Untargeted Mutation Triggered by Ribonucleoside Embedded in DNA

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• DNA polymerases can misincorporate ribonucleoside 5'-triphosphates into DNA strands.
• Incorporated ribonucleosides can lead to DNA damage and mutations.

Purpose of the Study:

• To investigate the impact of incorporated ribonucleosides on untargeted mutations in human cells.
• To elucidate the mechanism by which ribonucleosides induce mutations.

Main Methods:

• Introduction of riboguanosine (rG) into the supF gene in a plasmid.
• Transfection into U2OS cells and DNA recovery after 48 hours.
• Mutation analysis using Escherichia coli RF01 strain and RNA interference for APOBEC3B knockdown.

Main Results:

• Frequent untargeted base substitutions were observed at G bases within 5'-GpA-3' dinucleotides.
• These mutations were reduced by approximately 80% upon APOBEC3B knockdown.
• The mutation pattern resembled action-at-a-distance mutations induced by oxidative damage or APOBEC3 activity.

Conclusions:

• Embedded ribonucleosides induce base substitution mutations at G bases through an APOBEC3B-dependent mechanism.
• Ribonucleosides may contribute to APOBEC3-dependent cancer initiation events.