Cell-surface protein disulfide isomerase catalyzes transnitrosation and regulates intracellular transfer of nitric oxide
Adrian Zai, … , Anne Ward Scribner, Joseph Loscalzo
Adrian Zai, … , Anne Ward Scribner, Joseph Loscalzo
Published February 1, 1999
Citation Information: J Clin Invest. 1999;103(3):393-399. https://doi.org/10.1172/JCI4890.
View: Text | PDF
Article

Cell-surface protein disulfide isomerase catalyzes transnitrosation and regulates intracellular transfer of nitric oxide

Abstract

Since thiols can undergo nitrosation and the cell membrane is rich in thiol-containing proteins, we considered the possibility that membrane surface thiols may regulate cellular entry of NO. Recently, protein disulfide isomerase (PDI), a protein that catalyzes thio–disulfide exchange reactions, has been found on the cell-surface membrane. We hypothesized that cell-surface PDI reacts with NO, catalyzes S-nitrosation reactions, and facilitates NO transfer from the extracellular to intracellular compartment. We observed that PDI catalyzes the S-nitrosothiol–dependent oxidation of the heme group of myoglobin (15-fold increase in the rate of oxidation compared with control), and that NO reduces the activity of PDI by 73.1 ± 21.8% (P < 0.005). To assess the role of PDI in the cellular action of NO, we inhibited human erythroleukemia (HEL) cell-surface PDI expression using an antisense phosphorothioate oligodeoxynucleotide directed against PDI mRNA. This oligodeoxynucleotide decreased cell-surface PDI content by 74.1 ± 9.3% and PDI folding activity by 46.6 ± 3.5% compared with untreated or “scrambled” phosphorothioate oligodeoxynucleotide–treated cells (P < 0.0001). This decrease in cell-surface PDI was associated with a significant decrease in cyclic guanosine monophosphate (cGMP) generation after S-nitrosothiol exposure (65.4 ± 26.7% reduction compared with control; P < 0.05), with no effect on cyclic adenosine monophosphate (cAMP) generation after prostaglandin E1 exposure. These data demonstrate that the cellular entry of NO involves a transnitrosation mechanism catalyzed by cell-surface PDI. These observations suggest a unique mechanism by which extracellular NO gains access to the intracellular environment.

Authors

Adrian Zai, M. Audrey Rudd, Anne Ward Scribner, Joseph Loscalzo

×

Figure 5

Options: View larger image (or click on image) Download as PowerPoint
Reduction in cell-surface PDI folding activity with antisense treatment....
Reduction in cell-surface PDI folding activity with antisense treatment. Refolding of GS-RNase T1 was performed at 25°C in PBS (pH 7.4). RNase folding activity was measured as extent of folding at 45 min; 100% activity corresponds to the maximal folding by control HEL cells. GS-RNase T1 (6 μM final concentration) was added to incubation mixtures containing 10,000 cells/μl. In antisense-treated cells, folding activity decreased by 46.6 ± 3.5% compared with control. GS-RNase, glutathionyl-RNase T1.