HERC2 Facilitates BLM and WRN Helicase Complex Interaction with RPA to Suppress G-Quadruplex DNA

BLM and WRN are RecQ DNA helicases essential for genomic stability. Here, we demonstrate that HERC2, a HECT E3 ligase, is critical for their functions to suppress G-quadruplex (G4) DNA. HERC2 interacted with BLM, WRN, and replication protein A (RPA) complexes during the S-phase of the cell cycle. Depletion of HERC2 dissociated RPA from BLM and WRN complexes and significantly increased G4 formation. Triple depletion revealed that HERC2 has an epistatic relationship with BLM and WRN in their G4-suppressing function. In vitro, HERC2 released RPA onto single-stranded DNA (ssDNA) rather than anchoring onto RPA-coated ssDNA. CRISPR/Cas9-mediated deletion of the catalytic ubiquitin-binding site of HERC2 inhibited ubiquitination of RPA2, caused RPA accumulation in the helicase complexes, and increased G4, indicating an essential role for E3 activity in the suppression of G4. Both depletion of HERC2 and inactivation of E3 sensitized cells to the G4-interacting compounds telomestatin and pyridostatin. Overall, these results indicate that HERC2 is a master regulator of G4 suppression that affects the sensitivity of cells to G4 stabilizers. Given that HERC2 expression is frequently reduced in many types of cancers, G4 accumulation as a result of HERC2 deficiency may provide a therapeutic target for G4 stabilizers. Significance: HERC2 is revealed as a master regulator of G-quadruplex, a DNA secondary structure that triggers genomic instability and may serve as a potential molecular target in cancer therapy. Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/22/6371/F1.large.jpg. (C) 2018 AACR.