The glutamate transporter gene, EAAT2/GLT-1, is induced by epidermal growth factor (EGF) and downregulated by tumor necrosis factor α (TNFα). While TNFα is generally recognized as a positive regulator of NF-κB-dependent gene expression, its ability to control transcriptional repression is not well characterized. Additionally, the regulation of NF-κB by EGF is poorly understood. Herein, we demonstrate that both TNFα-mediated repression and EGF-mediated activation of EAAT2 expression require NF-κB. We show that EGF activates NF-κB independently of signaling to IκB. Furthermore, TNFα can abrogate IKKβ- and p65-mediated activation of EAAT2. Our results suggest that NF-κB can intrinsically activate EAAT2 and that TNFα mediates repression through a distinct pathway also requiring NF-κB. Consistently, we find that N-myc is recruited to the EAAT2 promoter with TNFα and that N-myc-binding sites are required for TNFα-mediated repression. Moreover, N-myc overexpression inhibits both basal and p65-induced activation of EAAT2. Our data highlight the remarkable specificity of NF-κB activity to regulate gene expression in response to diverse cellular signals and have implications for glutamate homeostasis and neurodegenerative disease.