Flower initiation and flower development

Once floral induction took place, flower initiation and further differentiation will take place. There are two major groups of genes that regulate floral development: meristem identity genes which encode transcription factors that are necessary for the initial induction of the second group of genes, the organ identity genes. These floral organ identity genes directly control floral identity (Irish, 2010).

- Floral meristem identity genes

In Arabidopsis, the critical meristem identity genes that must be activated are: SOC1, AP1 and LFY.

SOC1 and LFY play a central role in flower initiation by integrating the signals from different

pathways of environmental and endogenous regulators of floral induction (Figure 1.2). If SOC1 is activated, it turns on the expression of LFY, which triggers the expression of AP1. AP1 then stimulates the expression of LFY, which creates a positive feedback loop. In Rhododendron, homologues of LFY and SOC1 have been isolated (Cheon et al., 2012).

- Floral organ identity genes

The floral organs arise from proliferation of the floral meristem. The genetic pathways controlling the specification of different organ identities are well studied. The floral organ identity genes fall into three classes: types A, B and C and the control of organ identity relies on these three types of genes and is referred to as the ABC model (Figure 1.3). Type A activity, encoded by AP1 and AP2, controls the organ identity in the first and second whorls. Type B genes, APETALA3 (AP3) and

PISTILLATA (PI) control the second and third whorls. Type C gene AGAMOUS (AG) controls the

third and fourth whorls. The activity of type A genes alone specifies the sepals. For petal formation, both A and B genes are required. Type B and C genes form stamens and the activity of C alone defines carpels. Of these floral organ identity genes, only a homologue to AP3 has been isolated in Rhododendron (Cheon et al., 2011).

Loss of function of one of the ABC-genes will lead to flower malformation. Woody plants have a variety of floral modifications. In Rhododendron, sepals, stamens and carpels can become petaloïd, which results in beautiful doubled-flowers but leaving them infertile.

Figure 1.3 ABC model in Arabidopsis to determine the floral organ identity based on the interactions of type A, B and C genes (Taiz and Zeiger, 2010)

After the activation of the floral organ identity genes, the formation of floral parts occurs sequentially and can be followed microscopically in azalea. Bodson (1983) defined 9 stages in the development of the flower bud (Figure 1.4). Stage 0: vegetative bud; stage 1: initiation of bud scales, stage 2: initiation of flower primordia, stage 3: sepal initiation, stage 4: petal initiation, stage 5: stamen initiation, stage 6: carpel initiation, stage 7: initiation of style elongation, stage 8: the ovary contains ovules. The initiation phase comprises stages 1-2. Differentiation starts from stage 3.

Figure 1.4 Flower development of azalea. Top: stage 1, 2 and 4; bottom: stage 5, 7 and 8

1.5.3 Dormancy

Before further development of the flower buds to anthesis, there is a period of rest, i.e. dormancy. Dormancy is a common mechanism in woody plants of the temperate zone to prevent buds from resuming growth in unfavourable conditions. To break dormancy and to be able to continue further development, buds require a prolonged exposure to low temperatures. This chilling requirement is different among species and among cultivars (Cooke et al., 2012).

Three types of dormancy have been defined by Lang (1987) depending on which factors play a role. During ecodormancy, environmental factors prevent growth, while for paradormancy and endodormancy the growth inhibition arises from the plant itself. Paradormancy refers to factors from other plant parts inhibiting growth (e.g. apical dominance); endodormancy refers to factors inside the dormant structure itself which prevent initiation of growth from meristems (with the capacity to resume growth), under favourable environmental conditions (Rohde and Bhalerao, 2007). The regulation and mechanisms of dormancy are complex and still not fully understood. The induction of endodormancy can be regulated by the environmental factors shortening day length and lower temperatures. This makes it difficult to distinguish physiological and molecular changes associated with dormancy regulation from those of cold acclimation. Bud dormancy has been extensively studied and a number of reviews are present dealing with physiological and molecular aspects of bud dormancy in trees (Arora et al., 2003; Horvath et al., 2003; Chao et al., 2007; Rohde and Bhalerao, 2007; Ruttink et al., 2007). Research on flower bud dormancy focused mainly on temperate fruit trees and it is believed that similar processes as for vegetative buds play a role. However, the depth of dormancy can be different between vegetative and generative buds (Horvath, 2009). Flower buds seem to be less dormant than vegetative buds, but still require an equal amount of chilling units (Gariglio et al., 2006). Research on Rhododendron mainly

focusses on cold acclimation and cold hardiness of flower buds, but not on the process of flower bud dormancy (Arora and Rowland, 2011).

1.5.4 Anthesis

The last step in the flowering process is anthesis, i.e. the opening of the flower. The opening of flowers is characterised by 2 steps: petal growth by cell division, and unfolding of the petals by cell expansion. In these processes sugar supply is essential as flower development is associated with increased soluble carbohydrates in the petal cells. These carbohydrates provide energy for growth respiration and also lower the osmotic potential which promotes the influx of water (Reid, 2003). The research on flower opening has mainly been done on ornamental plants such as roses (Kumar et al., 2007), lily (van Doorn and Han, 2011), tulip and Alstroemeria (Collier 1997); and focussed mainly on source-sink relations and sucrose metabolism, as they prove to be essential for petal growth and expansion (Kumar et al., 2008b). For cut flowers, post-harvest quality has been thoroughly investigated with attention to sucrose metabolism, and ethylene induced senescence (van Doorn and Han, 2011).

During flowering of azalea, different stages in flower opening can be distinguished which are used to determine the quality of flowering. Buds develop from a closed (green) bud, to a colour- showing bud (CS), candle stage bud (CA) and ultimately an open flower (OF) (Figure 1.5). The commercial forcing period ends when plants have buds in the colour-showing stage or in the candle stage.

Figure 1.5 Four stages during flowering, from left to right: green bud (G), colour-showing bud (CS), candle- stage bud (CA), open flower (OF)