Formation of cobalt protoporphyrin in the liver of rats. A mechanism for the inhibition of liver haem biosynthesis by inorganic cobalt

Sinclair, Peter R.; Gibbs, Anthony H.; Sinclair, Jacqueline F.; Matteis, Federico De

doi:10.1042/bj1780529

Cited by 64 publications

(35 citation statements)

References 40 publications

(29 reference statements)

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“…In contrast to the results under oxidative and nitrosative stress conditions, the transcriptional activities of both the glbN and glbO promoters increased under transition metal-induced hypoxia. It has been observed that transition metals, like nickel or cobalt, induce a number of hypoxic genes under normoxic conditions (5,21,53) by serving as substrates for ferrochelatase and being incorporated into protoporphyrin IX in place of iron (48). Nickel-substituted hemoproteins, therefore, do not bind oxygen and cannot function as oxygen sensors.…”

Section: Discussionmentioning

confidence: 99%

Responses ofMycobacterium tuberculosisHemoglobin Promoters to In Vitro and In Vivo Growth Conditions

Pawaria

Lama

Raje

et al. 2008

Appl Environ Microbiol

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The success of Mycobacterium tuberculosis as one of the dreaded human pathogens lies in its ability to utilize different defense mechanisms in response to the varied environmental challenges during the course of its intracellular infection, latency, and reactivation cycle. Truncated hemoglobins trHbN and trHbO are thought to play pivotal roles in the cellular metabolism of this organism during stress and hypoxia. To delineate the genetic regulation of the M. tuberculosis hemoglobins, transcriptional fusions of the promoters of the glbN and glbO genes with green fluorescent protein were constructed, and their responses were monitored in Mycobacterium smegmatis and M. tuberculosis H37Ra exposed to environmental stresses in vitro and in M. tuberculosis H37Ra after in vivo growth inside macrophages. The glbN promoter activity increased substantially during stationary phase and was nearly 3-to 3.5-fold higher than the activity of the glbO promoter, which remained more or less constant during different growth phases in M. smegmatis, as well as in M. tuberculosis H37Ra. In both mycobacterial hosts, the glbN promoter activity was induced 1.5-to 2-fold by the general nitrosative stress inducer, nitrite, as well as the NO releaser, sodium nitroprusside (SNP). The glbO promoter was more responsive to nitrite than to SNP, although the overall increase in its activity was much less than that of the glbN promoter. Additionally, the glbN promoter remained insensitive to the oxidative stress generated by H 2 O 2 , but the glbO promoter activity increased nearly 1.5-fold under similar conditions, suggesting that the trHb gene promoters are regulated differently under nitrosative and oxidative stress conditions. In contrast, transition metal-induced hypoxia enhanced the activity of both the glbN and glbO promoters at all growth phases; the glbO promoter was induced ϳ2.3-fold, which was found to be the highest value for this promoter under all the conditions evaluated. Addition of iron along with nickel reversed the induction in both cases. Interestingly, a concentration-dependent decrease in the activity of both trHb gene promoters was observed when the levels of iron in the growth media were depleted by addition of an iron chelator. These results suggested that an iron/heme-containing oxygen sensor is involved in the modulation of the trHb gene promoter activities directly or indirectly in conjunction with other cellular factors. The modes of promoter regulation under different physiological conditions were found to be similar for the trHbs in both M. smegmatis and M. tuberculosis H37Ra, indicating that the promoters might be regulated by components that are common to the two systems. Confocal microscopy of THP-1 macrophages infected with M. tuberculosis carrying the trHb gene promoter fusions showed that there was a significant level of promoter activity during intracellular growth in macrophages. Time course evaluation of the promoter activity after various times up to 48 h by fluorescence-activated cell sorting analysis of th...

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Section: Discussionmentioning

confidence: 99%

Responses ofMycobacterium tuberculosisHemoglobin Promoters to In Vitro and In Vivo Growth Conditions

Pawaria

Lama

Raje

et al. 2008

Appl Environ Microbiol

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“…Cobalt has been shown to be a substrate for ferrochelatase, the enzyme responsible for the incorporation of iron into protoporphyrin IX to make heme (Labbe and Hubbard, 1961). Radiolabeling studies both in vivo (Sinclair et al, 1979) and in cultured cells (Sinclair et al, 1982) have demonstrated the incorporation of transition metals including cobalt and nickel into heme. If the effect of cobalt and nickel depends on incorporation into the heme moiety, increased levels of iron should competitively inhibit the stimulatory effects of these metals on Epo gene expression.…”

Section: Evidence That the Sensor Is A Heme Proteinmentioning

confidence: 99%

Oxygen sensing and signaling: impact on the regulation of physiologically important genes

Zhu

Bunn

1999

Respiration Physiology

151

101

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A growing number of physiologically relevant genes are regulated in response to changes in intracellular oxygen tension. It is likely that cells from a wide variety of tissues share a common mechanism of oxygen sensing and signal transduction leading to the activation of the transcription factor hypoxia-inducible factor 1 (HIF-1). Besides hypoxia, transition metals (Co 2+ , Ni 2+ and Mn 2+ ) and iron chelation also promote activation of HIF-1. Induction of HIF-1 by hypoxia is blocked by the heme ligands carbon monoxide and nitric oxide. There is growing, albeit indirect, evidence that the oxygen sensor is a flavoheme protein and that the signal transduction pathway involves changes in the level of intracellular reactive oxygen intermediates. The activation of HIF-1 by hypoxia depends upon signaling-dependent rescue of its α-subunit from oxygendependent degradation in the proteasome, allowing it to form a heterodimer with HIF-1β (ARNT), which then translocates to the nucleus and impacts on the transcription of genes whose cis-acting elements contain cognate hypoxia response elements.

