ANALYSIS OF THE HUMAN FERROCHELATASE PROMOTER IN TRANSGENIC MICE

Ferrochelatase catalyzes the chelation of ferrous iron and protoporphy rin to form heme. It is expressed as a housekeeping gene in all cells, but is upregulated during erythropoiesis, Ferrochelatase activity is deficient in the inherited disease protoporphyria as a result of heter ogeneous mutations. Although human ferrochelatase is transcribed from a single promoter in both nonerythroid and erythroid cells, previous s tudies using transient transfection assays failed to demonstrate eryth roid-specific increased expression from 4.0 kb of the human ferrochela tase promoter containing the erythroid cis-elements, GATA and NF-E2. T he present study analyzes the in vivo regulation of the ferrochelatase gene to provide insights into the mechanism of its erythroid-specific enhancement. Transgenic (TG) mouse lines were generated in which the luciferase reporter gene was driven by either a 150-bp ferrochelatase minimal promoter (-0.15 To) or by a 4.0 kb extended 5' upstream region (-4.0 TG), Expression of the -4.0 TG transgene was generally consiste nt with the endogenous gene during embryonic development and in nonery throid and erythroid tissues as demonstrated by Northern blotting and mRNA in situ hybridization, The -4.0 TG was expressed at a higher leve l than the -0.15 TG in nonerythroid and erythroid tissues, including d uring extramedullary erythropoiesis induced by n-acetylphenylhydrazine injection. The enhanced erythroid expression of the -4,0 TG correlate s with the appearance of a DNase I hypersensitive site in the 5' flank ing region of the transgene, Therefore, in the context of chromosomal integration, the 5' flanking region of the ferrochelatase gene is nece ssary and sufficient to confer high levels of transgene expression in erythroid tissue. (C) 1998 by The American Society of Hematology.