title: Nuclear factor-κB contributes to neuron-dependent induction of glutamate transporter-1 expression in astrocytes.
abstract: The glutamate transporter-1 [GLT-1 (excitatory amino acid transporter 2)] subtype of glutamate transporter ensures crisp excitatory signaling and limits excitotoxicity in the CNS. Astrocytic expression of GLT-1 is regulated during development, by neuronal activity, and in neurodegenerative diseases. Although neurons activate astrocytic expression of GLT-1, the mechanisms involved have not been identified. In the present study, astrocytes from transgenic mice that express enhanced green fluorescent protein (eGFP) under the control of a bacterial artificial chromosome (BAC) containing a very large region of DNA surrounding the GLT-1 gene (BAC GLT-1 eGFP mice) were used to assess the role of nuclear factor-κB (NF-κB) in neuron-dependent activation of the GLT-1 promoter. We provide evidence that neurons activate NF-κB signaling in astrocytes. Transduction of astrocytes from the BAC GLT-1 eGFP mice with dominant-negative inhibitors of NF-κB signaling completely blocked neuron-dependent activation of a NF-κB reporter construct and attenuated induction of eGFP. Exogenous expression of p65 and/or p50 NF-κB subunits induced expression of eGFP or GLT-1 and increased GLT-1-mediated transport activity. Using wild-type and mutant GLT-1 promoter reporter constructs, we found that NF-κB sites at -583 or -251 relative to the transcription start site were required for neuron-dependent reporter activation. Electrophoretic mobility shift and supershift assays reveal that p65 and p50 interact with these same sites ex vivo. Finally, chromatin immunoprecipitation showed that p65 and p50 interact with these sites in adult cortex, but not in kidney (a tissue that expresses no detectable GLT-1). Together, these studies strongly suggest that NF-κB contributes to neuron-dependent regulation of astrocytic GLT-1 transcription.
authors.affiliation: Department of Pediatrics, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania 19104-4318, USA.
authors.forename: Mausam
authors.initials: M
authors.lastname: Ghosh
authors.forename: Yongjie
authors.initials: Y
authors.lastname: Yang
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Rothstein
authors.forename: Michael B
authors.initials: MB
authors.lastname: Robinson
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D002450:Cell Communication / Q000502:physiology*; D002478:Cells, Cultured; D027342:Excitatory Amino Acid Transporter 2 / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D027342:Excitatory Amino Acid Transporter 2; D016328:NF-kappa B