Time of day effects on post-exercise phosphoproteomic profiling in mouse hippocampus

At both the rest phase (ZT3) and the active phase (ZT15) for mice, the acute exercise mouse model and the sham-exercise group were established. Left hippocampi from every two mice were mixed as one sample and three replicates were adopted in each group. Then the hippocampal phosphoproteome and proteome were screened by affinity enrichment and LC-MS/MS. Aspects of differentially phosphorylated sites and proteins between the rest and active phase were compared, including functional enrichment analysis, PPI network analysis, motif analysis and kinase prediction, etc. as a result, exercise significantly changed 932 and 828 phosphosites at the early rest phase and the early active phase, respectively. There were only 49 overlapped differentially regulated sites on 44 proteins. Functional enrichment analysis displayed that the breadth of signaling pathways modulated by exercise at ZT3 was far greater than that at ZT15. The phosphorylation status of differentially phosphorylated proteins enriched in multiple pathways was vastly different after the timed exercise, so were the predicted upstream kinases. Up to 29% differentially phosphorylated proteins were related to the structure or function of synapses, and both exercise at ZT3 and ZT15 might significantly affect the pathway of the glutamate synapse-calcium signaling pathway-long-term potential in hippocampus.