexes (G4s) are crucial non-DNA/RNA secondary structures involved in biological regulation.
• Current G4 detection methods lack the sequence resolution for imaging within specific mRNA transcripts in living cells.

Purpose of the Study:

• To develop a novel technology for sequence-resolved imaging of G4 structures within specific mRNA transcripts in living cells.
• To investigate the functional role of G4 structures in mRNA regulation, specifically in BCL2 mRNA.

Main Methods:

• A smart and versatile orthogonal recognition strategy (SORS) was developed, combining sequence-specific (Seq-module) and structure-specific (Str-module) targeting.
• SORS utilizes hybridization chain reaction for amplified fluorescence signal generation upon dual recognition of target mRNA G4s.
• The method was applied to detect and image G4 structures within BCL2 mRNA in living cells.

Main Results:

• SORS enabled sequence-resolved detection and imaging of G4 structures within BCL2 mRNA in living cells.
• A strong correlation was observed between high G4-folding percentage and reduced BCL2 protein expression.
• Direct evidence was provided for the translational suppressive role of the G4 structure in BCL2 mRNA.

Conclusions:

• The SORS strategy offers a versatile and robust platform for studying mRNA secondary structure dynamics at the sequence level.
• This technology has potential for developing oligonucleotide-based molecular diagnostics and therapeutics.
• G4 structures in mRNA play a significant role in regulating gene expression at the translational level.