Mice lacking protein phosphatase 5 are defective in ataxia telangiectasia mutated (ATM)-mediated cell cycle arrest.
Protein phosphatase 5 (Ppp5), a tetratricopeptide repeat domain protein, has been implicated in multiple cellular functions, including cellular proliferation, migration, differentiation and survival, and cell cycle checkpoint regulation via the ataxia telangiectasia mutated/ATM and Rad3-related (ATM/ATR) signal pathway. However, the physiological functions of Ppp5 have not been reported. To confirm the role of Ppp5 in cell cycle checkpoint regulation, we generated Ppp5-deficient mice and isolated mouse embryonic fibroblast (MEF) cells from Ppp5-deficient and littermate control embryos. Although Ppp5-deficient mice can survive through embryonic development and postnatal life and MEF cells from the Ppp5-deficient mice maintain normal replication checkpoint induced by hydroxyurea, Ppp5-deficient MEF cells display a significant defect in G(2)/M DNA damage checkpoint in response to ionizing radiation (IR). To determine whether this defect in IR-induced G(2)/M checkpoint is due to altered ATM-mediated signaling, we measured ATM kinase activity and ATM-mediated downstream events. Our data demonstrated that IR-induced ATM kinase activity is attenuated in Ppp5-deficient MEFs. Phosphorylation levels of two known ATM substrates, Rad17 and Chk2, were significantly reduced in Ppp5-deficient MEFs in response to IR. Furthermore, DNA damage-induced Rad17 nuclear foci were dramatically reduced in Ppp5-deficient MEFs. These results demonstrate a direct regulatory linkage between Ppp5 and activation of the ATM-mediated G(2)/M DNA damage checkpoint pathway in vivo.
Yong
Weidong
W
Herman B Wells Center for Pediatric Research, Section of Pediatric Cardiology, Department of Pediatrics, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
Bao
Shideng
S
Chen
Hanying
H
Li
Dapei
D
Sánchez
Edwin R
ER
Shou
Weinian
W
D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D002478:Cells, Cultured; D064447:Checkpoint Kinase 2; D004249:DNA Damage / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism*; D047108:Embryonic Development / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D016195:G2 Phase / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D005720:Gamma Rays; D006918:Hydroxyurea / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D009687:Nuclear Proteins / Q000172:deficiency / Q000378:metabolism*; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000172:deficiency / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects / Q000528:radiation effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D025521:Tumor Suppressor Proteins / Q000378:metabolism*
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D025521:Tumor Suppressor Proteins; D064447:Checkpoint Kinase 2; D064007:Ataxia Telangiectasia Mutated Proteins; C576325:Atm protein, mouse; C578633:Chek2 protein, mouse; D017346:Protein Serine-Threonine Kinases; D010749:Phosphoprotein Phosphatases; C089641:protein phosphatase 5; D006918:Hydroxyurea