A TCP domain transcription factor controls flower type specification along the radial axis of the Gerbera (Asteraceae) inflorescence

Abstract

Several key processes in plant development are regulated by TCP transcription factors. CYCLOIDEA-like (CYC-like) TCP domain proteins have been shown to control flower symmetry in distantly related plant lineages. Gerbera hybrida, a member of one of the largest clades of angiosperms, the sunflower family (Asteraceae), is an interesting model for developmental studies because its elaborate inflorescence comprises different types of flowers that have specialized structures and functions. The morphological differentiation of flower types involves gradual changes in flower size and symmetry that follow the radial organization of the densely packed inflorescence. Differences in the degree of petal fusion further define the distinct shapes of the Gerbera flower types. To study the role of TCP transcription factors during specification of this complex inflorescence organization, we characterized the CYC-like homolog GhCYC2 from Gerbera. The expression of GhCYC2 follows a gradient along the radial axis of the inflorescence. GhCYC2 is expressed in the marginal, bilaterally symmetrical ray flowers but not in the centermost disk flowers, which are nearly radially symmetrical and have significantly less fused petals. Overexpression of GhCYC2 causes disk flowers to obtain morphologies similar to ray flowers. Both expression patterns and transgenic phenotypes suggest that GhCYC2 is involved in differentiation among Gerbera flower types, providing the first molecular evidence that CYC-like TCP factors take part in defining the complex inflorescence structure of the Asteraceae, a major determinant of the family's evolutionary success.

Fig. 1.

Expression of GhCYC2 in various Gerbera tissues. GhCYC2 showed strongest expression in young inflorescences, petals, and carpel (stigma and style). Weaker expression was also seen in bracts and ovary. The lower blot shows ethidium bromide-stained ribosomal RNA bands to control for RNA loading.

Fig. 2.

GhCYC2 is expressed in the outermost ray flower primordia but not in the centermost disk flower primordia. RNA-blot analysis (A) was done for ray (RF) and disk flower (DF) primordia in developmental stages 3, 5, 6, and 7. Outer (DF-o) and centermost (DF-c) disk flowers were dissected separately at stages 3 and 5, whereas at stages 6 and 7, outer, inner (DF-i), and centermost disk flowers were separated. At stages 3 and 5, GhCYC2 expression was detected only in ray flowers and not in outer or centermost disk flowers. At later stages 6 and 7, in addition to ray flowers, GhCYC2 expression was detected in the outer and inner disk flowers (DF-i), but GhCYC2 expression was still excluded from the centermost disk flowers. SEM pictures (B–E) show that at stage 3 the Gerbera ray and disk flower primordia have similar petal (pe) and stamen (st) morphology but that at stage 5 the two flower types have distinct characteristics, such as altered petal symmetry. (Scale bars: 100 μm.)

Fig. 3.

In situ analysis of GhCYC2 expression in ray flowers (A–C) and in the outer disk flowers (D) of the Gerbera inflorescence (diameter 12 mm). Sections were bridized with GhCYC2 antisense RNA probes labeled with digoxigenin-UTP. In ray flowers, GhCYC2 was expressed in the ventral ligule (vLi) but not in the rudimentary dorsal petals (dPe), as seen in the cross (A) and longitudinal (C) sections. GhCYC2 expression was detected ubiquitously in the basal tubular part of ray flower petals (tuPe) (B). GhCYC2 expression was also detected as well in rudimentary stamens (ruSt) and in carpels (Ca) (B and C). In the outer disk flowers, GhCYC2 was expressed most clearly in stamens but also in carpel and petals (D). (Scale bars: 100 μm.)

Fig. 4.

Vegetative phenotype of the transgenic 35S::GhCYC2 Gerbera lines. Overexpression lines (tr) showed delayed growth (A) and smaller, more roundly shaped leaves (B). In wild type (wt), the inflorescence stem is bent during growth (C), but in transgenic lines the developing inflorescence faced upward (D). (Scale bars: 1 cm.)

Fig. 5.

The effect of GhCYC2 overexpression on Gerbera disk flowers. Transgenic disk flowers (tr df) had a clearly distinct phenotype compared with wild-type disk flowers (wt df). (A) Disk flower petals were longer and petals had a more pronounced ligular structure. Development of disk flower stamens (St) was disrupted. At the same developmental stages, wild-type disk flowers release pollen (B), but no pollen was seen in the 35S::GhCYC2 lines (A and C). In one of the overexpression lines, disk flower petals were fused together to form tubular structures (C). (Scale bar: 0.5 cm.)

Fig. 6.

The effect of suppressed GhCYC2 expression on Gerbera inflorescence phenotype. In nontransformed Terra Regina (A and B), trans flowers (wt tf) are longer than in the transgenic lines (A and C). Only the length of the distal ligule and not the basal petal tube differs between wild-type and transgenic trans flowers (tr tf) (A). (Scale bar: 0.5 cm.)