Similar genetic mechanisms underlie the parallel evolution of floral phenotypes

The repeated origin of similar phenotypes is invaluable for studying the underlying genetics of adaptive traits; molecular evidence, however, is lacking for most examples of such similarity. The floral morphology of neotropical Malpighiaceae is distinctive and highly conserved, especially with regard to symmetry, and is thought to result from specialization on oil-bee pollinators. We recently demonstrated that CYCLOIDEA2-like genes (CYC2A and CYC2B) are associated with the development of the stereotypical floral zygomorphy that is critical to this plant-pollinator mutualism. Here, we build on this developmental framework to characterize floral symmetry in three clades of Malpighiaceae that have independently lost their oil bee association and experienced parallel shifts in their floral morphology, especially in regard to symmetry. We show that in each case these species exhibit a loss of CYC2B function, and a strikingly similar shift in the expression of CYC2A that is coincident with their shift in floral symmetry. These results indicate that similar floral phenotypes in this large angiosperm clade have evolved via parallel genetic changes from an otherwise highly conserved developmental program.     

Floral zygomorphy, the recurring evolution of a successful trait

The flowers of the primitive angiosperm plants were radially symmetrical (actinomorphic). Flowers with bilateral symmetry (zygomorphic) evolved in several clades independently as an adaptation to specialized methods of pollination and played an important role in the diversification of flowering plants. In the model species Antirrhinum majus (snapdragon), the related genes CYCLOIDEA (CYC) and DICHOTOMA (DICH) are key in the development of this trait. This raises the question of whether they played a role in the evolution of floral bilateral symmetry. To address this, the evolution of CYC in relation to the evolution of zygomorphy is being investigated. Phylogenetic and functional analyses of CYC-like genes are being carried out in groups either closely related to Antirhinum or in families where zygomorphy evolved as an independent event. In addition, the origin of zygomorphy is being studied by comparing the function of CYC-like genes in species with zygomorphic flowers with their function in species with radially symmetrical flowers.     

RETARDED PALEA1 controls palea development and floral zygomorphy in rice

Poaceae, one of the largest flowering plant families in angiosperms, evolved distinct inflorescence and flower morphology diverging from eudicots and other monocots. However, the mechanism underlying the specification of flower morphology in grasses remains unclear. Here we show that floral zygomorphy along the lemma-palea axis in rice (Oryza sativa) is partially or indirectly determined by the CYCLOIDEA (CYC)-like homolog RETARDED PALEA1 (REP1), which regulates palea identity and development. The REP1 gene is only expressed in palea primordium during early flower development, but during later floral stages is radially dispersed in stamens and the vascular bundles of the lemma and palea. The development of palea is significantly retarded in the rep1 mutant and its palea has five vascular bundles, which is similar to the vascular pattern of the wild-type lemma. Furthermore, ectopic expression of REP1 caused the asymmetrical overdifferentiation of the palea cells, altering their floral asymmetry. This work therefore extends the function of the TCP gene family members in defining the diversification of floral morphology in grasses and suggests that a common conserved mechanism controlling floral zygomorphy by CYC-like genes exists in both eudicots and the grasses.     

Significance of consensus CYC-binding sites found in the promoters of both ChCYC and ChRAD genes in Chirita heterotricha (Gesneriaceae)

CYC-like genes are widely conserved in controlling floral dorsoventral asymmetry (zygomorphy) through persistent expression in corresponding domains in core eudicots. To understand how CYC-like gene expression is maintained during flower development, we selected Chirita heterotricha as a material and isolated the promoter sequences of the ChCYCIC and ChCYCID genes, homologs of CYC, by inverse polymerase chain reaction. Further promoter analyses led to the identification of a putative cis-regulatory element in each promoter matching the consensus DNA binding site for Antirrhinum CYC protein: GGCCCCTC at-165 for ChCYC1C, and GGCCCCCC at-163 for ChCYCID. This indicates that both the ChCYCIC and ChCYC1D genes have probably evolved autoregulatory loops to sustain their expression in developing flowers. We also isolated the coding and promoter sequences of the ChRAD gene, a homolog of Antirrhinum RAD. Promoter analysis showed that the ChRAD gene promoter also contained a putative CYC-binding site (GGCCCAC at -134). Therefore, ChRAD is likely a direct target of the ChCYC1 genes, which is similar to Antirrhinum RAD. These results imply that the establishment of floral zygomorphy in Chirita may have been achieved by the evolution of an autoregulatory loop for CYC-like genes,which was probably accompanied by simultaneous co-option of the RAD-like gene into their regulatory network.     

