Limits...
Proteomic analysis of grape berry cell cultures reveals that developmentally regulated ripening related processes can be studied using cultured cells.

Sharathchandra RG, Stander C, Jacobson D, Ndimba B, Vivier MA - PLoS ONE (2011)

Bottom Line: 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.The 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Viticulture and Oenology, Institute for Wine Biotechnology, Stellenbosch University, Stellenbosch, South Africa.

ABSTRACT

Background: This 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 findings: In 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.

Conclusions: The 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.

Show MeSH

Related in: MedlinePlus

Growth curve analysis of suspension cells derived from berry explants.Shown are growth curves recorded for stage I (from green berries) and III (from ripe berries) suspension cultures that were monitored over a 21 day period. Samples for proteomic evaluation were taken at 7 (log phase) and 14 days (early stationary phase) after sub-culture.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3040747&req=5

pone-0014708-g003: Growth curve analysis of suspension cells derived from berry explants.Shown are growth curves recorded for stage I (from green berries) and III (from ripe berries) suspension cultures that were monitored over a 21 day period. Samples for proteomic evaluation were taken at 7 (log phase) and 14 days (early stationary phase) after sub-culture.

Mentions: The berry explants spanned the major stages of berry development and ripening (as reported [20]) and exhibited the typical double sigmoidal growth curve characteristic of berry development. The phenotypical berry characteristics at each of the sampling points are described in Table 1. Somatic callus cultures were successfully initiated from all nine sampling points, but explants in the green and ripe stages callused more readily than the samples in the lag and véraison stage of berry development (Table 1). Non-embryogenic callus developed from the skin and pulp sections of the explants (berry slices) within three to six weeks of culturing. The callus was translucent to cream in colour, loose and watery with a shiny appearance (Figure 1). The suspension cultures were successfully initiated with these callus clumps and consisted of friable, fast dividing callus cells that dispersed homogenously into the culture medium. The suspension cultures consisted of actively growing, homogenous, thin walled and highly vacuolated cells (Figure 2, and Supporting Information S1). The suspension cultures exhibited a typical sigmoidal growth curve with a lag-phase (0–2 days), log-phase (3–10 days) and a stationary phase (11–21 days) (Figure 3). The cultures could be maintained by weekly sub-culturing to provide actively proliferating mid-log phase cells for initiating fresh cell suspension cultures.


Proteomic analysis of grape berry cell cultures reveals that developmentally regulated ripening related processes can be studied using cultured cells.

Sharathchandra RG, Stander C, Jacobson D, Ndimba B, Vivier MA - PLoS ONE (2011)

Growth curve analysis of suspension cells derived from berry explants.Shown are growth curves recorded for stage I (from green berries) and III (from ripe berries) suspension cultures that were monitored over a 21 day period. Samples for proteomic evaluation were taken at 7 (log phase) and 14 days (early stationary phase) after sub-culture.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3040747&req=5

pone-0014708-g003: Growth curve analysis of suspension cells derived from berry explants.Shown are growth curves recorded for stage I (from green berries) and III (from ripe berries) suspension cultures that were monitored over a 21 day period. Samples for proteomic evaluation were taken at 7 (log phase) and 14 days (early stationary phase) after sub-culture.
Mentions: The berry explants spanned the major stages of berry development and ripening (as reported [20]) and exhibited the typical double sigmoidal growth curve characteristic of berry development. The phenotypical berry characteristics at each of the sampling points are described in Table 1. Somatic callus cultures were successfully initiated from all nine sampling points, but explants in the green and ripe stages callused more readily than the samples in the lag and véraison stage of berry development (Table 1). Non-embryogenic callus developed from the skin and pulp sections of the explants (berry slices) within three to six weeks of culturing. The callus was translucent to cream in colour, loose and watery with a shiny appearance (Figure 1). The suspension cultures were successfully initiated with these callus clumps and consisted of friable, fast dividing callus cells that dispersed homogenously into the culture medium. The suspension cultures consisted of actively growing, homogenous, thin walled and highly vacuolated cells (Figure 2, and Supporting Information S1). The suspension cultures exhibited a typical sigmoidal growth curve with a lag-phase (0–2 days), log-phase (3–10 days) and a stationary phase (11–21 days) (Figure 3). The cultures could be maintained by weekly sub-culturing to provide actively proliferating mid-log phase cells for initiating fresh cell suspension cultures.

Bottom Line: 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.The 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Viticulture and Oenology, Institute for Wine Biotechnology, Stellenbosch University, Stellenbosch, South Africa.

ABSTRACT

Background: This 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 findings: In 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.

Conclusions: The 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.

Show MeSH
Related in: MedlinePlus