Quantitative Phosphoproteomic and Physiological Analyses Provide Insights into the Formation of the Variegated Leaf in <i>Catalpa fargesii</i>

Variegated plants are valuable materials for investigating leaf color regulated mechanisms. To unveil the role of posttranslational modification in the variegated phenotype, we conducted global quantitative phosphoproteomic analysis on different leaf color sectors of Maiyuanjinqiu and the corresponding of Catalpa fargesii using Ti4+-IMAC phosphopeptide enrichment. A total of 3778 phosphorylated sites assigned to 1646 phosphoproteins were identified, and 3221 in 1434 proteins were quantified. Differential phosphoproteins (above 1.5 or below 1/1.5) in various leaf color sectors were selected for functional enrichment analyses. Gene ontology (GO) enrichment revealed that processes of photosynthesis, regulation of the generation of precursor metabolites, response to stress, homeostasis, amino acid metabolism, transport&#8211;related processes, and most of the energy metabolisms might contribute to leaf color. KEGG pathway enrichment analysis was performed based on differential phosphoproteins (DPs) in different organelles. The result showed that most enriched pathways were located in the chloroplasts and cytosol. The phosphorylation levels of glycometabolism enzymes might greatly affect leaf variegation. Measurements of fluorescence parameters and enzyme activities confirmed that protein phosphorylation could affect plant physiology by regulating enzyme activity. These results provide new clues for further study the formation mechanisms of naturally variegated phenotype