Sepiapterin reductase regulation of endothelial tetrahydrobiopterin and nitric oxide bioavailability

Ling Gao, Yuh Fen Pung, Jun Zhang, Peng Chen, Ting Wang, Min Li, Miguel Meza, Ligia Toro, Hua Cai

Research output: Contribution to journalArticle

25 Scopus citations

Abstract

Sepiapterin reductase (SPR) catalyzes the final step of tetrahydrobiopterin (H4B) biosynthesis and the first step of H4B regeneration from an exogenous precursor sepiapterin. Despite the potential significance of SPR in regulating H4B-dependent nitric oxide (NO) production, the endothelium-specific sequence and functions of SPR remain elusive. We first cloned endothelial SPR cDNA from bovine aortic endothelial cells (Genebank: DQ978331). In cells transiently transfected with SPR gene, SPR activity (HPLC) was dramatically increased by 19-fold, corresponding to a significant increase in endothelial H4B content (HPLC) and NO production (electron spin resonance). In vivo delivery of SPR gene significantly increased vascular SPR protein expression (mouse vs. bovine antibodies to differentiate endogenous vs. exogenous), activity, H4B content, and NO production, as well as NO-dependent vasorelaxation. In endothelial cells transfected with small interfering RNA specific for SPR, ∼87% of mRNA were attenuated (real-time quantitative RT-PCR), corresponding to a significant reduction in SPR protein expression and activity, which was associated with decreases in both intracellular H 4B content and NO level. Exogenous administration of sepiapterin to endothelial cells significantly upregulated H4B and NO levels, which were attenuated by SPR RNA interference (RNAi). H4B-stimulated increase in NO production, however, was SPR RNAi independent. GTP cyclohydrolase 1 expression and activity, as well as dihydrofolate reductase expression, were not affected by SPR RNAi, whereas dihydrofolate reductase activity was significantly downregulated. These data represent the first to study endothelial SPR functionally and clearly demonstrate an important role of endothelial SPR in modulating H4B and NO bioavailability.

Original languageEnglish (US)
Pages (from-to)H331-H339
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume297
Issue number1
DOIs
StatePublished - Jul 2009

Keywords

  • Aortic endothelial cells
  • Dihydrofolate reductase
  • Endothelial nitric oxide synthase
  • Guanosine 5′-triphosphate cyclohydrolase 1

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

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