Phosphoproteome profiling of hippocampal synaptic plasticity

• Published in: Biochemical and biophysical research communications Vol. 626; pp. 92 - 99
• Main Authors: Lim, So-Hee, Lee, Na-Yoon, Ryu, Ju Yeon, An, Jin Hua, Lee, Ga Seul, Min, Sun Seek, Moon, Jeonghee, Lee, Jae-Ran
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 20.10.2022
• Subjects: Hippocampal synaptic plasticity; Phosphopeptide enrichment; Phosphoproteomic analysis; Semi-quantification; Phosphoproteomic analysis; Hippocampal synaptic plasticity; Semi-quantification; Phosphopeptide enrichment
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2022.07.051

The balance between the actions of protein kinases and phosphatases is crucial for neuronal functions, including synaptic plasticity. Although the phosphorylation and dephosphorylation of neuronal proteins are regulated by synaptic plasticity, no systematic analyses of this have yet been conducted. We performed a phosphoproteomic analysis of hippocampal synaptic plasticity using a nano-Acquity/Synapt LC-MS/MS system. Neuronal proteins were extracted from hippocampal tissues and cultured neurons exposed to long-term potentiation (LTP) or long-term depression (LTD). Filter-aided sample preparation (FASP) was performed to remove residual anionic detergents for complete tryptic digestion. Phosphopeptides were then enriched using TiO2 chromatography, followed by immunoaffinity chromatography with an anti-phosphotyrosine antibody. Among the 1500 phosphopeptides identified by LC-MS/MS, 374 phosphopeptides were detected simultaneously in both hippocampal tissues and cultured neurons. Semi-quantification counting the number of spectra of each phosphopeptide showed that 42 of 374 phosphopeptides changed significantly depending on synaptic plasticity. In conclusion, a new proteomic method using sequential enrichment of phosphopeptides and semi-quantification enabled the phosphoproteomic analysis of hippocampal synaptic plasticity. •Phosphoproteomic analysis of hippocampal synaptic plasticity was performed using nano-Acquity/Synapt LC-MS/MS system.•New proteomic method using sequential enrichment of phosphopeptides and semi-quantification was conducted.•Filter-aided sample preparation enabled complete tryptic digestion of synaptic membrane-bound proteins.