Institute for Protein Research, Osaka University, Japan.
We studied the transcriptional cis-acting elements of the myeloperoxidase gene, which is expressed during the promyelocyte stage of granulocyte development by assay of transient expression of the chloramphenicol acetyltransferase (CAT) gene in myeloid leukemia SKM-1 cells and analysis of the DNA binding sites for HL-60 nuclear factors. Assay of CAT expression dependent on restriction fragments isolated from genomic clones indicated that the fragments located on introns 7 and 9 enhanced the expression. Methylation interference experiments showed that the guanine residues in a consensus sequence of an estrogen response element in the intron 7 fragment interacted with a nuclear factor. Gel retardation analysis indicated that this interaction of the intron 7 fragment with the nuclear factor was specifically inhibited by an oligodeoxynucleotide containing the 21-base pair (bp) estrogen response element. DNase I footprint analysis revealed that a 36-bp-specific sequence of the intron 9 fragment was protected from DNase I by nuclear extracts. This sequence contained a palindromic sequence consisting of the conserved half-motif of an estrogen response element with 5-bp spacing. The interaction of the intron 9 fragment with the nuclear extracts was specifically inhibited by an oligodeoxynucleotide of the 36-bp sequence. Furthermore, the 21- and 36-bp oligodeoxynucleotides in the constructs enhanced CAT expression in the cells. These results suggest that these elements in introns 7 and 9 are involved in expression of the myeloperoxidase gene.