%0 Journal Article %A Ibiza, Sales %A Pérez-Rodríguez, Andrea %A Ortega, Ángel %A Martínez Ruiz, Antonio %A Barreiro, Olga %A Carlota A. García-Domínguez %A Víctor M. Víctor %A Juan V. Esplugues %A José M. Rojas %A Francisco Sánchez-Madrid %A Juan M. Serrador %T Endothelial nitric oxide synthase regulates N-Ras activation on the Golgi complex of antigen-stimulated T cells %D 2008 %@ 0027-8424 %@ 1091-6490 %U https://hdl.handle.net/20.500.14352/104316 %X Ras/ERK signaling plays an important role in T cell activation and development. We recently reported that endothelial nitric oxide synthase (eNOS)-derived NO regulates T cell receptor (TCR)- dependent ERK activation by a cGMP-independent mechanism. Here, we explore the mechanisms through which eNOS exerts this regulation. We have found that eNOS-derived NO positively regulates Ras/ERK activation in T cells stimulated with antigen on antigenpresenting cells (APCs). Intracellular activation of N-, H-, and K-Ras was monitored with fluorescent probes in T cells stably transfected with eNOS-GFP or its G2A point mutant, which is defective in activity and cellular localization. Using this system, we demonstrate thateNOS selectively activates N-Ras but not K-Ras on the Golgi complex of T cells engaged with APC, even though Ras isoforms are activated in response to NO from donors. We further show that activation of N-Ras involves eNOS-dependent S-nitrosylation on Cys118, suggesting that upon TCR engagement, eNOS-derived NO directly activates N-Ras on the Golgi. Moreover, wild-type but not C118S N-Ras increased TCR-dependent apoptosis, suggesting that S-nitrosylation of Cys118 contributes to activation-induced T cell death. Our data define a signaling mechanism for the regulation of the Ras/ERK pathway based on the eNOS-dependent differential activation of N-Ras and K-Ras at specific cell compartments. %~