Microarray Analysis Supports a Role for CCAAT/Enhancer-binding Protein-β in Brain Injury

Abstract

CCAAT/enhancer-binding protein-β (C/EBPβ) is a transcription factor that plays an important role in regulating cell growth and differentiation. This protein plays a central role in lymphocyte and adipocyte differentiation and hepatic regeneration and in the control of inflammation and immunity in the liver and in cells of the myelomonocytic lineage. Our previous studies suggested that this protein could also have important functions in the brain. Therefore, we were interested in the identification of downstream targets of this transcription factor in cells of neural origin. We performed cDNA microarray analysis and found that a total of 48 genes were up-regulated in C/EBPβ-overexpressing neuronal cells. Of the genes that displayed significant changes in expression, several were involved in inflammatory processes and brain injury. Northern blot analysis confirmed the up-regulation of ornithine decarboxylase, 24p3/LCN2, GRO1/KC, spermidine/spermine N1-acetyltransferase, xanthine dehydrogenase, histidine decarboxylase, decorin, and TM4SF1/L6. Using promoter-luciferase reporter transfection assays, we showed the ornithine decarboxylase and 24p3 genes to be biological downstream targets of C/EBPβ in neuroblastoma cells. Moreover, the levels of C/EBPβ protein were significantly induced after neuronal injury, which was accompanied by increased levels of cyclooxygenase-2 enzyme. This strongly supports the concept that C/EBPβ may play an important role in brain injury. CCAAT/enhancer-binding protein-β (C/EBPβ) is a transcription factor that plays an important role in regulating cell growth and differentiation. This protein plays a central role in lymphocyte and adipocyte differentiation and hepatic regeneration and in the control of inflammation and immunity in the liver and in cells of the myelomonocytic lineage. Our previous studies suggested that this protein could also have important functions in the brain. Therefore, we were interested in the identification of downstream targets of this transcription factor in cells of neural origin. We performed cDNA microarray analysis and found that a total of 48 genes were up-regulated in C/EBPβ-overexpressing neuronal cells. Of the genes that displayed significant changes in expression, several were involved in inflammatory processes and brain injury. Northern blot analysis confirmed the up-regulation of ornithine decarboxylase, 24p3/LCN2, GRO1/KC, spermidine/spermine N1-acetyltransferase, xanthine dehydrogenase, histidine decarboxylase, decorin, and TM4SF1/L6. Using promoter-luciferase reporter transfection assays, we showed the ornithine decarboxylase and 24p3 genes to be biological downstream targets of C/EBPβ in neuroblastoma cells. Moreover, the levels of C/EBPβ protein were significantly induced after neuronal injury, which was accompanied by increased levels of cyclooxygenase-2 enzyme. This strongly supports the concept that C/EBPβ may play an important role in brain injury. CCAAT/enhancer-binding proteins (C/EBPs) 1The abbreviations used are: C/EBP, CCAAT/enhancer-binding protein; IL, interleukin; MOPS, 4-morpholinepropanesulfonic acid; ODC, ornithine decarboxylase; COX, cyclooxygenase. 1The abbreviations used are: C/EBP, CCAAT/enhancer-binding protein; IL, interleukin; MOPS, 4-morpholinepropanesulfonic acid; ODC, ornithine decarboxylase; COX, cyclooxygenase. are a family of transcription factors that belong to the (bZIP) (basic leucine zipper) class and are known to couple extracellular signal transduction pathways to numerous cellular processes. C/EBPs have been involved in the regulation of various aspects of cell differentiation and function in multiple tissues. Six different members of the family (C/EBPα to C/EBPζ) that give rise to different isoforms corresponding to full-length and amino-terminally truncated proteins have been isolated (1Calkhoven C.F. Muller C. Leutz A. Genes Dev. 2000; 14: 1920-1932Google Scholar) and found to play a role in growth arrest and cell differentia