Dynamic N -glycoproteome analysis of maize seedling leaves during de-etiolation using Concanavalin A lectin affinity chromatography and a nano-LC–MS/MS-based iTRAQ approach

Key message The identification of N -glycosylated proteins with information about changes in the level of N -glycosylation during de-etiolation provides a database that will aid further research on plant N -glycosylation and de-etiolation. Abstract N -glycosylation is one of the most prominent and abundant protein post-translational modifications in all eukaryotes and in plants it plays important roles in development, stress tolerance and immune responses. Because light-induced de-etiolation is one of the most dramatic developmental processes known in plants, seedlings undergoing de-etiolation are an excellent model for investigating dynamic proteomic profiles. Here, we present a comprehensive, quantitative N -glycoproteomic profile of maize seedlings undergoing 12 h of de-etiolation obtained using Concanavalin A (Con A) lectin affinity chromatography enrichment coupled with a nano-LC–MS/MS-based iTRAQ approach. In total, 1084 unique N -glycopeptides carrying 909 N -glycosylation sites and corresponding to 609 proteins were identified and quantified, including 186 N -glycosylation sites from 162 proteins that were significantly regulated over the course of the 12 h de-etiolation period. Based on hierarchical clustering analysis, the significantly regulated N -glycopeptides were divided into seven clusters that showed different N -glycosylation patterns during de-etiolation. We found no obvious difference in the enriched MapMan bincode categories for each cluster, and these clustered significantly regulated N -glycoproteins (SRNPs) are enriched in miscellaneous, protein, cell wall and signaling, indicating that although the N -glycosylation regulation patterns of these SRNPs might differ, they are involved in similar biological processes. Overall, this study represents the first large-scale quantitative N -glycoproteome of the model C4 plant, maize, which is one of the most important cereal and biofuel crops. Our results greatly expand the maize N -glycoproteomic database and also shed light on the potential roles of N -glycosylation modification during the greening of maize leaves.