Phosphorylation of TPL-2 on serine 400 is essential for lipopolysaccharide activation of extracellular signal-regulated kinase in macrophages.
Tumor progression locus 2 (TPL-2) kinase is essential for Toll-like receptor 4 activation of the mitogen-activated protein kinase extracellular signal-regulated kinase (ERK) and for upregulation of the inflammatory cytokine tumor necrosis factor (TNF) in lipopolysaccharide (LPS)-stimulated macrophages. LPS activation of ERK requires TPL-2 release from associated NF-kappaB1 p105, which blocks TPL-2 access to its substrate, the ERK kinase MEK. Here we demonstrate that TPL-2 activity is also regulated independently of p105, since LPS stimulation was still needed for TPL-2-dependent activation of ERK in Nfkb1(-/-) macrophages. In wild-type macrophages, LPS induced the rapid phosphorylation of serine (S) 400 in the TPL-2 C-terminal tail. Mutation of this conserved residue to alanine (A) blocked the ability of retrovirally expressed TPL-2 to induce the activation of ERK in LPS-stimulated Nfkb1(-/-) macrophages. TPL-2(S400A) expression also failed to reconstitute LPS activation of ERK and induction of TNF in Map3k8(-/-) macrophages, which lack endogenous TPL-2. Consistently, the S400A mutation was found to block LPS stimulation of TPL-2 MEK kinase activity. Thus, induction of TPL-2 MEK kinase activity by LPS stimulation of macrophages requires TPL-2 phosphorylation on S400, in addition to its release from NF-kappaB1 p105. Oncogenic C-terminal truncations of TPL-2 that remove S400 could promote its transforming potential by eliminating this critical control step.
Robinson
M J
MJ
Division of Immune Cell Biology, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, United Kingdom.
Beinke
S
S
Kouroumalis
A
A
Tsichlis
P N
PN
Ley
S C
SC
D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis / Q000187:drug effects; D002460:Cell Line; D051766:Early Growth Response Protein 1 / Q000235:genetics; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D020930:MAP Kinase Kinase Kinases / Q000737:chemistry / Q000378:metabolism*; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects* / Q000201:enzymology*; D051379:Mice; D008969:Molecular Sequence Data; D052002:NF-kappa B p50 Subunit / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D048069:Tumor Necrosis Factors / Q000235:genetics; D015854:Up-Regulation / Q000187:drug effects
D051766:Early Growth Response Protein 1; D008070:Lipopolysaccharides; D052002:NF-kappa B p50 Subunit; D011518:Proto-Oncogene Proteins; D048069:Tumor Necrosis Factors; D012694:Serine; D051057:Proto-Oncogene Proteins c-akt; D051550:I-kappa B Kinase; D048049:Extracellular Signal-Regulated MAP Kinases; D020930:MAP Kinase Kinase Kinases; C489843:Map3k8 protein, mouse
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