SMI#9 attenuated cisplatin-induced PCNA monoubiquitination (TLS marker), FANCD2 (Fanconi anemia (FA) activation marker), and TLS polymerase POL η. SMI#9-induced decreases in γH2AX levels were associated with concomitant inhibition of H2AX monoubiquitination, suggesting a key role for RAD6 in modulating cisplatin-induced γH2AX via H2AX monoubiquitination.
Importantly, Pol kappa-mediated DNA synthesis during hydroxyurea (HU)-dependent fork restart is regulated by both the Fanconi Anemia (FA) pathway and PCNA polyubiquitination.
The compromised DNA repair and chemosensitization induced by SMI#9 or <i>RAD6B</i> depletion were associated with decreased platinum drug-induced proliferating cell nuclear antigen (PCNA) and FANCD2 monoubiquitinations (surrogate markers of TLS and FA pathway activation, respectively) and with attenuated FANCD2, RAD6, γH2AX, and POL η foci formation and cisplatin-adduct removal.
Interestingly, proliferating cell nuclear antigen (PCNA) monoubiquitination, known to contribute to Rev1 recruitment, does not require FA core complex function.