Induction of DNA damage responses by adozelesin is S phase-specific and dependent on active replication forks.
Adozelesin is an alkylating minor groove DNA binder that is capable of rapidly inhibiting DNA replication in treated cells through a trans-acting mechanism and preferentially arrests cells in S phase. It has been shown previously that in cells treated with adozelesin, replication protein A (RPA) activity is deficient, and the middle subunit of RPA is hyperphosphorylated. The adozelesin-induced RPA hyperphosphorylation can be blocked by the replicative DNA polymerase inhibitor, aphidicolin, suggesting that adozelesin-triggered cellular DNA damage responses require active DNA replication forks. These data imply that cellular DNA damage responses to adozelesin treatment are preferentially induced in S phase. Here, we show that RPA hyperphosphorylation, RPA intranuclear focalization, and gamma-H2AX intranuclear focalization induced by adozelesin treatment are all dependent on DNA replication fork progression, and focalization is only induced in S phase cells. These findings are similar to those seen with the S phase-specific DNA-damaging agent, camptothecin. Conversely, all three DNA damage responses are independent of either S phase or replication fork progression when induced by treatment with the DNA strand scission agent, C-1027. Furthermore, we demonstrate that adozelesin-induced RPA and gamma-H2AX intranuclear foci appear to colocalize within the nuclei of S phase cells.
Liu
Jen-Sing
JS
Departments of Microbiology and Biochemistry and the Witebsky Center for Microbial Pathogenesis and Immunology, University at Buffalo, SUNY, School of Medicine and Biomedical Sciences, Buffalo, New York 14214, USA.
Kuo
Shu-Ru
SR
Beerman
Terry A
TA
Melendy
Thomas
T
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents / Q000633:toxicity; D000903:Antibiotics, Antineoplastic / Q000633:toxicity; D018906:Antineoplastic Agents, Alkylating / Q000633:toxicity*; D016590:Aphidicolin / Q000494:pharmacology; D001572:Benzofurans; D002166:Camptothecin / Q000633:toxicity*; D003509:Cyclohexanecarboxylic Acids / Q000633:toxicity*; D053138:Cyclohexenes; D004249:DNA Damage; D004261:DNA Replication / Q000187:drug effects*; D000080890:Duocarmycins; D053281:Enediynes; D006367:HeLa Cells; D006801:Humans; D007211:Indoles; D016196:S Phase / Q000187:drug effects
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D000903:Antibiotics, Antineoplastic; D018906:Antineoplastic Agents, Alkylating; D001572:Benzofurans; D003509:Cyclohexanecarboxylic Acids; D053138:Cyclohexenes; D000080890:Duocarmycins; D053281:Enediynes; D007211:Indoles; C058024:C 1027; D016590:Aphidicolin; C060090:adozelesin; D002166:Camptothecin