Abstract
Grapevine (Vitis vinifera) is an economically important fruit crop that is subject to many types of insect and pathogen attack. To better elucidate the plant response to Lobesia botrana pathogen infection, we initiated a global comparative proteomic study monitoring steady-state protein expression as well as changes in N-glycosylation, phosphorylation, and Lys-acetylation in control and infected mesocarp and exocarp from V. vinifera cv Italia. A multi-parallel, large-scale proteomic approach employing iTRAQ labeling prior to three peptide enrichment techniques followed by tandem mass spectrometry led to the identification of a total of 3059 proteins, 1135 phosphorylation sites, 323 N-linked glycosylation sites and 138 Lys-acetylation sites. Of these, we could identify changes in abundance of 899 proteins. The occupancy of 110 phosphorylation sites, 10 N-glycosylation sites and 20 Lys-acetylation sites differentially changed during L. botrana infection. Sequence consensus analysis for phosphorylation sites showed eight significant motifs, two of which containing up-regulated phosphopeptides (X-G-S-X and S-X-X-D) and two containing down-regulated phosphopeptides (R-X-X-S and S-D-X-E) in response to pathogen infection. Topographical distribution of phosphorylation sites within primary sequences reveal preferential phosphorylation at both the N- and C termini, and a clear preference for C-terminal phosphorylation in response to pathogen infection suggesting induction of region-specific kinase(s). Lys-acetylation analysis confirmed the consensus X-K-Y-X motif previously detected in mammals and revealed the importance of this modification in plant defense. The importance of N-linked protein glycosylation in plant response to biotic stimulus was evident by an up-regulated glycopeptide belonging to the disease resistance response protein 206. This study represents a substantial step toward the understanding of protein and PTMs-mediated plant-pathogen interaction shedding light on the mechanisms underlying the grape infection.
Highlights
Grapevine (Vitis vinifera) is considered one of the most commercially important fruit crops because of the variety of derived food products including wine, table grape, and raisins
Our aims were to gain insight into the sequence of events that occur in the V. vinifera “Italia cv” proteome and post-translational modifications (PTMs) during infection, and to identify proteins that contribute to the susceptibility and response of this grapevine to L. botrana
We used state-of-the-art proteomics and PTMomics tools to gain insights into the sequence of events underlying the V. vinifera infection, and to identify proteins that contribute to the susceptibility and response of this grapevine to L. botrana
Summary
Grapevine (Vitis vinifera) is considered one of the most commercially important fruit crops because of the variety of derived food products including wine, table grape, and raisins. Plants dramatically change at the molecular and biochemical levels as reflected by global transcriptome (2– 4), proteome (5, 6), and metabolome (7) system analyses These studies demonstrate the importance of the complex modulation of plant metabolism and defense responses during pathogen infection. These changes activate a series of biological signaling mechanisms such as Ca2ϩ influx (8, 9), kinase cascades (10, 11), reactive oxygen species (12, 13), and phytohormone signaling pathways (14). Our aims were to gain insight into the sequence of events that occur in the V. vinifera “Italia cv” proteome and PTMs during infection, and to identify proteins that contribute to the susceptibility and response of this grapevine to L. botrana. This work represents the first comprehensive study of protein phosphorylation, N-glycosylation, and Lys-acetylation on a plant system after infection with a pathogen
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