Proteomics and metabolomics represent powerful tools of functional genomics, giving direct access to the molecular mechanisms underlying plant response on developmental signals, metabolic pathways, and regulatory networks. The combination of these approaches might give deeper insights into the molecular mechanisms contributing on mutualistic plant-microbial interactions, in particular, legume-rhizobial symbiosis. Here we address age-related changes in determinate and indeterminate root nodules of common bean (Phaseolus vulgaris) and pea (Pisum sativum). Juvenile, flowering, and senescent plants were harvested, the proteins were isolated by phenol extraction and digested with trypsin, isolating metabolites by a multi-step extraction with solvents of different polarity. Protein hydrolyzates were analyzed by nanoLC-ESI-Q- and LIT-Orbitrap-MS in data-dependent acquisition (DDA) mode. Database search and annotation of PTMs were based on the SEQUEST algorithm; quantification was performed using a label-free strategy. Functions and localization of the proteins were annotated by an in-house developed workflow, combined with MapMan, Mercator, LocTree3 and String tools. Analysis of primary metabolome relied on the combination of GC-MS and IP-RP-UHPLC-MS, with a secondary metabolite profiling by RP-UHPLC-MS. The analysis revealed 656 and 92 differentially regulated proteins in common bean and pea nodules, respectively. Plant proteins were mostly down-regulated and represented by the molecules involved in signaling and protein metabolism, while most of the bacterial polypeptides were up-regulated and found to be involved in energy metabolism and nitrogen fixation. Agedependent changes in metabolite profiles were characterized based on the accumulation of carbohydrates and amino acids. Interestingly, mutation in the gene sym27 resulted in the suppression of signaling and protein biosynthesis in both symbiosis partners. Remarkably, nodule ageing was accompanied with enhanced glycation and carbonylation. In total, 36 age-related glycation hotspots were identified in the P. vulgaris nodule proteome. This project was supported by the Russian Science Foundation (project 17-16-01042).
|Name||ACTA NATURAE (РУССКОЯЗЫЧНАЯ ВЕРСИЯ)|
|Conference||II ОБЪЕДИНЕННЫЙ НАУЧНЫЙ ФОРУМ, VI СЪЕЗД ФИЗИОЛОГОВ СНГ,|
VI СЪЕЗД БИОХИМИКОВ РОССИИ, IX РОССИЙСКИЙ СИМПОЗИУМ «БЕЛКИ И ПЕПТИДЫ»
|Period||1/10/19 → 6/10/19|