jabbrv-ltwa-all.ldf jabbrv-ltwa-en.ldf Pistachio ( Pistacia vera L.) thrives in semi-arid and arid environments and is highly adaptable to various abiotic stresses. However, soil salinization significantly threatens productivity, leading to considerable osmotic and ionic stress for these plants. This study investigates the changes in root protein expression and metabolic pathways in response to sodium chloride-induced salt stress through biochemical and proteomics analyses. One-year-old pistachio rootstocks were treated with four different saline water regimes over 100 days, and the total proteins were isolated from these samples. Over 1,600 protein identifiers were detected, with comparative analysis revealing 245 proteins that were more abundant and 190 that were less abundant across three stress levels. Key pathways associated with stress tolerance, such as protein modification, folding, and heat shock protein (HSP) protection, were upregulated. An increase in secondary metabolites played a crucial role in detoxification. As salt stress intensified, the abundance of trafficking proteins increased, enhancing transporter activities. Active signaling pathways were observed at lower stress levels, while structural proteins became more critical at higher stress levels for maintaining cell membrane integrity. This cultivar demonstrated enhanced kinase activities that regulate lipid and carbohydrate metabolism, aiding in ion homeostasis and redox balance. The protein interaction network, mapped to orthologous proteins in Arabidopsis thaliana, revealed clusters associated with cytosolic, carbohydrate, and amino acid metabolism contributing to salinity stress tolerance. The validation of proteomic data was conducted through corresponding changes in transcript levels, marking this study as the first to map root proteomic changes under salt stress in pistachio.