Cdk2-dependent phosphorylation of p21 regulates the role of Cdk2 in cisplatin cytotoxicity

Am J Physiol Renal Physiol. 2011 May;300(5):F1171-9. doi: 10.1152/ajprenal.00507.2010. Epub 2011 Feb 16.

Abstract

Cisplatin cytotoxicity is dependent on cyclin-dependent kinase 2 (Cdk2) activity in vivo and in vitro. We found that an 18-kDa protein identified by mass spectrometry as p21(WAF1/Cip1) was phosphorylated by Cdk2 starting 12 h after cisplatin exposure. The analysis showed it was phosphorylated at serine 78, a site not previously identified. The adenoviral transduction of p21 before cisplatin exposure protects from cytotoxicity by inhibiting Cdk2. Although cisplatin causes induction of endogenous p21, the protection is inefficient. We hypothesized that phosphorylation of p21 at serine 78 could affect its role as a Cdk inhibitor, and thereby lessen its ability to protect from cisplatin cytotoxicity. To investigate the effect of serine 78 phosphorylation on p21 activity, we replaced serine 78 with aspartic acid, creating the phosphomimic p21(S78D). Mutant p21(S78D) was an inefficient inhibitor of Cdk2 and was inefficient at protecting TKPTS cells from cisplatin-induced cell death. We conclude that phosphorylation of p21 by Cdk2 limits the effectiveness of p21 to inhibit Cdk2, which is the mechanism for continued cisplatin cytotoxicity even after the induction of a protective protein.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Acute Kidney Injury / chemically induced*
  • Acute Kidney Injury / enzymology
  • Acute Kidney Injury / pathology
  • Adenoviridae / genetics
  • Amino Acid Sequence
  • Animals
  • Antineoplastic Agents / toxicity*
  • Cisplatin / toxicity*
  • Cyclin A / genetics
  • Cyclin A / metabolism
  • Cyclin-Dependent Kinase 2 / genetics
  • Cyclin-Dependent Kinase 2 / metabolism*
  • Cyclin-Dependent Kinase Inhibitor p21 / genetics
  • Cyclin-Dependent Kinase Inhibitor p21 / metabolism*
  • Disease Models, Animal
  • Genetic Vectors
  • HEK293 Cells
  • Humans
  • Kidney Tubules, Proximal / drug effects*
  • Kidney Tubules, Proximal / enzymology
  • Kidney Tubules, Proximal / pathology
  • Mice
  • Mice, 129 Strain
  • Molecular Sequence Data
  • Mutation
  • Phosphorylation
  • Recombinant Fusion Proteins / metabolism
  • Serine
  • Tandem Mass Spectrometry
  • Time Factors
  • Transduction, Genetic
  • Transfection

Substances

  • Antineoplastic Agents
  • Cdkn1a protein, mouse
  • Cyclin A
  • Cyclin-Dependent Kinase Inhibitor p21
  • Recombinant Fusion Proteins
  • Serine
  • CDK2 protein, human
  • Cdk2 protein, mouse
  • Cyclin-Dependent Kinase 2
  • Cisplatin