Proteomic analysis of strawberry achenes reveals active synthesis and recycling of L-ascorbic acid.

Although the commonly named strawberry fruit (Fragaria×ananassa) is the sum of achenes and receptacles, the true fruit in the botanical sense is the achene. Here we report the protein changes occurring in the achene when developing from immature to mature stage. We have used 2-DE followed by image analysis, and protein identification by PMF combined with MS/MS, to investigate the protein variations associated to this transition. From a total of 331 spots analyzed, the corresponding 315 proteins have been identified. Differentially accumulated proteins between immature and mature achenes mostly reflect the physiological events associated to seed development and maturation, with only a few changes related to the development of the dry pericarp. We have focused our attention on vitamin C biosynthesis. Interestingly, GDP-mannose 3',5'-epimerase, a key enzyme in the l-ascorbate biosynthesis pathway, and ascorbate peroxidase, involved in l-ascorbic acid oxidation, accumulate in immature achenes. The higher amount of these enzymes found in the green achene is coincident with a higher content of l-ascorbate, and higher expression levels of these and other gene encoding enzymes of the l-ascorbic acid biosynthesis pathway. Altogether our results suggest an important role of l-ascorbic acid at the early developmental stage of the achene. BIOLOGICAL SIGNIFICANCE: In this manuscript we report the identification of the most abundant proteins in strawberry (F.×ananassa) achenes at early and late stages of development, thus providing a proteomic view of the events that occur during the development of this organ. Despite the importance of strawberry as a commercial fruit, the molecular changes governing its growth and ripening processes are largely unknown. The lack of information is even greater in the case of the achenes, which are the true fruit and play a critical role in the developmental process of the receptacle. Our original proteomic study reported here, restricted to the achenes, completes the previous transcriptomic (very limited) and metabolomic maps of this organ, adding clarity to the role of the achene in the global ripening process. The results obtained not only complement the previous "omics" studies significantly, but also open new key questions that deserve further research (role of hormones). We finally focus on the biosynthesis of l-ascorbic acid, which appears to be tightly regulated by some specific pathways, and whose content is important in the achene. The information provided here will be of interest not only for the groups studying strawberry, but also for many other groups interested in the fruit ripening process, as well as for groups studying the regulation of l-ascorbic acid content in different plant tissues.