DNA damage triggers p21WAF1-dependent Emi1 down-regulation that maintains G2 arrest.
Several regulatory proteins control cell cycle progression. These include Emi1, an anaphase-promoting complex (APC) inhibitor whose destruction controls progression through mitosis to G1, and p21(WAF1), a cyclin-dependent kinase (CDK) inhibitor activated by DNA damage. We have analyzed the role of p21(WAF1) in G2-M phase checkpoint control and in prevention of polyploidy after DNA damage. After DNA damage, p21(+/+) cells stably arrest in G2, whereas p21(-/-) cells ultimately progress into mitosis. We report that p21 down-regulates Emi1 in cells arrested in G2 by DNA damage. This down-regulation contributes to APC activation and results in the degradation of key mitotic proteins including cyclins A2 and B1 in p21(+/+) cells. Inactivation of APC in irradiated p21(+/+) cells can overcome the G2 arrest. siRNA-mediated Emi1 down-regulation prevents irradiated p21(-/-) cells from entering mitosis, whereas concomitant down-regulation of APC activity counteracts this effect. Our results demonstrate that Emi1 down-regulation and APC activation leads to stable p21-dependent G2 arrest after DNA damage. This is the first demonstration that Emi1 regulation plays a role in the G2 DNA damage checkpoint. Further, our work identifies a new p21-dependent mechanism to maintain G2 arrest after DNA damage.
Lee
Jinho
J
Institut de Biologie Structurale J-P Ebel, F38027 Grenoble, France.
Kim
Jin Ah
JA
Barbier
Valerie
V
Fotedar
Arun
A
Fotedar
Rati
R
D064173:Anaphase-Promoting Complex-Cyclosome; D015703:Antigens, CD; D015820:Cadherins / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism*; D004249:DNA Damage; D015536:Down-Regulation / Q000235:genetics* / Q000528:radiation effects; D004789:Enzyme Activation / Q000528:radiation effects; D044783:F-Box Proteins / Q000235:genetics* / Q000378:metabolism; D016195:G2 Phase / Q000528:radiation effects; D005720:Gamma Rays; D045325:HCT116 Cells; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism
D015703:Antigens, CD; C513384:CDH1 protein, human; D015820:Cadherins; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D044783:F-Box Proteins; C498960:FBXO5 protein, human; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome