Malate is a major organic acid that accumulates in grape berries (Vitis spp.) and contributes to sourness and other related fruit quality. In commercial cultivars (vinifera), malate accumulates at the green stage followed by a dramatic decline toward ripeness. However, the identification of genes involved in these shifts in fruit malate levels has been hampered by the complexity and time-constraint involved in developing transformed, fruit-bearing grapevines.
This research aims to establish fruit-derived cell culture lines as a platform to elucidate the metabolic processes and candidate genes determining malate levels in the fruit. For this purpose, we have characterized the changes in malate and related metabolites during the development of cells, cultured in liquid suspension. We have generated cell-culture lines that have contrasting levels of malate in ripe fruit, and developed protocols for the transfection and agrobacterium tumefaciens-mediated transient transformation of cells. As in the developing fruit, cells exhibited a marked increase in malate accumulation during the initial period of growth (six days), however, the decline in malate (days 10-18) occurred concomitantly with progressive cell death, increase in media pH, and a decrease in media oxygen levels. We conclude that berry-derived cell culture is a good model to study accumulation-related metabolism, and that the degradation of malate seems to result from cell death rather than active catabolism. Trials involving transient expression in fruit cell culture indicate it as a promising approach for the rapid screening of candidate genes involved in fruit physiology. Future trials to test the involvement of specific genes in the accumulation of malate in three contrasting berry-derived cell lines are underway.