MutSβ regulates G4-associated telomeric R-loops to maintain telomere integrity in ALT cancer cells

Area of Science:

• Oncology
• Molecular Biology
• Genetics

Background:

• Alternative lengthening of telomeres (ALT) is a telomerase-independent mechanism maintaining telomere length in ~15% of human cancers.
• Telomere maintenance in ALT cancers involves homologous recombination and is linked to RNA:DNA hybrids (R-loops) formed by telomeric repeat-containing RNA (TERRA).

Purpose of the Study:

• To investigate the role of the mismatch repair protein MutSβ in telomere maintenance within ALT cancer cells.
• To elucidate how MutSβ influences G-quadruplex (G4) structures and R-loops at ALT telomeres.

Main Methods:

• Immunofluorescence to detect MutSβ enrichment at telomeres in ALT cancer cells.
• CRISPR/Cas9-mediated depletion of MSH3 to assess telomere integrity and C-circle formation.
• In vitro biochemical assays using purified MutSβ to study G4 structure destabilization.

Main Results:

• MutSβ, composed of MSH2 and MSH3, is enriched at telomeres in ALT cancer cells.
• MSH3 depletion leads to increased R-loop-dependent telomere fragility and C-circle accumulation.
• Purified MutSβ destabilizes G4 structures in vitro.

Conclusions:

• MutSβ plays a critical role in ALT telomere maintenance by preventing the accumulation of G4 structures and TERRA R-loops.
• Destabilization of G4 structures by MutSβ is essential for regulating R-loops and maintaining telomere integrity in ALT cancers.