Studies of the C/EBP family of transcription factors : cloning, regulation, and expression

Abstract: The CCAAT/Enhancer Binding Protein (C/EBP) family belongs to the basic leucine zipperclass of transcription factors with a total of 6 members having been identified in this familyso far. This gene family has been mainly implicated in regulation of cell specific geneexpression and differentiation.In order to identify factors important for the regulation of the mouse C/EBPa gene thepromoter region was characterised using nuclear extracts prepared from liver. Several siteswithin the promoter which bound nuclear proteins were identified. One of these sites wasidentified as a C/EBP binding site and it was further determined that the gene is autoregulated.Upstream of this site an E-box motif was identified which binds the transcription factor USF.It was also demonstrated that purified Myc and Max proteins can bind to this site, suggestinga role for these proteins in regulating C/EBPa expression.The expression of C/EBPa and C/EBPb was studied in brown adipose tissue. Dramaticchanges of gene expression during cold exposure were observed. The C/EBPa expressionwas transiently downregulated whereas C/EBPb expression was strongly induced. Alsonorepinephrine affected the expression of both genes and it was concluded that both genes areunder andrenergic control in this tissue.Following up the studies of the C/EBPa promoter, the role of Myc and Max in regulation ofthe C/EBP¶ gene was investigated. It was determined that overexpression of Myc stronglyrepresses expression of the C/EBPa gene, and also blocks differentiation of the brownadipocyte cell line HIB-lB. The Myc repression however was not mediated via the Mycbinding site, but through the core promoter region.The last two studies describe the cloning of the human C/EBPa and C/EBPe genes. Bothgenes were sequenced and the sequences were submitted to the EMBL/GenBank databases.The coding region of the C/EBPa gene was seen to have very high similarity to the rat andmouse genes, but the promoter region was less conserved. By using Northern blot analysis onseveral human tissues, the C/EBPa gene was found to have a tissue restricted expressionpattern. Sequence determination of the C/EBPe gene revealed that it shared homology to therat gene termed CRPl . Its chromosomal localisation was determined both with fluorescent insitu hybridisation (FISH) and by linkage analysis to chromosome 14qll.2, between theTCRA and CTLA genes. This chromosomal region is frequently rearranged in T-cellleukaemias . Expression in normal human tissues was detected in peripheral blood mononuclearcells, ovaries and in bone marrow. Expression was also found in the human cell lines Jurkatand HL60. This was the first time expression of this C/EBP isoform has been reported. Theseresults show that the C/EBPe gene has a very cell specific expression pattern and that it hasan intriguing chromosome location.Key words: Basic leucine zipper, CCAAT/Enhancer Binding Protein, C/EBP, DNA binding protein,Gene expression, Transcription factorISBN 91-628-2038-9