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Appropriate aliquots of cell extracts were used for measurements of luciferase and β-galactosidase activity, as recommended by Promega.To correct for transfection efficiency, light units from the luciferase assay were divided by the OD reading from the β-galactosidase assay. Sites that are hypersensitive to digestion can also be seen between these footprinted regions (Fig. Other footprints further upstream of FP6 were observed in similar experiments, but these were not defined further because maximal activity of the human ).

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These promoter deletion constructs were transiently transfected into Hep G2 cells that produce SHBG (20).

The results indicate that footprints 1–4 are required for transcriptional activity in these cells and that the region upstream of nt ).

The SLC2A9 genotypes also showed significant association with self-reported gout.

The proportion of the variance of serum uric acid concentrations explained by genotypes was about 1.2% in men and 6% in women, and the percentage accounted for by expression levels was 3.5% in men and 15% in women.

Testicular androgen-binding protein has the same primary structure and steroid-binding properties as plasma SHBG and is thought to control androgen-dependent sperm maturation events in the male reproductive tract (1, 9).

In rodents, the testis appears to be the major site of expression in adult animals, but the gene is also expressed transiently in the fetal rat liver, and this may serve to influence the differentiation of reproductive tissues during late fetal development (10).It is produced by hepatocytes, and blood levels of SHBG in humans are influenced by a variety of hormones, as well as several nutritional and metabolic regulators, including sex steroids (2), thyroid hormone (2, 3), insulin (4), dietary lipids (5), isoflavanoid phytoestrogens (6), and body weight (7).Low plasma levels of SHBG are associated with several sex steroid hormone-dependent diseases (2) and have been reported to be an early indicator of the onset of Type II diabetes (8) and cardiovascular disease (7).A Maxam Gilbert sequencing (G/A) reaction was used as a size marker ( sequences −299/−137, whereas footprints 1–4 were removed individually by further 5′-deletions.The 3′-end of these deletion mutants was maintained at nt 7 relative to the transcription start site (Fig.3).In experiments where promoter-luciferase reporter gene constructs were co-transfected into Hep G2 cells with COUP-TF and/or HNF-4 expression vectors, HNF-4 markedly increased transcription, whereas COUP-TF suppressed this probably by displacing HNF-4 from their common FP1-binding site.