Photoregulating RNA digestion using azobenzene linked dumbbell antisense oligodeoxynucleotides

Area of Science:

• Biochemistry
• Molecular Biology
• Oligonucleotide Chemistry

Background:

• Antisense oligodeoxynucleotides (asODNs) are therapeutic agents targeting specific RNA molecules.
• Controlling the stability and activity of asODNs spatially and temporally is crucial for effective gene silencing.
• Azobenzene moieties can undergo photoisomerization, offering a potential mechanism for light-induced structural changes.

Purpose of the Study:

• To design and synthesize azobenzene-linked dumbbell asODNs for light-regulated antisense activity.
• To investigate the impact of azobenzene photoisomerization on hairpin stability and RNA binding.
• To evaluate the photomodulation of RNA digestion by RNase H.

Main Methods:

• Synthesis of azobenzene-modified dumbbell asODNs.
• Thermal melting studies (Tm) to assess hairpin stability.
• UV/visible light irradiation to induce azobenzene trans-cis isomerization.
• Assessment of RNA binding affinity and RNase H-mediated RNA digestion.

Main Results:

• Azobenzene photoisomerization induced significant differences in thermal stability (ΔTm = 12.1-21.3 °C).
• Light irradiation modulated the RNA binding ability of the asODNs.
• Trans-azobenzene asODNs showed limited target RNA binding, which was recovered upon irradiation, enabling RNA digestion by RNase H.

Conclusions:

• Azobenzene-linked dumbbell asODNs offer a promising platform for reversible spatial and temporal regulation of antisense activities.
• The system utilizes both steric binding and RNA digestion by RNase H for controlled gene silencing.
• This technology holds potential for advanced therapeutic applications requiring precise control over gene expression.