Positive and negative regulation of EAAT2 by NF-kappaB: a role for N-myc in TNFalpha-controlled repression.
The glutamate transporter gene, EAAT2/GLT-1, is induced by epidermal growth factor (EGF) and downregulated by tumor necrosis factor alpha (TNFalpha). While TNFalpha is generally recognized as a positive regulator of NF-kappaB-dependent gene expression, its ability to control transcriptional repression is not well characterized. Additionally, the regulation of NF-kappaB by EGF is poorly understood. Herein, we demonstrate that both TNFalpha-mediated repression and EGF-mediated activation of EAAT2 expression require NF-kappaB. We show that EGF activates NF-kappaB independently of signaling to IkappaB. Furthermore, TNFalpha can abrogate IKKbeta- and p65-mediated activation of EAAT2. Our results suggest that NF-kappaB can intrinsically activate EAAT2 and that TNFalpha mediates repression through a distinct pathway also requiring NF-kappaB. Consistently, we find that N-myc is recruited to the EAAT2 promoter with TNFalpha and that N-myc-binding sites are required for TNFalpha-mediated repression. Moreover, N-myc overexpression inhibits both basal and p65-induced activation of EAAT2. Our data highlight the remarkable specificity of NF-kappaB activity to regulate gene expression in response to diverse cellular signals and have implications for glutamate homeostasis and neurodegenerative disease.
Sitcheran
Raquel
R
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, NC 27599-7295, USA.
Gupta
Pankaj
P
Fisher
Paul B
PB
Baldwin
Albert S
AS
D027322:Amino Acid Transport System X-AG / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology; D027342:Excitatory Amino Acid Transporter 2; D005786:Gene Expression Regulation / Q000187:drug effects; D050491:Glutamate Plasma Membrane Transport Proteins; D006801:Humans; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D027981:Symporters / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology
D027322:Amino Acid Transport System X-AG; D027342:Excitatory Amino Acid Transporter 2; D050491:Glutamate Plasma Membrane Transport Proteins; D016328:NF-kappa B; D016271:Proto-Oncogene Proteins c-myc; D011994:Recombinant Proteins; C493397:SLC1A2 protein, human; D027981:Symporters; D014409:Tumor Necrosis Factor-alpha; D004815:Epidermal Growth Factor; D004247:DNA