Gene expression profiling of candidate genes for flowering inititation and dormancy breaking in Rhododendron simsii hybrids

Pot azalea (R. simsii hybrids) production in the Ghent area (Belgium) accounts approximately 40 million plants per year. The past decade, growers and their associations have made efforts to improve both the quality of the vegetative as well as the flowering plants; however, suboptimal flowering is often observed. Different potential causes have been quoted but a clear cut direct cause is seldom found. However, we believe that the fact that pot azaleas are grown to flower almost year-round is an important causal factor. In nature, Rhododendron plants flower in spring and bud initiation occurs in September. In autumn, buds become dormant and a cold period during winter is required for dormancy-breaking and flowering in spring. In pot azalea growing, these conditions are artificially induced. Flower initiation is stimulated by the application of plant growth regulators and dormancy breaking by means of a cold-treatment for 6 to 8 weeks at 7°C. However, environmental factors such as deviating day lengths compared to the natural conditions in the wild can have an impact as well. Therefore we aimed at identifying the influential factors related to flowering quality by an integrated approach focusing on the induction of processes at the level of RNA expression and physiological measurements. Candidate genes for flower initiation (FT, TOC, CO, LFY, AG, AP3 and CRY) have been isolated in azalea, as well as genes potentially involved in the process of dormancy breaking (ELIP, dehydrin, ARP) and the carbohydrate metabolism (SUSY and ?-amylase). The expression of these genes was determined in ‘Hellmut Vogel’ plants grown under different experimental culture conditions. Long and short day lengths (8h and 16h) were applied at the time of flower initiation (in combination with plant growth regulators). The expression profiles of both FT and LFY clearly indicated when bud formation was initiated, but day-length was not a determining factor for initiation. Only minor differences could be seen between treatments and therefore we assume that azalea is a day-neutral plant. In a second experiment, we compared the impact of decreasing day-length after initiation, as occurs in nature, with a constant day-length, but again no significant differences could be seen at the level of initiation.
To evaluate gene expression under cold treatment, plants with fully developed buds (stage 7) were grown at 7°C (refrigerator) and 18°C (greenhouse) for 6 weeks. The expression of ELIP, ARP and dehydrin is clearly upregulated under cold conditions and decreases again after 5 weeks when dormancy has been released. When we look at the sugar metabolism, the expression of both SUSY and ?-amylase appears to be upregulated in plants grown at cold temperatures compared to plants kept in the greenhouse. These plants have a higher need for carbohydrates to protect the tissues against the cold temperatures and this was clearly demonstrated by the results of the gene expression studies. Measurements of the starch and sugar content in both leaves and buds confirmed these results. Although the gene expression profiles in this study were only implemented in pot azaleas, the same analysis could be done easily on Rhododendron plants as well.