Inhibition of human Chk1 causes increased initiation of DNA replication, phosphorylation of ATR targets, and DNA breakage.

TY - JOUR

T1 - Inhibition of human Chk1 causes increased initiation of DNA replication, phosphorylation of ATR targets, and DNA breakage.

AU - Syljuåsen, Randi G

AU - Sørensen, Claus Storgaard

AU - Hansen, Lasse Tengbjerg

AU - Fugger, Kasper

AU - Lundin, Cecilia

AU - Johansson, Fredrik

AU - Helleday, Thomas

AU - Sehested, Maxwell

AU - Lukas, Jiri

AU - Bartek, Jiri

N1 - Keywords: Caffeine; Cell Cycle Proteins; Chromosomal Proteins, Non-Histone; DNA Damage; DNA Replication; DNA, Single-Stranded; DNA-Binding Proteins; Histones; Humans; Phosphorylation; Protein Kinase Inhibitors; Protein Kinases; Protein-Serine-Threonine Kinases; Purines; RNA, Small Interfering; Replication Protein A; Staurosporine; Tumor Suppressor Protein p53

PY - 2005

Y1 - 2005

N2 - Human checkpoint kinase 1 (Chk1) is an essential kinase required to preserve genome stability. Here, we show that Chk1 inhibition by two distinct drugs, UCN-01 and CEP-3891, or by Chk1 small interfering RNA (siRNA) leads to phosphorylation of ATR targets. Chk1-inhibition triggered rapid, pan-nuclear phosphorylation of histone H2AX, p53, Smc1, replication protein A, and Chk1 itself in human S-phase cells. These phosphorylations were inhibited by ATR siRNA and caffeine, but they occurred independently of ATM. Chk1 inhibition also caused an increased initiation of DNA replication, which was accompanied by increased amounts of nonextractable RPA protein, formation of single-stranded DNA, and induction of DNA strand breaks. Moreover, these responses were prevented by siRNA-mediated downregulation of Cdk2 or the replication initiation protein Cdc45, or by addition of the CDK inhibitor roscovitine. We propose that Chk1 is required during normal S phase to avoid aberrantly increased initiation of DNA replication, thereby protecting against DNA breakage. These results may help explain why Chk1 is an essential kinase and should be taken into account when drugs to inhibit this kinase are considered for use in cancer treatment.

AB - Human checkpoint kinase 1 (Chk1) is an essential kinase required to preserve genome stability. Here, we show that Chk1 inhibition by two distinct drugs, UCN-01 and CEP-3891, or by Chk1 small interfering RNA (siRNA) leads to phosphorylation of ATR targets. Chk1-inhibition triggered rapid, pan-nuclear phosphorylation of histone H2AX, p53, Smc1, replication protein A, and Chk1 itself in human S-phase cells. These phosphorylations were inhibited by ATR siRNA and caffeine, but they occurred independently of ATM. Chk1 inhibition also caused an increased initiation of DNA replication, which was accompanied by increased amounts of nonextractable RPA protein, formation of single-stranded DNA, and induction of DNA strand breaks. Moreover, these responses were prevented by siRNA-mediated downregulation of Cdk2 or the replication initiation protein Cdc45, or by addition of the CDK inhibitor roscovitine. We propose that Chk1 is required during normal S phase to avoid aberrantly increased initiation of DNA replication, thereby protecting against DNA breakage. These results may help explain why Chk1 is an essential kinase and should be taken into account when drugs to inhibit this kinase are considered for use in cancer treatment.

U2 - 10.1128/MCB.25.9.3553-3562.2005

DO - 10.1128/MCB.25.9.3553-3562.2005

M3 - Journal article

C2 - 15831461

SN - 0270-7306

VL - 25

SP - 3553

EP - 3562

JO - Molecular and Cellular Biology

JF - Molecular and Cellular Biology

IS - 9

ER -