Abstract
The endogenous bronchodilator, S-nitrosoglutathione (GSNO), increases expression, maturation, and function of both the wild-type and the ΔF508 mutant of the cystic fibrosis transmembrane conductance regulatory protein (CFTR). Though transcriptional mechanisms of action have been identified, GSNO seems also to have post-transcriptional effects on CFTR maturation. Here, we report that 1) GSNO is only one of a class of S-nitrosylating agents that, at low micromolar concentrations, increase ΔF508 and wild-type CFTR expression and maturation; 2) NO itself (at these concentrations) and 8-bromocyclic GMP are minimally active on CFTR; 3) a novel agent, S-nitrosoglutathione diethyl ester, bypasses the need for GSNO bioactivation by γ-glutamyl transpeptidase to increase CFTR maturation; 4) surprisingly, expression—but not S-nitrosylation—of cysteine string proteins (Csp) 1 and 2 is increased by GSNO; 5) the effect of GSNO to increase full maturation of wild-type CFTR is inhibited by Csp silencing (si)RNA; 6) proteins relevant to CFTR trafficking are SNO-modified, and SNO proteins traffic through the endoplasmic reticulum (ER) and Golgi after GSNO exposure; and 7) GSNO alters the interactions of ΔF508 CFTR with Csp and Hsc70 in the ER and Golgi. These data suggest that GSNO is one of a class of S-nitrosylating agents that act independently of the classic NO radical/cyclic GMP pathway to increase CFTR expression and maturation. They also suggest that the effect of GSNO is dependent on Csp and on intracellular SNO trafficking. We speculate that these data will be of relevance to the development of NO donor-based therapies for CF.
Footnotes
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This research was supported by grants 2R01-HL59337 and 5K12-HD001421-05 from the National Institutes of Health, by grant Zaman 04GO from the Cystic Fibrosis Foundation, and by the Ivy Foundation.
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ABBREVIATIONS: CF, cystic fibrosis; CFTR, cystic fibrosis transmembrane conductance regulatory protein; wt, wild-type; GSNO, S-nitrosoglutathione; GNODE, S-nitrosoglutathione diethyl ester; GGT, γ-glutamyl transpeptidase; ENO, ethyl nitrite; SNO, S-nitrosothiol; ER, endoplastic reticulum; MS, mass spectrometry; Csp, cysteine string protein; siRNA, small interfering RNA; CGSNO, S-nitrosocysteinyl glycine; SNOAC, S-nitroso-N-acetylcysteine; CHIP, carboxyl terminus Hsc70 interacting protein; ER, endoplasmic reticulum.
- Received February 6, 2006.
- Accepted July 11, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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