Liver-specific DNase I-hypersensitive sites and DNA methylation pattern in the promoter region of a 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene

Abstract

The mRNA for the liver isozyme of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase is transcribed from the L promoter of gene A. We show here that L-promoter activity is tissue specific. To identify on the gene in situ potential cis-acting sequences, we have examined 15 kb of its 5' region for DNase I-hypersensitive sites detectable on chromatin. We have also evaluated the DNA methylation status of the 3.7-kb encompassing the L promoter. Five DNase I-hypersensitive sites were detected on liver chromatin, three upstream (M1 at position -4500, L2 at position -1000, L1 at position -200) and two downstream (I1 at position +3000, I2 at position +3500) from the L-type mRNA transcription initiation site. Their presence correlated with transcriptional activity as they were not observed on chromatin from kidney, a tissue where gene A is not expressed. Sites M1 and L1 corresponded to the M and L promoters, respectively, providing in vivo evidence for a promoter localization obtained earlier with cloned DNA only. Site I2 coincided with a glucocorticoid-responsive unit described by others, but its presence did not depend on glucocorticoids. Thus, sites L2 and I1 could correspond to novel control elements. While DNA was methylated around position -2000 both in liver and kidney, downstream from that position it was fully demethylated in liver but not in kidney. This pattern changed during development of fetal liver. The data suggest mechanisms for the lack of activity of the L promoter in kidney and for its activation in developing and adult liver.