Glial-specific cAMP response of the glial fibrillary acidic protein gene cell lines.
Expression of the rat glial fibrillary acidic protein (GFAP) gene is responsive to the intracellular level of cAMP. We have examined the sequence 5'-upstream of the transcription start site of the rat GFAP-encoding gene to determine the elements responsible for regulating the cAMP response. The RT4 cell lines consist of a neural stem-cell type RT4-AC and its three derivative cell types, one glial-cell type, RT4-D, and two neuronal-cell types, RT4-B and RT4-E. GFAP is expressed in the stem-cell type and the glial-cell type but is not expressed in the neuronal-cell types. Luciferase expression vectors containing various areas of the 10.8-kb region upstream of the transcription start site of the GFAP gene were transiently transfected into these RT4 cells. The effect of cAMP was examined by quantitating the transient expression of luciferase. We found that (i) the 5'-upstream region alone (up to 10.8 kb) allows expression of the GFAP gene in the stem-cell type, the glial-cell type, and a neuronal-cell type; (ii) there are negative and positive cAMP-responsive elements that are juxtaposed within the region between -240 bp and -110 bp upstream and are functional in the stem-cell and glial-cell types but are not functional in the neuronal-cell type RT4-E; (iii) there may be elements that respond to dibutyryl-cAMP in all three RT4 cell types within the region from 2 kb to 10.8 kb upstream of the transcription start site; and (iv) a regulatory luciferase plasmid pRLgfap-1, containing both the upstream and downstream regulatory regions of the GFAP gene, not only expresses luciferase but also responds to forskolin in the stem-cell type and the glial-cell type. This regulatory plasmid, however, does not express in the neuronal-cell type with or without the forskolin treatment.
Kaneko
R
R
Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder 80309-0347.
Hagiwara
N
N
Leader
K
K
Sueoka
N
N
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D005822:Genetic Vectors; D005904:Glial Fibrillary Acidic Protein / Q000096:biosynthesis* / Q000235:genetics*; D008156:Luciferases / Q000096:biosynthesis / Q000378:metabolism; D008969:Molecular Sequence Data; D009457:Neuroglia / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism; D010957:Plasmids; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D013234:Stem Cells / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection
D005904:Glial Fibrillary Acidic Protein; D005576:Colforsin; D004247:DNA; D000242:Cyclic AMP; D008156:Luciferases