苦苣苔科大叶石上莲CYC类基因RT-PCR表达模式研究

CYC类基因的分子系统学研究已经在苦苣苔科Gesneriaceae中展开,但是还缺乏对这些基因表达和功能的研究。因此,我们选择苦苣苔科大叶石上莲Oreocharis benthamii作为实验材料,分离出了CYC类基因的两个拷贝,经过分子系统学分析这两个基因分别属于苦苣苔科GCYC1和GCYC2两个分支,故命名为ObCYC1和ObCYC2。分区的RT-PCR实验结果显示这两个基因拥有不同的时间空间表达模式。ObCYC1与模式植物金鱼草Antirrhinum majus中的CYC基因类似,集中在花冠背部区域表达,这与它们拥有保守的功能区TCP和R相一致。但是,ObCYC1与CYC表达模式仍有区别,即,和CYC相比ObCYC1在花冠背部区域的表达提前减弱。这可能和大叶石上莲花冠微弱的两侧对称性相关。另外,由于大叶石上莲的背部花瓣较两侧和腹部花瓣小,因此,在功能上ObCYC1可能起抑制背部花瓣生长作用而CYC基因则促进背部花瓣生长。与ObCYC1不同,ObCYC2的保守功能区有更多的氨基酸位点变化,而且在RT-PCR实验中也没有检测到它的表达。因此,需要开展更深入的实验研究分析ObCYC2的基本功能,这将有助于了解GCYC2类基因在苦苣苔科及其近缘科中的功能和进化途径。     

利用农杆菌介导的瞬时表达系统研究CYC类TCP蛋白的亚细胞定位

CYCLOIDEA（CYC）类TCP基因在豆科与玄参科植物中都参与了两侧对称花型的发育,但它们的具体功能有明显的差异。通过在豌豆花瓣中建立农杆菌EHA105介导的瞬时表达系统,观察到豆科植物百脉根和玄参科植物金鱼草的CYC类TCP蛋白亚细胞定位有明显区别。     

Evolution of regulatory interactions controlling floral asymmetry

A key challenge in evolutionary biology is to understand how new morphologies can arise through changes in gene regulatory networks. For example, floral asymmetry is thought to have evolved many times independently from a radially symmetrical ancestral condition, yet the molecular changes underlying this innovation are unknown. Here, we address this problem by investigating the action of a key regulator of floral asymmetry, CYCLOIDEA (CYC), in species with asymmetric and symmetric flowers. We show that CYC encodes a DNA-binding protein that recognises sites in a downstream target gene RADIALIS (RAD) in Antirrhinum. The interaction between CYC and RAD can be reconstituted in Arabidopsis, which has radially symmetrical flowers. Overexpression of CYC in Arabidopsis modifies petal and leaf development, through changes in cell proliferation and expansion at various stages of development. This indicates that developmental target processes are influenced by CYC in Arabidopsis, similar to the situation in Antirrhinum. However, endogenous RAD-like genes are not activated by CYC in Arabidopsis, suggesting that co-option of RAD may have occurred specifically in the Antirrhinum lineage. Taken together, our results indicate that floral asymmetry may have arisen through evolutionary tinkering with the strengths and pattern of connections at several points in a gene regulatory network.     

Genetic analysis of floral symmetry in Van Gogh's sunflowers reveals independent recruitment of CYCLOIDEA genes in the Asteraceae

The genetic basis of floral symmetry is a topic of great interest because of its effect on pollinator behavior and, consequently, plant diversification. The Asteraceae, which is the largest family of flowering plants, is an ideal system in which to study this trait, as many species within the family exhibit a compound inflorescence containing both bilaterally symmetric (i.e., zygomorphic) and radially symmetric (i.e., actinomorphic) florets. In sunflower and related species, the inflorescence is composed of a single whorl of ray florets surrounding multiple whorls of disc florets. We show that in double-flowered (dbl) sunflower mutants (in which disc florets develop bilateral symmetry), such as those captured by Vincent van Gogh in his famous nineteenth-century sunflower paintings, an insertion into the promoter region of a CYCLOIDEA (CYC)-like gene (HaCYC2c) that is normally expressed specifically in WT rays is instead expressed throughout the inflorescence, presumably resulting in the observed loss of actinomorphy. This same gene is mutated in two independent tubular-rayed (tub) mutants, though these mutations involve apparently recent transposon insertions, resulting in little or no expression and radialization of the normally zygomorphic ray florets. Interestingly, a phylogenetic analysis of CYC-like genes from across the family suggests that different paralogs of this fascinating gene family have been independently recruited to specify zygomorphy in different species within the Asteraceae.     

Significance of consensus CYC‐binding sites found in the promoters of both ChCYC and ChRAD genes in Chirita heterotricha (Gesneriaceae)

Abstract CYC‐like genes are widely conserved in controlling floral dorsoventral asymmetry (zygomorphy) through persistent expression in corresponding domains in core eudicots. To understand how CYC‐like gene expression is maintained during flower development, we selected Chirita heterotricha as a material and isolated the promoter sequences of the ChCYC1C and ChCYC1D genes, homologs of CYC, by inverse polymerase chain reaction. Further promoter analyses led to the identification of a putative cis‐regulatory element in each promoter matching the consensus DNA binding site for Antirrhinum CYC protein: GGCCCCTC at ?165 for ChCYC1C, and GGCCCCCC at ?163 for ChCYC1D. This indicates that both the ChCYC1C and ChCYC1D genes have probably evolved autoregulatory loops to sustain their expression in developing flowers. We also isolated the coding and promoter sequences of the ChRAD gene, a homolog of Antirrhinum RAD. Promoter analysis showed that the ChRAD gene promoter also contained a putative CYC‐binding site (GGCCCAC at ?134). Therefore, ChRAD is likely a direct target of the ChCYC1 genes, which is similar to Antirrhinum RAD. These results imply that the establishment of floral zygomorphy in Chirita may have been achieved by the evolution of an autoregulatory loop for CYC‐like genes, which was probably accompanied by simultaneous co‐option of the RAD‐like gene into their regulatory network.