科研之友
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摘要
Objective. CD4+ T cells from patients with active lupus have impaired ERK pathway signaling that decreases DNA methyltransferase expression, resulting in DNA demethylation, overexpression of immune genes, and autoimmunity. The ERK pathway defect is due to impaired phosphorylation of T-505 in the protein kinase C delta (PKC delta) activation loop. However, the mechanisms that prevent PKC delta T-505 phosphorylation in lupus T cells are unknown. Others have reported that oxidative modifications, and nitration in particular, of T cells as well as serum proteins correlate with lupus disease activity. We undertook this study to test our hypothesis that nitration inactivates PKC delta, contributing to impaired ERK pathway signaling in lupus T cells.
Methods. CD4+ T cells were purified from lupus patients and controls and then stimulated with phorbol myristate acetate (PMA). Signaling protein levels, nitration, and phosphorylation were quantitated by immunoprecipitation and immunoblotting of T cell lysates. Transfections were performed by electroporation.
Results. Treating CD4+ T cells with peroxynitrite nitrated PKC delta, preventing PKC delta T-505 phosphorylation and inhibiting ERK pathway signaling similar to that observed in lupus T cells. Patients with active lupus had higher nitrated T cell PKC delta levels than did controls, which correlated directly with disease activity, and antinitrotyrosine immunoprecipitations demonstrated that nitrated PKC delta, but not unmodified PKC delta, was refractory to PMA-stimulated T-505 phosphorylation, similar to PKC delta in peroxynitrite-treated cells.
Conclusion. Oxidative stress causes PKC delta nitration, which prevents its phosphorylation and contributes to the decreased ERK signaling in lupus T cells. These results identify PKC delta as a link between oxidative stress and the T cell epigenetic modifications in lupus.
- 出版日期2012-9
