title: CHK1 activity is required for continuous replication fork elongation but not stabilization of post-replicative gaps after UV irradiation.
abstract: Ultraviolet (UV)-induced DNA damage causes an efficient block of elongating replication forks. The checkpoint kinase, CHK1 has been shown to stabilize replication forks following hydroxyurea treatment. Therefore, we wanted to test if the increased UV sensitivity caused by the unspecific kinase inhibitor caffeine--inhibiting ATM and ATR amongst other kinases--is explained by inability to activate the CHK1 kinase to stabilize replicative structures. For this, we used cells deficient in polymerase η (Polη), a translesion synthesis polymerase capable of properly bypassing the UV-induced cis-syn TT pyrimidine dimer, which blocks replication. These cells accumulate gaps behind progressing replication forks after UV exposure. We demonstrate that both caffeine and CHK1 inhibition, equally retards continuous replication fork elongation after UV treatment. Interestingly, we found more pronounced UV-sensitization by caffeine than with the CHK1 inhibitor in clonogenic survival experiments. Furthermore, we demonstrate an increased collapse of replicative structures after caffeine treatment, but not after CHK1 inhibition, in UV-irradiated cells. This demonstrates that CHK1 activity is not required for stabilization of gaps induced during replication of UV-damaged DNA. These data suggest that elongation and stabilization of replicative structures at UV-induced DNA damage are distinct mechanisms, and that CHK1 is only involved in replication elongation.
authors: Department of Genetics, Microbiology, and Toxicology, Stockholm University, S-10691, Stockholm, Sweden.
authors: Ingegerd
authors: I
authors: Elvers
authors: Anna
authors: A
authors: Hagenkort
authors: Fredrik
authors: F
authors: Johansson
authors: Tatjana
authors: T
authors: Djureinovic
authors: Anne
authors: A
authors: Lagerqvist
authors: Niklas
authors: N
authors: Schultz
authors: Ivaylo
authors: I
authors: Stoimenov
authors: Klaus
authors: K
authors: Erixon
authors: Thomas
authors: T
authors: Helleday
mesh_terms: D002110:Caffeine / Q000494:pharmacology; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D002461:Cell Line, Transformed; D002470:Cell Survival; D000071877:Checkpoint Kinase 1; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D004261:DNA Replication / Q000187:drug effects / Q000528:radiation effects; D004259:DNA-Directed DNA Polymerase / Q000172:deficiency; D006801:Humans; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000528:radiation effects; D014466:Ultraviolet Rays; D000099270:Y-Family DNA Polymerases
chemical_list: D002110:Caffeine; D000071877:Checkpoint Kinase 1; D004259:DNA-Directed DNA Polymerase; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; C000605867:CHEK1 protein, human; C112338:Rad30 protein; D000099270:Y-Family DNA Polymerases