Tpl2 (tumor progression locus 2) phosphorylation at Thr290 is induced by lipopolysaccharide via an Ikappa-B Kinase-beta-dependent pathway and is required for Tpl2 activation by external signals.
The serine-threonine protein kinase encoded by the tumor progression locus 2 (Tpl2) proto-oncogene transduces Toll-like receptor and death receptor signals in a variety of cell types. Here we show that Tpl2 undergoes phosphorylation at Thr(290) both in cells overexpressing Tpl2 and in cells stimulated with lipopolysaccharide (LPS) or tumor necrosis factor-alpha and that phosphorylation on this site parallels Tpl2 activation. Reconstitution of Tpl2(-/-) macrophages with wild type Tpl2 or Tpl2 T290D restored ERK activation by LPS, whereas reconstitution of the same cells with Tpl2 T290A did not, suggesting that phosphorylation at Thr(290) is required for the physiological activation of Tpl2 by external signals. Both the wild type Tpl2 and the kinase-inactive mutant Tpl2 K167M undergo Thr(290) phosphorylation, suggesting that Thr(290) may be a site of trans-phosphorylation rather than auto-phosphorylation. Pretreatment of 293 cells and primary macrophages with the Ikappa-B kinase-beta (IKKbeta) inhibitor PS-1145 blocked Tpl2 phosphorylation at Thr(290), suggesting that phosphorylation depends on IKKbeta, an obligatory positive regulator of Tpl2. We conclude that Tpl2 phosphorylation at Thr(290) is induced by LPS, depends on IKKbeta, and is required for the physiological activation of Tpl2 by external signals.
Cho
Jeonghee
J
Molecular Oncology Research Institute, Tufts-New England Medical Center and Tufts Cancer Center, Boston, Massachusetts 02111, USA.
Melnick
Michael
M
Solidakis
Georgios P
GP
Tsichlis
Philip N
PN
D000818:Animals; D001665:Binding Sites; D001854:Bone Marrow Cells; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006575:Heterocyclic Compounds, 3-Ring / Q000494:pharmacology; D051550:I-kappa B Kinase; D007163:Immunosorbent Techniques; D008070:Lipopolysaccharides / Q000494:pharmacology*; D020930:MAP Kinase Kinase Kinases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D011725:Pyridines / Q000494:pharmacology; D034741:RNA, Small Interfering; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D013912:Threonine / Q000378:metabolism*; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D004791:Enzyme Inhibitors; D006575:Heterocyclic Compounds, 3-Ring; D008070:Lipopolysaccharides; C456319:PS1145; D011518:Proto-Oncogene Proteins; D011725:Pyridines; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D013912:Threonine; D017346:Protein Serine-Threonine Kinases; C496564:Chuk protein, mouse; D051550:I-kappa B Kinase; C496562:Ikbkb protein, mouse; C496566:Ikbke protein, mouse; D048049:Extracellular Signal-Regulated MAP Kinases; D020930:MAP Kinase Kinase Kinases; C489843:Map3k8 protein, mouse