Peroxynitrite induces tyrosine residue modifications in synaptophysin C-terminal domain, affecting its interaction with src.
Peroxynitrite is a potent oxidant that contributes to tissue damage in neurodegenerative disorders. We have previously reported that treatment of rat brain synaptosomes with peroxynitrite induced post-translational modifications in pre- and post-synaptic proteins and stimulated soluble N-ethylmaleimide sensitive fusion proteins attachment receptor complex formation and endogenous glutamate release. In this study we show that, following peroxynitrite treatment, the synaptic vesicle protein synaptophysin (SYP) can be both phosphorylated and nitrated in a dose-dependent manner. We found that tyrosine-phosphorylated, but not tyrosine-nitrated, SYP bound to the src tyrosine kinase and enhanced its catalytic activity. These effects were mediated by direct and specific binding of the SYP cytoplasmic C-terminal tail with the src homology 2 domain. Using mass spectrometry analysis, we mapped the SYP C-terminal tail tyrosine residues modified by peroxynitrite and found one nitration site at Tyr250 and two phosphorylation sites at Tyr263 and Tyr273. We suggest that peroxynitrite-mediated modifications of SYP may be relevant in modulating src signalling of synaptic terminal in pathophysiological conditions.
Mallozzi
Cinzia
C
Department of Cell Biology and Neuroscience, Istituto Superiore di Sanità, Viale Regina Elena, Rome, Italy.
Ceccarini
Marina
M
Camerini
Serena
S
Macchia
Gianfranco
G
Crescenzi
Marco
M
Petrucci
Tamara Corinna
TC
Di Stasi
Anna Maria Michela
AM
D000818:Animals; D001921:Brain / Q000648:ultrastructure; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects; D016708:Synaptophysin / Q000737:chemistry* / Q000378:metabolism*; D013574:Synaptosomes / Q000187:drug effects*; D014443:Tyrosine / Q000378:metabolism*; D018909:src Homology Domains / Q000235:genetics / Q000502:physiology*; D019061:src-Family Kinases / Q000378:metabolism*
D016708:Synaptophysin; D030421:Peroxynitrous Acid; D014443:Tyrosine; D019061:src-Family Kinases