S-Nitrosation of Cellular Proteins by NO Donors in Rat Embryonic Fibroblast 3Y1 Cells: Factors Affecting S-Nitrosation

Norihiro Ryuman,Takao Arai,Nobuo Watanabe

doi:10.1155/2011/450317

Norihiro Ryuman, Takao Arai + Show 1 more

Open Access

https://doi.org/10.1155/2011/450317

Copy DOI

Abstract

The mechanism of protein S-nitrosation in cells is not fully understood. Using rat 3Y1 cells, we addressed this issue. Among S-nitrosothiols and NO donors tested, only S-nitrosocysteine (CysNO) induced S-nitrosation when exposed in Hanks' balanced salt solution (HBSS) and not in serum-containing general culture medium. In HBSS, NO release from CysNO was almost completely abolished by sequestering metal ions with a metal chelator without affecting cellular S-nitrosation. In contrast, L-leucine, a substrate of L-type amino acid transporters (LATs), significantly inhibited S-nitrosation. The absence of S-nitrosation with CysNO in general culture medium resulted not only from a competition with amino acids in the medium for LATs but also from transnitrosation of cysteine residues in serum albumin. Collectively, these results suggest that in simple buffered saline, CysNO-dependent S-nitrosation occurs through a cellular incorporation-dependent mechanism, but if it occurs in general culture media, it may be through an NO-dependent mechanism.

Highlights

Nitric oxide (NO) plays diverse roles in physiological processes, such as vasodilatation, host defenses against infection, and neuromodulation, some of which are mediated by the activation of the guanylate cyclase (GS)/cGMP pathway [1, 2]
To clarify whether NO derived from CysNO degradation in the medium was responsible for S-nitrosation of cellular proteins, we examined the effect of diethylenetriaminepentaacetic acid (DETAPAC) on S-nitrosation by CysNO in cells maintained in Hanks’ balanced salt solution (HBSS)
The present study demonstrates that in buffered saline only CysNO can cause significant levels of protein S-nitrosation in rat 3Y1 cells when exposed for a short period of time

Summary

Introduction

Nitric oxide (NO) plays diverse roles in physiological processes, such as vasodilatation, host defenses against infection, and neuromodulation, some of which are mediated by the activation of the guanylate cyclase (GS)/cGMP pathway [1, 2]. More than 100 proteins have been identified to undergo S-nitrosation [3]

Methods

Results

Conclusion