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“…No direct evidence for the latter theory could be found however, in spite of the well-known ability of ferrochelatase (EC 4.99.1 .l), the metal ion inserting enzyme of haem biosynthesis, to insert a variety of metal ions into a number of porphyrin co-substrates [4]. Recently cobalt protophorphyrin has been isolated from rats injected with large doses of cobalt chloride [5], rat liver homogenates have also been shown to produce this metalloporphyrin [6]. Isolated hepatocytes in culture, which are increasingly used to study hepatic function, have so far been reported unable to produce cobalt protoporphyrin when supplied with the metal ion [7,8].…”

Section: Introductionmentioning

confidence: 99%

Effects of cobalt chloride on haem synthesis in isolated hepatocytes

Lodola

1981

FEBS Letters

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Administration of cobalt to ratsleads to the inhibition of amino-laevulinate synthetase (EC 2.3.1.37) and the induction of haem oxygenase (EC 1.14.99.3) synthesis [ 1,2]. These effects have been ascribed to a direct action by the metal ion [l] or the formation of a cobalt porphyrin [3]. No direct evidence for the latter theory could be found however, in spite of the well-known ability of ferrochelatase (EC 4.99.1 .l), the metal ion inserting enzyme of haem biosynthesis, to insert a variety of metal ions into a number of porphyrin co-substrates [4]. Recently cobalt protophorphyrin has been isolated from rats injected with large doses of cobalt chloride [5], rat liver homogenates have also been shown to produce this metalloporphyrin [6]. Isolated hepatocytes in culture, which are increasingly used to study hepatic function, have so far been reported unable to produce cobalt protoporphyrin when supplied with the metal ion [7,8]. In view of the findings in [_5,6] the effects of this metal ion on hepatocyte cultures was re-examined.In the presence of cobalt chloride it was found that there was a decreased rate of haem synthesis from exogenously supplied amino-laevulinate (ALA), a specific haem precursor [9]. Under these conditions the metalloporphyrin concentration associated with the cell fraction increased relative to the control. A metalloporphyrin was extracted from cells cultured in the presence of ALA and CoCIZ which had spectral properties consistent with its being cobalt protoporphyrin. Methods and materials Isolation of hepatocytesHepatocytes were isolated as in [lo] modified to exclude the post-perfusion incubation step. The iso- ElsevierlNorth-Holland Biomedical Presslated cells were washed (3 X 40 ml) in Waymouth MB 752/l tissue culture medium supplemented with 10% foetal calf serum, penicillin (100 units/ml), streptomycin (100 mcg/ml), mycostatin (50 units/ml) and Hepes buffer (final cont. 2.5 mM) at pH 7.3. After the final wash the cells were resuspended in 40-50 ml of this medium. Hepatocyte cultureThe cell suspension was diluted to 3.5 X lo6 cells/ inl, with tissue culture medium, and 1.5 ml of this suspension seeded into one 9.5 cm2 well of a Costar 6 well cluster dish and incubated at 37°C.After 4 h fresh medium was added. Experimental additions were made either at this point (for 24 h incubations) or after 18 h at the next media change (O-10 h time courses). Haem determinationsReaction was stopped by harvesting the cells in: (a) the reaction medium; or (b) after removal of the medium into 0.9% (w/v) NaCl. The total haem concentration was determined as in [I 11. Metalloporphyrin extractionAfter 24 h incubation the cells and media from 10 cluster dishes were harvested, combined and the metalloporphyrin extracted as in [5].2.5. Spectroscopy Spectra were obtained using a chopped split-beam spectrophotometer (Applied Photophysics Ltd., London) interfaced to an Apple ITT 2020 Microprossesor used for baseline corrections and spectral manipulations. Protein assaysProtein concentrations were determined as ...

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Formation of cobalt protoporphyrin in the liver of rats. A mechanism for the inhibition of liver haem biosynthesis by inorganic cobalt

Cited by 64 publications

References 40 publications

Responses ofMycobacterium tuberculosisHemoglobin Promoters to In Vitro and In Vivo Growth Conditions

Responses ofMycobacterium tuberculosisHemoglobin Promoters to In Vitro and In Vivo Growth Conditions

Oxygen sensing and signaling: impact on the regulation of physiologically important genes

Effects of cobalt chloride on haem synthesis in isolated hepatocytes

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