Response of tetraploid Citrus wilsonii Tanaka to drought stress by phosphoproteomics analysis

Abstract [Objective] Using phosphoproteomics, expression patterns of the phosphorylated proteins were analyzed with tetraploid Citrus wilsonii leaves under drought stress, aiming to reveal the mechanism of tetraploid C. wilsonii in response to drought stress and provide support for the improvement of droughttolerant citrus rootstock varieties. [Methods] Phosphorylated proteins in the leaves of tetraploid C. wilsonii after drought stress were identified and analyzed using IMAC affinity enrichment and TMT labeling technology. Functional annotation and metabolic pathway analysis were performed for the differentially expressed phosphorylated proteins. [Results] (1) A total of 3 794 phosphorylation sites and 1 521 phosphorylated proteins were quantified. There were 662 phosphorylated proteins with a fold change exceeding 1.3 (αFC>1.3), which were mainly located in the nucleus (46.07%) and chloroplasts (24.62%). (2) The differentially expressed phosphorylated proteins were mainly involved in binding RNA and Ca2+ , and participating in metabolic pathways such as RNA splicing, photosynthesis, and SNARE interaction in vesicle transport. (3) RT-qPCR results showed that 92.86% of the genes coding the differentially expressed phosphorylated proteins showed similar trend of change in transcriptional and protein levels with the Pearson correlation coefficient of 0.893. [Conclusion] Tetraploid C. wilsonii regulates the proteins involved in RNA splicing and photosynthesis pathway through phosphorylation in response to drought stress.