iTRAQ-based quantitative proteomic analysis reveals the role of the tonoplast in fruit senescence.

UNLABELLED: The vacuole is by far the largest multifunctional organelle in fruits and plays a functional role in fruit development and fruit quality. Despite its significance, little information exists pertaining to the role of the vacuolar membrane (tonoplast) in the process of fruit senescence. In the present study, an iTRAQ-based quantitative proteomic approach was used to characterize the dynamic alterations in the tonoplast proteome during fruit senescence. Tonoplasts were purified from apple fruit at various stages of senescence using an iodixanol step gradient protocol. A total of 345 tonoplast-related proteins were identified with diverse functions such as transporters and proton pumps, signal transduction, membrane fusion or vesicle trafficking, cellular metabolic process, defense response, protein folding and degradation, and cytoskeleton. Changes in protein abundance during storage were characterized for the identified proteins. A total of 22 proteins displayed differential levels of abundance during storage. The senescence-related tonoplast proteins mostly function in the transportation of metabolites, signal transduction, membrane trafficking, and stress response. RT-qPCR analysis was used to quantify the level of expression of nine genes encoding some of the differentially abundant proteins. The results of this study provide new information regarding the function of the tonoplast during fruit senescence. BIOLOGICAL SIGNIFICANCE: Studies on the postharvest physiology and biochemistry of apple fruit have been conducted for several decades. Little proteomic information is available, however, pertaining to the role of the vacuole in fruit ripening and senescence. In the present study, an iTRAQ-based quantitative proteomic analysis was conducted on tonoplasts isolated from apple fruit in order to gain a global view of alterations in the tonoplast proteome during fruit senescence. The information obtained in the present study not only provides basic information about the tonoplast proteome in apple fruit but also characterized the quantitative changes that occur in the abundance of tonoplast proteins during the course of fruit senescence. This study provides a deeper insight into the cellular functions of the vacuole during fruit senescence that can serve as a basis for the development of future biotechnological strategies for the improvement of fruit quality.