TGF-beta-induced matrix proteins inhibit p42/44 MAPK and JNK activation and suppress TNF-mediated IkappaBalpha degradation and NF-kappaB nuclear translocation in L929 fibroblasts.
The role of transforming growth factor beta1 (TGF-beta1)-induced extracellular matrix proteins in the modulation of cellular response to the cytotoxic effect of tumor necrosis factor (TNF) or Fas ligand was investigated. Murine L929 fibroblasts were prestimulated with or without TGF-beta1 for 1-24 h and the resulting extracellular protein matrices were prepared. Unstimulated control L929 cells were then cultured on these matrices. Compared to control matrix-stimulated L929 cells, the TGF-beta1 matrix-stimulated cells resisted TNF killing in the presence of actinomycin D (ActD), but became more susceptible to killing by anti-Fas antibodies/ActD. The induced TNF resistance is independent of the NF-kappaB antiapoptotic effect. For example, exposure of TGF-beta1 matrix-stimulated L929 cells to TNF failed to result in IkappaBalpha degradation and NF-kappaB nuclear translocation or activation. Also, control matrix stimulated the activation of p42/44 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) in L929 cells, whereas TGF-beta1 matrix suppressed the activation. Nonetheless, in response to TNF, JNK activation was restored in the TGF-beta1 matrix-stimulated cells. By metabolic labeling, ammonium sulfate precipitation and N-terminal amino acid microsequencing, TGF-beta1 was shown to induce a novel matrix protein of 46 kDa (p46) from L929 cells. Adsorption of p46 by peptide antibodies against its N-terminus removed the TGF-beta1 matrix protein-mediated protection against TNF/ActD cytotoxicity and its enhancement of anti-Fas/ActD killing, indicating that p46 is responsible for these effects. Immunostaining of L929 cells revealed that the antibodies were bound to a membrane protein of 100 kDa (p100). Thus, the matrix p46 is likely derived from the released membrane p100.
Chang
N S
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Guthrie Research Institute, Guthrie Medical Center, 1 Guthrie Square, Sayre, Pennsylvania, 18840, USA. nschang@inet.guthrie.org
D000595:Amino Acid Sequence; D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003609:Dactinomycin / Q000494:pharmacology; D016326:Extracellular Matrix Proteins / Q000737:chemistry / Q000378:metabolism*; D020672:I-kappa B Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D007700:Kinetics; D007739:L Cells; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors*; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008969:Molecular Sequence Data; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D019014:fas Receptor / Q000187:drug effects / Q000502:physiology
D004268:DNA-Binding Proteins; D016326:Extracellular Matrix Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606205:Nfkbia protein, mouse; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; D000072000:NF-KappaB Inhibitor alpha; D003609:Dactinomycin; D011505:Protein-Tyrosine Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases