Cell-to-cell transport through plasmodesmata in tree callus cultures

One factor that contributes to a successful fruit tree grafting is the establishment of symplasmic contacts in the graft interface to facilitate the transfer of compounds between scion and stock. Using novel experimental and theoretical approaches we investigated whether the localized incompatibility, experienced in some Prunus grafts, could be related to insufficient plasmodesmal coupling at an early stage of development within one of the partners. Dye-coupling analysis using fluorescent tracers combined with confocal laser scanning microscopy were performed in cultured callus from either the plum rootstock (Prunus cerasifera Ehrh.Prunus munsoniana W. Wight et Hedr.) cv. Marianna 2624 or from the apricot (Prunus armeniaca L.) cv. Moniqui growing in vitro. Fluorescein was loaded into callus cells in a caged form. Following photoactivation of fluorescence within single cells, the uncaged fluorescein could be traced as it was spreading cell-to-cell revealing the existence of functional plasmodesmata. This set of experiments was performed within the stock partner in callus fusions (callus grafts) as well as in ungrafted callus. The results indicated species-related as well as developmental-related differences in plasmodesmal conductivity. The results further pointed to a novel control factor of connectivity that reaches the graft partner and changes its innate rate of communication: when combining the poorly transporting apricot cultivar with the well-transporting plum cultivar, communication between plum callus cells was much reduced, compared to that in plum homografts. For further support of the hypothesis, we carried out a quantitative analysis in which fluorescein was esterloaded into the callus. Fluorescence redistribution after photobleaching of fluorescein in individual cells gave a measure for the plasmodesmal contact between the cells. We found significant differences between the species with regard to mobile fraction and halftime of redistribution, which confirmed that callus cells are not interconnected to the same extent in Marianna 2624 and Moniqui.