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

The CCAAT/Enhancer Binding Protein (C/EBP) family belongs to the basic leucine zipper class of transcription factors with a total of 6 members having been identified in this family so far. This gene family has been mainly implicated in regulation of cell specific gene expression and differentiation. In order to identify factors important for the regulation of the mouse C/EBPa gene the promoter region was characterised using nuclear extracts prepared from liver. Several sites within the promoter which bound nuclear proteins were identified. One of these sites was identified 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, suggesting a role for these proteins in regulating C/EBP a expression. The expression of C/EBPa and C/EBPb was studied in brown adipose tissue. Dramatic changes of gene expression during cold exposure were observed. The C/EBPa expression was transiently downregulated whereas C/EBPb expression was strongly induced.

Also norepinephrine affected the expression of both genes and it was concluded that both genes are under andrenergic control in this tissue. Following up the studies of the C/EBPa promoter, the role of Myc and Max in regulation of the C/EBPb gene was investigated. It was determined that overexpression of Myc strongly represses expression of the C/EBPa gene, and also blocks differentiation of the brown adipocyte 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. Both genes 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 and mouse genes, but the promoter region was less conserved. By using Northern blot analysis on several human tissues, the C/EBPa gene was found to have a tissue restricted expression pattern. Sequence determination of the C/EBPe gene revealed that it shared homology to the rat gene termed CRPl. Its chromosomal localisation was determined both with fluorescent in situ hybridisation (FISH) and by linkage analysis to chromosome 14qll.2, between the TCRA and CTLA genes. This chromosomal region is frequently rearranged in T-cell leukaemias. Expression in normal human tissues was detected in peripheral blood mononuclear cells, 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. These results show that the C/EBPe gene has a very cell specific expression pattern and that it has an intriguing chromosome location.