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Cytokine activation of p38 mitogen-activated protein kinase and apoptosis is opposed by alpha-4 targeting of protein phosphatase 2A for site-specific dephosphorylation of MEK3.

alpha-4 is an essential gene and is a dominant antiapoptotic factor in various tissues that is a regulatory subunit for type 2A protein phosphatases. A multiplexed phosphorylation site screen revealed that knockdown of alpha-4 by small interfering RNA (siRNA) increased p38 mitogen-activated protein kinase (MAPK) and c-Jun phosphorylation without changes in JNK or ERK. FLAG-alpha-4 coprecipitated hemagglutinin-MEK3 plus endogenous protein phosphatase 2A (PP2A) and selectively enhanced dephosphorylation of Thr193, but not Ser189, in the activation loop of MEK3. Overexpression of alpha-4 suppressed p38 MAPK activation in response to tumor necrosis factor alpha (TNF-alpha). The alpha-4 dominant-negative domain (DND) (residues 220 to 340) associated with MEK3, but not PP2A, and its overexpression sensitized cells to activation of p38 MAPK by TNF-alpha and interleukin-1beta, but not by ansiomycin or sorbitol. The response was diminished by nocodazole or by siRNA knockdown of the Opitz syndrome protein Mid1 that binds alpha-4 to microtubules. Interference by alpha-4 DND or alpha-4 siRNA increased caspase 3/7 activation in response to TNF-alpha. Growth of transformed cells in soft agar was enhanced by alpha-4 and suppressed by alpha-4 DND. The results show that alpha-4 targets PP2A activity to MEK3 to suppress p38 MAPK activation by cytokines, thereby inhibiting apoptosis and anoikis.

Pubmed ID: 17438131

Authors

  • Prickett TD
  • Brautigan DL

Journal

Molecular and cellular biology

Publication Data

June 28, 2007

Associated Grants

  • Agency: NCI NIH HHS, Id: CA77584

Mesh Terms

  • Actinin
  • Animals
  • Apoptosis
  • COS Cells
  • Caspase 3
  • Caspase 7
  • Cell Line
  • Cercopithecus aethiops
  • Dose-Response Relationship, Drug
  • Enzyme Activation
  • Gene Targeting
  • Genes, Essential
  • HeLa Cells
  • Hemagglutinins
  • Humans
  • Interleukin-1beta
  • JNK Mitogen-Activated Protein Kinases
  • MAP Kinase Kinase 3
  • Microfilament Proteins
  • Microtubule Proteins
  • Mitogen-Activated Protein Kinase 14
  • Nocodazole
  • Nuclear Proteins
  • Oligopeptides
  • Peptides
  • Phosphoprotein Phosphatases
  • Phosphorylation
  • Promoter Regions, Genetic
  • Protein Phosphatase 2
  • Protein Structure, Tertiary
  • RNA, Small Interfering
  • Threonine
  • Transcription Factors
  • Tumor Necrosis Factor-alpha