Over-expression of the Gerbera hybrida At-SOC1-like1 gene Gh-SOC1 leads to floral organ identity deterioration

Abstract

Background and aims: The family of MADS box genes is involved in a number of processes besides controlling floral development. In addition to supplying homeotic functions defined by the ABC model, they influence flowering time and transformation of vegetative meristem into inflorescence meristem, and have functions in roots and leaves. Three Gerbera hybrida At-SOC1-like genes (Gh-SOC1-Gh-SOC3) were identified among gerbera expressed sequence tags.

Methods: Evolutionary relationships between SOC1-like genes from gerbera and other plants were studied by phylogenetic analysis. The function of the gerbera gene Gh-SOC1 in gerbera floral development was studied using expression analysis, protein-protein interaction assays and reverse genetics. Transgenic gerbera lines over-expressing or downregulated for Gh-SOC1 were obtained using Agrobacterium transformation and investigated for their floral phenotype.

Key results: Phylogenetic analysis revealed that the closest paralogues of At-SOC1 are Gh-SOC2 and Gh-SOC3. Gh-SOC1 is a more distantly related paralogue, grouping together with a number of other At-SOC1 paralogues from arabidopsis and other plant species. Gh-SOC1 is inflorescence abundant and no expression was seen in vegetative parts of the plant. Ectopic expression of Gh-SOC1 did not promote flowering, but disturbed the development of floral organs. The epidermal cells of ray flower petals appeared shorter and their shape was altered. The colour of ray flower petals differed from that of the wild-type petals by being darker red on the adaxial side and greenish on the abaxial surface. Several protein-protein interactions with other gerbera MADS domain proteins were identified.

Conclusions: The At-SOC1 paralogue in gerbera shows a floral abundant expression pattern. A late petal expression might indicate a role in the final stages of flower development. Over-expression of Gh-SOC1 led to partial loss of floral identity, but did not affect flowering time. Lines where Gh-SOC1 was downregulated did not show a phenotype. Several gerbera MADS domain proteins interacted with Gh-SOC1.

Fig. 1.

Phylogenetic relationships of SOC1-like genes from Gerbera hybrida (Gh-SOC1, FR716023; Gh-SOC2, FR846140; Gh-SOC3, FR846141), Arabidopsis thaliana (At-SVP, At2g22540; At-SOC1, At2g45660; At-AGL71, At5g51870; At-AGL72, At5g51860; At-AGL42, At5g62165; At-AGL14, At4g11880; At-AGL19, At4g22950), Petunia hybrida (Ph-UNS, AF335238; Ph-FBP21, AF335239; Ph-FBP22, AF335240; Ph-FBP28, AF335244), Dendranthema grandiflorum (Cm-SOC1, AB481225) and Helianthus annuus (Ha-SOC1, GU985593). Gh-SOC2 and Gh-SOC3 are the putative orthologues of At-SOC1. Bootstrap support is shown when it exceeds 50 %.

Fig. 2.

Expression pattern of Gh-SOC1 in different gerbera organs (A) and at different petal developmental stages (B) (for stages, see Helariutta et al., 1993). R, roots; LP, leaf petiole; LB, leaf blade; SC, scape; BR, involucral bracts; YI, young inflorescence (6–16 mm in diameter); RE, receptacle; PB, pappus bristles; P, petals; ST, stamens; C, carpel; O, ovary; RF, ray flower; DF, disc flower.

Fig. 3.

RNA in situ expression analysis of Gh-SOC1 in developing gerbera inflorescences (inflorescence diameter 6–13·6 mm, antisense probe) showing ray flowers (A) and disc flowers (B). Abbreviations: P, petal; S, stamen; C, carpel; O, ovary; VB, vascular bundles; PB, pappus bristles. Expression is seen in petals, ovary, carpel, stamen and pappus bristles. Controls were hybridized with a Gh-SOC1 sense probe (C). Scale bars = 100 µm.

Fig. 4.

Gerbera hybrida wild-type ‘Terra Regina’ disc flower vs. Gh-SOC1 over-expression line SAT17 TR6 disc flower. The petals of the over-expression line lack anthocyanin and the stamen structure is abnormal.

Fig. 5.

Gerbera hybrida wild-type ‘Terra Regina’ flower types vs. Gh-SOC1 over-expression line SAT17 TR6 flower types. All the flower types of the over-expression line were shorter than the respective flower types in the wild type. Abbreviations: RF, ray flower; TR, trans flower; DF, disc flower. The scale is in cm.

Fig. 6.

Scanning electron microscopy analysis of the Gh-SOC1 over-expression line SAT17 TR6. Petal adaxial epidermal cells (A, D), petal abaxial epidermal cells (B, E) and carpel epidermal cells (C, F) of disc flowers from theGh-SOC1 over-expression line (A–C) and non-transgenic control plants (D–F). The epidermal cell structure of both the adaxial and abaxial petal surfaces of the Gh-SOC1 over-expression line differs from the wild-type petal epidermal cell structure. The carpel of the Gh-SOC1 over-expression line displays guard cell-like structures not normally found in wild-type carpels. Scale bars = 100 µm.

Fig. 7.

Yeast two-hybrid analysis of protein–protein interactions between Gh-SOC1 and other gerbera MADS domain proteins. Red, strong interaction; pink, weak interaction; blue, no interaction detected.

Fig. 8.

Yeast three-hybrid analysis of ternary protein complex formation among Gh-SOC1 and other gerbera MADS domain proteins. The strength of interactions is indicated.