Mitotic DNA damage response: Polo-like kinase-1 is dephosphorylated through ATM-Chk1 pathway.
DNA damage during the cell division cycle can activate ATM/ATR and their downstream kinases that are involved in the checkpoint pathway, and cell growth is halted until damage is repaired. As a result of DNA damage induced in mitotic cells by doxorubicin treatment, cells accumulate in a G2-like phase, not in mitosis. Under these conditions, two mitosis-specific kinases, Cdk1 and Plk1, are inhibited by inhibitory phosphorylation and dephosphorylation, respectively. G2-specific phosphorylation of Cdc25 was increased during incubation after mitotic DNA damage. Inhibition of Plk1 through dephosphorylation was dependent on ATM/Chk1 activity. Depleted expression of ATM and Chk1 was achieved using small hairpin RNA (shRNA) plasmid constructs. In this condition, damaged mitotic cells did not accumulated in a G2-like stage, and entered into G1 phase without delay. Protein phosphatase 2A was responsible for dephosphorylation of mitotic Plk1 in response to DNA damage. In knockdown of PP2A catalytic subunits, Plk1 was not dephosphorylated, but rather degraded in response to DNA damage, and cells did not accumulate in G2-like phase. The effect of ATM/Chk1 inhibition was counteracted by overexpression of PP2A, indicated that PP2A may function as a downstream target of ATM/Chk1 at a mitotic DNA damage checkpoint, or may have a dominant effect on ATM/Chk1 function at this checkpoint. Finally, we have shown that negative regulation of Plk1 by dephosphorylation is important to cell accumulation in G2-like phase at the mitotic DNA damage checkpoint, and that this ATM/Chk1/PP2A pathway independent on p53 is a novel mechanism of cellular response to mitotic DNA damage.
Lee
Hee-Jae
HJ
Laboratory of Cell Cycle & Signal Transduction, World Class University Research Department of Nanobiomedical Science, and Institute of Tissue Regeneration Engineering, Dankook University, Cheonan, Korea.
Hwang
Hyo-In
HI
Jang
Young-Joo
YJ
D064007:Ataxia Telangiectasia Mutated Proteins; D002453:Cell Cycle / Q000187:drug effects; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D000071877:Checkpoint Kinase 1; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004317:Doxorubicin / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000187:drug effects; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D054648:Protein Phosphatase 2 / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D034741:RNA, Small Interfering; D015398:Signal Transduction; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D020687:cdc25 Phosphatases / Q000378:metabolism; D000097869:Polo-Like Kinase 1
D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D000071877:Checkpoint Kinase 1; D004268:DNA-Binding Proteins; D004317:Doxorubicin; D011494:Protein Kinases; D054648:Protein Phosphatase 2; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D020687:cdc25 Phosphatases; D000097869:Polo-Like Kinase 1; C576324:ATM protein, human; C000605867:CHEK1 protein, human