Structural organization of the human heme oxygenase gene and the function of its promoter

Abstract

Human heme oxygenase is induced by its substrate heme, but not induced by heat shock [Yoshida et al. (1988) Eur. J . Biochem. 1 7 / , 457-4611. In order to study the molecular mechanisms of heme-mediated induction of human hcmc oxygenase, we have isolated and characterized the genomic clones for heme oxygenase. The human heme oxygenase gene (HO gene) is about 14 kb long and organized into five exons. The transcription initiation site was identified by S1 nuclease mapping and primer-extension analyses. Using HeLa whole cell extracts, we confirmed that the transcription of the cloned HO gene is initiated accurately at the assigned initiation site. In its 5'-flanking region, a potential heat-shock element (HSE) is present 367 bp upstream from the initiation site, although, in contrast to rat heme oxygenase, human enzyme is not induced by heat shock. We therefore analyzed the effects of heat shock on the transient expression of chimeric fusion genes harboring the promoter of the human HO gene ligated to the Escherichia coli gene gpt in mouse amelanotic melanoma cells. The 5'-flanking region of the human HO gene bearing a potential HSE failed to confer the heat-inducibility of gpt RNA production, suggesting that the HSE of the human HO gene is not functional.Heme oxygenase, an essential enzyme in heme catabolism, cleaves heme to form biliverdin [l], which is subsequently converted to bilirubin by biliverdin reductase in mammals [2]. The activity of heme oxygenase is highest in the spleen, where senescent erythrocytes (nearly 1 YO of erythrocytes in man) are sequestrated and destroyed every day. Heme oxygenase also plays an important role in determining the availability of cellular heme for various hemoproteins, such as cytochrome P-450, catalase and tryptophan pyrrolase [3]. It is therefore of particular significance that the activity of heme oxygenase is increased by its substrate heme [4, 51 as well as by various non-heme substances [3]. The increased activity of heme oxygenase may result in the depletion of cellular free heme and cause functional disturbances of such hemoproteins (reviewed in [6]).Recently, we have cloned and characterized the rat heme oxygenase gene (HO gene) [7]. Its 5'-flanking region contains a potential heat-shock element (HSE), which was originally found in the 5'-flanking regions of heat-shock protein genes and conferred the transcriptional induction of these genes by environmental derangements (reviewed in [S, 91). We thus found that rat heme oxygenase is a heat-shock protein and hemin induces heme oxygenase by a different mechanism from heat shock [lo]. In contrast to rat enzyme, human heme oxygenase is not induced by heat shock, suggesting that human enzyme is not a heat-shock protein, although hemin increases both mRNA levels and activity of heme oxygenase in human macrophages and glioma cells [I 11.In order to study the regulation of its expression, we have cloned and characterized the human HO gene. In this report, we show the structural organization of the gene and present the nucl...

Interaction of upstream stimulatory factor with the human heme oxygenase gene promoter

“…The cis-acting element of the rat heme oxygenase gene (-48 to -37; Fig. 1B) is almost identical to the nucleotide residues (-45 to -34) of the human heme oxygenase gene [12], suggesting that the same nuclear protein, possibly USF, may interact with this element of the human heme oxygenase gene. In order to investigate whether USF is indeed involved in the expression of the human heme oxygenase gene, we partially purified USF from HeLa cells, which was shown to bind to the upstream promoter sequence of Ad2MLP ( Fig.…”

“…The products were identified by S1 nuclease mapping analysis. The S1 probe was the PstI/HinfI fragment (nucleotide residues -283 to intron 1) end-labelled at the HinfI site, which is located 221 bp downstream from the transcription initiation site [12].…”

“…The subcloned plasmid, harbouring the 5'-flanking region of about 1.4 kb, exons 1, 2, and 3, and a part of intron 3 of the human heme oxygenase gene [12], was transcribed using HeLa cell lysates according to the instructions of the supplier. The products were identified by S1 nuclease mapping analysis.…”

Study of GT-repeat expansion in Heme oxygenase-1 gene promoter as genetic cause of male infertility

“…Pair of primers was obtained from SHIBAHARA et al 1989 to amplify a 137 to 200 bp fragment, depending on the number of repeats, containing GT-n repeats region from promoter of HO-1 gene by PCR. The 5′ end of forward primer was labeled with FAM 5′-FAMAGAGCCTGCAGCTTCTCAGA-3′ for using in analysis of DNA fragmentation by ABI capillary genotyping and an unlabeled antisense primer 5′-ACAAAGTCTG GCCATAGGAC-3′, which were designed on the basis of the published sequence [24]. The PCR cycle of 94°C for 45 s, 57°C for 30 s and 72°C for 30 s was carried out for a total of 32 cycles.…”

Care for Older People

“…Weinvestigated genes to determine what kind of people tend to have emphysema. There is an enzyme called "hemoxygenase-1", which all cells in our body possess to knock-off active oxygen from smoking or other factors (78). Those who have the type L of hemoxygenase-1 gene are apt to comedownto emphysemathree times as often as those whodo not have (79).…”