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 RIS Download
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