DNA2 and MSH2 cooperatively repair stabilized G4 and allow efficient telomere replication

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• G-quadruplexes (G4s) are stable DNA secondary structures crucial for cellular processes.
• Timely resolution of G4s is hypothesized to be essential for efficient and accurate DNA replication.
• The DNA2 nuclease-mediated G4 resolution pathway's biological significance remains largely unknown.

Purpose of the Study:

• To investigate the biological role of the DNA2-mediated G4 removal pathway.
• To elucidate the regulatory mechanism of G4 resolution by DNA2.
• To understand the impact of G4 stabilizers on telomere replication.

Main Methods:

• Single molecular analysis of replicating DNA (SMARD) to detect G4s and stalled replication forks.
• Assessing the effects of DNA2 deficiency or inhibition on G4 accumulation and telomere replication.
• Investigating the interaction between MutSα (MSH2-MSH6) and G4s.
• Evaluating the impact of G4-stabilizing compounds on DNA2 and helicase activity.

Main Results:

• DNA2 deficiency or inhibition led to significant G4 accumulation and stalled replication forks at telomeres.
• The MutSα complex binds G4s and promotes their resolution via DNA2-mediated G4 excision.
• MSH2 deficiency mirrored DNA2 deficiency effects, causing G4 accumulation and defective telomere replication.
• G4 stabilizers blocked helicase unwinding but not DNA2 cleavage, impairing telomere replication and causing instability, particularly in DNA2- or MSH2-deficient cells.

Conclusions:

• DNA2 and MutSα are critical for resolving G-quadruplexes during DNA replication.
• The DNA2-mediated pathway is essential for preventing G4 accumulation and maintaining telomere replication fidelity.
• G4 stabilizers pose a significant risk to telomere stability, especially in cells with compromised G4 resolution pathways.