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Regulation of heme oxygenase-1 gene expression in vascular smooth muscle cells by nitric oxide
C L Hartsfield1, J Alam, J L Cook
1Division of Pulmonary and Critical Care Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland 21224, USA.
Insights
Nitric oxide (NO) upregulates heme oxygenase-1 (HO-1) expression in vascular cells. This NO-mediated HO-1 induction involves new protein synthesis and is independent of cyclic GMP signaling, revealing a key NO-CO regulatory link.
Area of Science:
- Biochemistry
- Molecular Biology
- Physiology
Background:
- Heme oxygenase (HO) catalyzes heme degradation, producing carbon monoxide (CO).
- Carbon monoxide (CO) and nitric oxide (NO) are gaseous signaling molecules with overlapping physiological functions.
- The relationship between NO and HO-mediated CO production suggests potential regulatory interactions.
Purpose of the Study:
- To investigate whether nitric oxide (NO) regulates heme oxygenase (HO) expression.
- To elucidate the molecular mechanisms underlying NO-induced HO-1 expression.
Main Methods:
- Treatment of aortic vascular smooth muscle cells (aSMC) with NO donor spermine NONOate (SNN).
- Analysis of HO-1 mRNA and protein levels.
- Assessment of gene transcription and mRNA turnover rates.
- Inhibition studies using cycloheximide, cGMP analogs, guanylate cyclase inhibitors, and antioxidants.
Main Results:
- Spermine NONOate (SNN) significantly increased HO-1 mRNA and protein in aSMC.
- HO-1 induction resulted from increased gene transcription and decreased mRNA degradation.
- NO-induced HO-1 expression requires new protein synthesis and is independent of cyclic GMP (cGMP).
- Antioxidant N-acetyl-L-cysteine inhibited NO-induced HO-1 expression, while peroxynitrite did not.
Conclusions:
- Nitric oxide (NO) positively regulates heme oxygenase-1 (HO-1) expression in vascular smooth muscle cells.
- This regulation involves transcriptional and post-transcriptional mechanisms.
- The findings highlight a novel regulatory network between NO and CO signaling pathways.
Abstract:
Heme oxygenase (HO)-mediated heme degradation is the primary mechanism for production of cellular carbon monoxide (CO). Analogous to nitric oxide (NO), CO mediates physiological and cellular functions such as vasodilation, stimulation of guanylate cyclase, and neuronal transmission. In view of accumulating data demonstrating a correlation between the activity of these two gaseous molecules and that the predominant source of CO is via HO catalysis, we hypothesized that NO regulates HO expression. We demonstrate that the NO donor spermine NONOate (SNN) increases steady-state levels of HO-1 mRNA in aortic vascular smooth muscle cells (aSMC) in both a time- and dose-dependent manner. The accumulation of HO-1 mRNA that correlated with increased HO-1 protein synthesis resulted from both an increased rate of gene transcription and a decreased rate of mRNA turnover. Inhibition of the NO-induced HO-1 mRNA expression by cycloheximide suggests that new protein synthesis is required for increased HO-1 gene expression. Induction of HO-1 expression by SNN occurs in a guanosine 3',5'-cyclic monophosphate (cGMP)-independent manner because exposure of cells to 8-bromoguanosine 3',5'-cyclic monophosphate, a cGMP analog, did not increase HO-1 mRNA levels, and pretreatment of cells with 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, a selective guanylate cyclase inhibitor, did not prevent SNN-induced HO-1 mRNA accumulation. The antioxidant N-acetyl-L-cysteine markedly inhibited SNN-induced HO-1 mRNA expression, whereas peroxynitrite did not induce HO-1 expression in aSMC. Interestingly, CO did not attenuate NO-induced HO-1 expression through an autocrine negative feedback mechanism as had been observed for hypoxia-induced HO-1 expression. These data provide evidence for an important regulatory network between NO and CO via HO-1.