Proteomic Analysis of Grape Berry Cell Cultures Reveals that Developmentally Regulated Ripening Related Processes Can Be Studied Using Cultured Cells

Abstract

BackgroundThis work describes a proteomics profiling method, optimized and applied to berry cell suspensions to evaluate organ-specific cultures as a platform to study grape berry ripening. Variations in berry ripening within a cluster(s) on a vine and in a vineyard are a major impediment towards complete understanding of the functional processes that control ripening, specifically when a characterized and homogenous sample is required. Berry cell suspensions could overcome some of these problems, but their suitability as a model system for berry development and ripening needs to be established first.Methodology/Principal FindingsIn this study we report on the proteomic evaluation of the cytosolic proteins obtained from synchronized cell suspension cultures that were established from callus lines originating from green, véraison and ripe Vitis vinifera berry explants. The proteins were separated using liquid phase IEF in a Microrotofor cell and SDS PAGE. This method proved superior to gel-based 2DE. Principal component analysis confirmed that biological and technical repeats grouped tightly and importantly, showed that the proteomes of berry cultures originating from the different growth/ripening stages were distinct. A total of twenty six common bands were selected after band matching between different growth stages and twenty two of these bands were positively identified. Thirty two % of the identified proteins are currently annotated as hypothetical. The differential expression profile of the identified proteins, when compared with published literature on grape berry ripening, suggested common trends in terms of relative abundance in the different developmental stages between real berries and cell suspensions.ConclusionsThe advantages of having suspension cultures that accurately mimic specific developmental stages are profound and could significantly contribute to the study of the intricate regulatory and signaling networks responsible for berry development and ripening.

Response of Vitis vinifera cell cultures to Eutypa lata and Trichoderma atroviride culture filtrates: expression of defence-related genes and phenotypes

“…Proteomic analysis of these cells showed low expression levels of PR proteins (Sharathchandra et al 2011), making them ideal for elicitation studies. Gene expression and metabolite production by the cell cultures proved to be dependent on the elicitor, further confirming the suitability of the cell cultures.…”

“…Dauphine were established from callus derived from pea size green berries as previously described by Sharathchandra et al (2011). The suspension cultures were grown in 250-mL Erlenmeyer flasks containing 100 mL of Calderon medium (Calderon et al 1994).…”

“…Dauphine, were used. These suspensions were previously characterised (Sharathchandra et al 2011) and shown to constitutively produce low levels of defence-related proteins. In this study, the cell suspension cultures were treated with extracts from the two fungi in a time-course experiment of molecular gene expression (q-RT-PCR) and metabolite profiling (enzyme activities and polyphenol content).…”

Chromatographic and Electrophoretic Prefractionation Tools in Proteome Analysis

“…On a similar line of thinking, Davidsson et al (2001) have subfractionated human cerebrospinal fluid and brain tissue, whereas Wang et al (2002) have mapped the proteome of ovarian carcinoma cells. This instrument is quite popular in proteome prefractionation and has been used, together with the miniaturized version, for a variety of applications, including the exploration of the proteome of grape berry cell cultures (Sharathchandra et al, 2011). All these attempts were made with the objective of discovering the maximum number of protein components, most of them being of low abundance and not detectable prior to fractionation.…”

Functional analysis of differentially expressed proteins in Chinese bayberry ( Myrica rubra Sieb. et Zucc.) fruits during ripening

“…Proteomics is a powerful approach to characterize biochemical networks and to establish functional correlations between genotype and phenotype. Differential proteomic studies have been accomplished on immature and mature fruits, including tomato (Rocco et al, 2006), grape (Sharathchandra, Stander, Jacobson, Ndimba, & Vivier, 2011), orange (Pan et al, 2012), apricot (D'Ambrosio et al, 2013), peach (Nilo, Campos-Vargas, & Orellana, 2012), strawberry (Bianco et al, 2009), mango (Andrade et al, 2012), papaya , and apple (Costa et al, 2010). The present work investigated Chinese bayberry fruits at three ripening stages by using a comparative proteomic approach based on 2-DE and MALDI-TOF/TOF-MS.…”