Blocking Tyr265 nitration of protein phosphatase 2A attenuates nitrosative stress-induced endothelial dysfunction in renal microvessels

摘要

Protein tyrosine (Tyr) nitration, the covalent addition of a nitro group ((NO2)-N-center dot) to Tyr residues, is emerging as a candidate mechanism of endothelial dysfunction. Previous studies have shown that Tyr nitration is primarily induced by nitrosative stress, a process characterized by the production of reactive nitrogen species, especially peroxynitrite anion (ONOO-), which is considered a secondary product of NO in the presence of superoxide radicals (O-2(center dot-)). However, the impact of nitrosative stress-induced Tyr nitration on endothelial dysfunction has not been thoroughly elucidated to date. We developed an endothelial dysfunction model, a process called endothelial-to-mesenchymal transition (EndMT), and evaluated the production of NO, O-2(center dot-), and protein nitration during EndMT. The results showed that TGF-1 stimulation induced EndMT and elevated endothelial NO and O-2(center dot-) production as well as nitration of the catalytic subunit of protein phosphatase (PP)2A. Mass spectrometry analysis showed that Tyr265 was the nitration site in the catalytic subunit of protein phosphatase (PP)2A, and this Tyr nitration increased PP2A activity and disrupted endothelial integrity. To devise an endothelial-targeted anti-PP2Ac nitration strategy, a mimic peptide, tyrosine 265 wild type (Y265WT), conjugated with the cell-penetrating peptide HIV-1 TAT protein (TAT) was synthesized. PP2Ac nitration and PP2A activity were significantly inhibited by pretreatment with TAT-265WT, and the integrity of endothelial cells was maintained. Furthermore, injection of TAT-265WT attenuated renal nitration formation and caused anticapillary rarefaction in a unilateral urethral obstructive nephropathy model. Taken together, these results offer preclinical proof of concept for TAT-265WT as a tractable agent to protect against nitrosative stress-induced endothelial dysfunction in renal microvessels.Deng,Y., Cai, Y., Liu, L., Lin, X., Lu, P., Guo, Y., Han, M., Xu, G. Blocking Tyr265 nitration of protein phosphatase 2A attenuates nitrosative stress-induced endothelial dysfunction in renal microvessels.