Functional Analysis of the Cell-Specific Enhancer in the Human Proopiomelanocortin Gene by β-Galactosidase Histochemical Staining

摘要

ABSTRACTNucleotide sequences responsible for the cell-specific expression of the human proopiomelanocortin (POMC) gene were analyzed by histochemical staining of β-galactosidase in culture cells transfected with chimeric genes containing the 5′-flanking regions of the human POMC gene fused to theEscherichia coli lacZgene. The chimeric genes were stably introduced into various culture cells, including AtT-20 cells, which express the endogenous mouse POMC gene. Whereas the control gene containing the cytomegalovirus enhancer was expressed in all cell lines tested, only AtT-20 cells supported the efficient transcription of the gene containing 2.9 kb of the human POMC 5′-flanking region. These results indicate that the stable transfection-expression system utilizing the histochemical detection of the gene expression is a useful method for the analysis of cell-specific gene expression. These results have also confirmed that thetrans-acting factors in mouse AtT-20 cells interact with the human POMC gene promoter region and activate the transcription of the gene. Deletion analysis has demonstrated that the profiles of the transcriptional activity of the various human POMC–lacZfusion genes are similar to those of the rat POMC gene described previously. Comparison of the human and the rat 5′-flanking sequences revealed close homology in several regions, which might be involved in the efficient transcription of the POMC gene in AtT-