Regulation of CCAAT/enhancer-binding protein family members by stimulation of glutamate receptors in cultured rat cortical astrocytes.
Regulation of mRNA levels, DNA binding activities, and phosphorylation of CCAAT/enhancer-binding protein (C/EBP) family members by stimulation of glutamate receptors were studied in cultured rat cortical astrocytes. Indirect immunofluorescence and immunoblot analyses with specific antibodies to C/EBP family members revealed that both C/EBPbeta and C/EBPdelta but not C/EBPalpha are expressed in the nuclei of astrocytes. After exposure to glutamate, C/EBPbeta mRNA levels increased within 10 min, reached the maximal level at about 1 h, and returned to the basal level within 6 h. In contrast, C/EBPdelta mRNA levels decreased by 6 h and were recovered within 12 h. These changes in mRNA levels were accompanied by an increase and a decrease in proteins for C/EBPbeta and C/EBPdelta, respectively. Elevation of C/EBPbeta mRNA levels by glutamate treatment required an increase in intracellular Ca2+ concentration and depended on activations of protein kinase C and calmodulin-dependent protein kinases. Gel mobility shift analysis using nuclear extracts from the glutamate-treated cells showed increases in C/EBP site binding activities 2 h after the exposure to glutamate. Moreover, glutamate stimulated phosphorylation of C/EBPbeta in 32P-labeled astrocytes in a Ca2+-dependent manner. These results suggest that glutamate regulates functions of C/EBP family members in brain astrocytes through changes in mRNA levels of C/EBPbeta and C/EBPdelta as well as through phosphorylation of C/EBPbeta.
Yano
S
S
Department of Pharmacology, Kumamoto University School of Medicine, Kumamoto 860, Japan.
Fukunaga
K
K
Takiguchi
M
M
Ushio
Y
Y
Mori
M
M
Miyamoto
E
E
D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D001667:Binding, Competitive; D022762:CCAAT-Enhancer-Binding Proteins; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D018698:Glutamic Acid / Q000494:pharmacology; D005810:Multigene Family; D009687:Nuclear Proteins / Q000096:biosynthesis* / Q000235:genetics; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D011494:Protein Kinases / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017470:Receptors, Glutamate / Q000378:metabolism*; D015398:Signal Transduction
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D017470:Receptors, Glutamate; D018698:Glutamic Acid; D011494:Protein Kinases