Supplementary Material for: Comparative phosphoproteomic profiling in the prefrontal cortex of prenatally stressed male offspring rats

Many investigations have indicated that prenatal stress (PS) causes depressive-like disturbances in offspring rats. However, the underlying pathogenic mechanisms have not yet been fully elucidated. The prefrontal cortex (PFC) has been shown to play a role in susceptibility to stress during fetal development; thus, we focused our attention on differential protein phosphorylation in this region of PS-S(Susceptibility to PS) offspring rats. The sucrose preference test was used to screen for susceptibility to PS. The validity of the prenatally stressed model was verified by other common depression-like behaviors. We used MS-based TMT quantitative proteomics in combination with the phosphopeptide enrichment method to compare phosphoproteomic profiling in the prefrontal cortex of PS-S and control male offspring rats. In total, 3418 phosphoproteins, 8404 phosphopeptides and 12175 phosphosites were identified in this analysis. According to the screening criteria, 902 phosphopeptides increased and 609 decreased in the PFC of the PS-S group compared to the control rats. GO enrichment analysis indicated that the main enriched terms in the cellular component category were ‘synapse part’, ‘myelin sheath’, ‘synapse’, ‘neuron part’ and ‘axon’. The phosphoproteins enriched in the molecular function and biological process categories were mainly related to cytoskeleton- and projection morphogenesis-associated proteins. KEGG pathway enrichment analyses identified 30 significant KEGG pathways; the top five pathways included salivary secretion, endocrine and other factor-regulated calcium reabsorption, pancreatic secretion and insulin secretion. Motifs such as......_S_P...RR, ......S_PE...., ......_S_PV...., ......_S_P.H... and ..S..._S_PT....were the top five motifs enriched in phosphorylated sites.PS may induce depressive-like behaviors in male offspring rats by regulating the phosphorylation of proteins mainly related to synapses, myelin sheaths, neurons and the cytoskeleton. The phosphorylation of related proteins may act as key pathogenic hits. Data are available via ProteomeXchange with identifier PXD026563.