Mechanism of melphalan-induced B7-1 gene expression in P815 tumor cells.
We have previously shown that exposure of P815 tumor cells to melphalan (L-phenylalanine mustard; L-PAM) leads to up-regulation of B7-1 surface expression, and this L-PAM-induced up-regulation requires de novo RNA synthesis and is associated with accumulation of B7-1 mRNA. Here we show that the effect of L-PAM on B7-1 surface expression can be mimicked by exposing P815 tumor cells to oxidative stress but not to heat shock. Moreover, the antioxidant N-acetyl-L-cysteine prevented the L-PAM-induced accumulation of B7-1 mRNA in P815 tumor cells, suggesting that reactive oxygen species are involved in the transcriptional regulation of L-PAM-induced B7-1 gene expression. Although AP-1 and NF-kappaB are regarded as redox-sensitive transcription factors and the promoter/enhancer region of the B7-1 gene contains an AP-1 and an NF-kappaB binding site, exposure of P815 tumor cells to L-PAM led to rapid and transient activation only of NF-kappaB, but not AP-1, that bound specifically to a probe containing the respective binding site in the murine or human B7-1 gene. Moreover, exposure of P815 tumor cells to a cell-permeable peptide that selectively inhibits NF-kappaB activation by blocking the activation of the IkappaB-kinase complex was found to inhibit the L-PAM-induced B7-1 mRNA accumulation, indicating that NF-kappaB activation is essential for the L-PAM-induced B7-1 gene expression. Taken together, these results indicate that L-PAM leads to activation of B7-1 gene expression by activating NF-kappaB via a pathway that involves reactive oxygen species.
Donepudi
M
M
Department of Biochemistry and Molecular Biology, University of Illinois, Chicago, IL 60612.
Raychaudhuri
P
P
Bluestone
J A
JA
Mokyr
M B
MB
D000111:Acetylcysteine / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis; D018906:Antineoplastic Agents, Alkylating / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000975:Antioxidants / Q000494:pharmacology; D018122:B7-1 Antigen / Q000096:biosynthesis* / Q000235:genetics*; D051940:B7-2 Antigen; D001667:Binding, Competitive; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D002463:Cell Membrane Permeability; D002467:Cell Nucleus / Q000737:chemistry; D004742:Enhancer Elements, Genetic / Q000187:drug effects / Q000276:immunology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000276:immunology*; D006358:Hot Temperature; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D051550:I-kappa B Kinase; D046911:Macromolecular Substances; D012515:Mast-Cell Sarcoma / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008558:Melphalan / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D008562:Membrane Glycoproteins / Q000096:biosynthesis; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000276:immunology; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism; D015345:Oligonucleotide Probes / Q000378:metabolism; D010455:Peptides / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000276:immunology; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000187:drug effects / Q000235:genetics / Q000276:immunology
D015703:Antigens, CD; D018906:Antineoplastic Agents, Alkylating; D000975:Antioxidants; D018122:B7-1 Antigen; D051940:B7-2 Antigen; C497964:CD86 protein, human; C497965:Cd86 protein, mouse; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D015345:Oligonucleotide Probes; D010455:Peptides; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D006861:Hydrogen Peroxide; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; C496564:Chuk protein, mouse; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human; C496562:Ikbkb protein, mouse; C496566:Ikbke protein, mouse; D008558:Melphalan; D000111:Acetylcysteine