Pds1 phosphorylation in response to DNA damage is essential for its DNA damage checkpoint function.
In Saccharomyces cerevisiae, Pds1 is an anaphase inhibitor and plays an essential role in DNA damage and spindle checkpoint pathways. Pds1 is phosphorylated in response to DNA damage but not spindle disruption, indicating distinct mechanisms delaying anaphase entry. Phosphorylation of Pds1 is Mec1 and Chk1 dependent in vivo. Here, we show that Pds1 is phosphorylated at multiple sites in vivo in response to DNA damage by Chk1. Mutation of the Chk1 phosphorylation sites on Pds1 abolished most of its DNA damage-inducible phosphorylation and its checkpoint function, whereas its anaphase inhibitor functions and spindle checkpoint functions remain intact. Loss of Pds1 phosphorylation correlates with APC-dependent Pds1 destruction in response to DNA damage. We also show that APC(Cdc20) is active in preanaphase arrested cells after DNA damage. This suggests that Pds1 is stabilized by phosphorylation in response to DNA damage, but APC(Cdc20) activity is not altered. Our results indicate that phosphorylation of Pds1 by Chk1 is the key function of Chk1 required to prevent anaphase entry.
Wang
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Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
Liu
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Wang
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Y
Qin
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Elledge
S J
SJ
D000595:Amino Acid Sequence; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D000071877:Checkpoint Kinase 1; D004249:DNA Damage / Q000235:genetics* / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D018816:Genes, cdc; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D064249:Securin
C577495:CDC20 protein, S cerevisiae; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins; D005656:Fungal Proteins; D009687:Nuclear Proteins; C578004:PDS1 protein, S cerevisiae; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins; D064249:Securin; D011494:Protein Kinases; D000071877:Checkpoint Kinase 1