Trans-activators regulating neuronal glucose transporter isoform-3 gene expression in mammalian neurons.
The murine facilitative glucose transporter isoform 3 is developmentally regulated and is predominantly expressed in neurons. By employing the primer extension assay, the transcription start site of the murine Glut 3 gene in the brain was localized to -305 bp 5' to the ATG translation start codon. Transient transfection assays in N2A neuroblasts using murine GLUT3-luciferase reporter constructs mapped enhancer activities to two regions located at -203 to -177 and -104 to -29 bp flanking a previously described repressor element (-137 to -130 bp). Dephosphorylated Sp1 and Sp3 proteins from the 1- and 21-day-old mouse brain nuclear extracts bound the repressor elements, whereas both dephosphorylated and phosphorylated cAMP-response element-binding protein (CREB) in N2A, 1- and 21-day-old mouse brain nuclear extracts bound the 5'-enhancer cis-elements (-187 to -180 bp) of the Glut 3 gene, and the Y box protein MSY-1 bound the sense strand of the -83- to -69-bp region. Sp3, CREB, and MSY-1 binding to the GLUT 3 DNA was confirmed by the chromatin immunoprecipitation assay, whereas CREB and MSY-1 interaction was detected by the co-immunoprecipitation assay. Furthermore, small interference RNA targeted at CREB in N2A cells decreased endogenous CREB concentrations, and CREB mediated GLUT 3 transcription. Thus, in the murine brain similar to the N2A cells, phosphorylated CREB and MSY-1 bound the Glut 3 gene trans-activating the expression in neurons, whereas Sp1/Sp3 bound the repressor elements. We speculate that phosphorylated CREB and Sp3 also interacted to bring about GLUT 3 expression in response to development/cell differentiation and neurotransmission.
Rajakumar
Augustine
A
Division of Neonatology and Developmental Biology, Department of Pediatrics, David Geffen School of Medicine, UCLA, Los Angeles, California 90095-1752, USA.
Thamotharan
Shanthie
S
Raychaudhuri
Nupur
N
Menon
Ram K
RK
Devaskar
Sherin U
SU
D000818:Animals; D001921:Brain / Q000196:embryology / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D018387:Codon, Initiator; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D017931:DNA Primers / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D017353:Gene Deletion; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D051274:Glucose Transporter Type 3; D051379:Mice; D008807:Mice, Inbred BALB C; D009004:Monosaccharide Transport Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D009419:Nerve Tissue Proteins; D009474:Neurons / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012313:RNA / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000378:metabolism; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection
D002843:Chromatin; D018387:Codon, Initiator; D017362:Cyclic AMP Response Element-Binding Protein; D017931:DNA Primers; D004268:DNA-Binding Proteins; D051274:Glucose Transporter Type 3; D009004:Monosaccharide Transport Proteins; D009419:Nerve Tissue Proteins; D034741:RNA, Small Interfering; C495689:Slc2a3 protein, mouse; D016329:Sp1 Transcription Factor; C497122:Sp3 protein, mouse; D014157:Transcription Factors; D051705:Sp3 Transcription Factor; D012313:RNA