Arabidopsis genes AS1, AS2, and JAG negatively regulate boundary-specifying genes to promote sepal and petal development

Boundary formation is crucial for organ development in multicellular eukaryotes. In higher plants, boundaries that separate the organ primordia from their surroundings have relatively low rates of cell proliferation. This cellular feature is regulated by the actions of certain boundary-specifying genes, whose ectopic expression in organs can cause inhibition of organ growth. Here, we show that the Arabidopsis thaliana ASYMMETRIC LEAVES1 and 2 (AS1 and AS2) and JAGGED (JAG) genes function in the sepal and petal primordia to repress boundary-specifying genes for normal development of the organs. Loss-of-function as1 jag and as2 jag double mutants produced extremely tiny sepals and petals. Analysis of a cell-cycle marker HISTONE4 revealed that cell division in sepal primordia of the double mutant was inhibited. Moreover, these abnormal sepals and petals exhibited ectopic overexpression of the boundary-specifying genes PETAL LOSS (PTL) and CUP-SHAPED COTYLEDON1 [corrected] and 2 (CUC1 and CUC2). Loss of PTL or CUC1 and CUC2 functions in the as1 jag background could partially rescue the tiny sepal and petal phenotypes, supporting the model that the tiny sepal/petal phenotypes are caused, at least in part, by ectopic expression of boundary-specifying genes. Together, our data reveal a previously unrecognized fundamental regulation by which AS1, AS2, and JAG act to define sepal and petal from their boundaries.     

Ectopic expression of a single homeotic gene, the Petunia gene green petal, is sufficient to convert sepals to petaloid organs.

Genetic studies in Arabidopsis and Antirrhinum showed that petal determination requires the concomitant expression of two homeotic functions, A and B, whereas the A function alone determines sepal identity. The B function is represented by at least two genes. The Petunia homeotic gene green petal (gp) is essential for petal determination as demonstrated by a Petunia gp mutant that has sepals instead of petals. We have used ectopic expression of the gp gene as a tool to study flower development in Petunia. CaMV 35S-gp expression leads to homeotic conversion of sepals into petaloid organs when expressed early in development. This demonstrates that a single homeotic gene is sufficient to induce homeotic conversion of sepals to petals, suggesting that other petal determining genes are regulated in part by ectopically expressed gp. Indeed, two other MADS-box-containing genes, pmads 2 and fbp 1, which show homology to the Antirrhinum B function gene globosa, are activated in the converted petal tissue. Furthermore, our data provide evidence for autoregulation of gp expression in the petaloid tissue and uncover the role of gp in fusion of petal tissues.     

FRL1 is required for petal and sepal development in Arabidopsis

A novel flower mutant, frl1 (frill 1) was isolated in Arabidopsis thaliana. The frl1 mutant has serrated petals and sepals but the other floral and vegetative organs appear to be normal. To analyse the role of the FRL1 gene, morphological, cytological and double mutant analyses were carried out. The frl1 flower had broader petals and sepals as compared with the wild-type. The distal region of frl1 petals contained fewer epidermal cells but their size was variable and generally larger than that in the wild-type. However, no significant difference was found in the basal region. Observations of the early petal development revealed that the morphology of the developing frl1 petal was normal until the middle of stage 9, but the frl1 phenotype became apparent in stages later than 10. Furthermore, larger nuclei with varied sizes were observed in the distal region of frl1 petals, but not in this region in wild-type petals. This strongly suggests that abnormal endo-reduplication had occurred. These observations indicate that the frl1 mutation affects the number of cell divisions and the subsequent cell expansion during the late stage of petal lamina formation, and that FRL1 might be maintaining the mitotic state or suppressing the transition to the endo-reduplication cycle. Double mutants with the homeotic mutants apetala3-1 and agamous showed additive phenotypes. Ectopic petals in the third whorl of fr11 ag flowers were serrated, indicating that the FRL1 gene acts in petal and sepal development in an organ-specific manner.     

APETALA1突变影响WRKY基因的基础表达

拟南芥APETALA1（AP1）既是一个花分生组织特征基因又是一个花器官特征基因,在花器官发育中控制花萼和花瓣的发育。通过GUS染色进一步证实AP1主要在茎尖、花萼、花瓣和花托的位置表达。启动子分析发现,AP1启动子区包含了包括W-box在内的大量顺式作用元件,暗示相关转录调控因子参与了对AP1的调控。21个WRKY基因单突变后并不改变AP1在花中的表达,但是AP1突变则增强了检测的10个WRKY基因中7个WRKY基因的表达,暗示AP1参与了对WRKY基因的基础表达的调控。这个结果也暗示AP1可能通过控制花萼和花瓣的发育从而参与了对花的基础抗性。     

Petal Development in Lotus japonicus

Previous studies have demonstrated that petal shape and size in legume flowers are determined by two separate mechanisms, dorsoventral (DV) and organ internal (IN) asymmetric mechanisms, respectively. However, little is known about the molecular mechanisms controlling petal development in legumes. To address this question, we investigated petal development along the floral DV axis in Lotus japonicus with respect to cell and developmental biology by comparing wild‐type legumes to mutants. Based on morphological markers, the entire course of petal development, from initiation to maturity, was grouped to define 3 phases or 13 stages. In terms of epidermal micromorphology from adaxial surface, mature petals were divided into several distinct domains, and characteristic epidermal cells of each petal differentiated at stage 9, while epidermal cells of all domains were observed until stage 12. TCP and MIXTA‐like genes were found to be differentially expressed in various domains of petals at stages 9 and 12. Our results suggest that DV and IN mechanisms interplay at different stages of petal development, and their interaction at the cellular and molecular level guides the elaboration of domains within petals to achieve their ideal shape, and further suggest that TCP genes determine petal identity along the DV axis by regulating MIXTA‐like gene expression.     

PETAL LOSS gene regulates initiation and orientation of second whorl organs in the Arabidopsis flower

PETAL LOSS is a new class of flower development gene whose mutant phenotype is confined mostly to the second whorl. Two properties are disrupted, organ initiation and organ orientation. Initiation is frequently blocked, especially in later-formed flowers, or variably delayed. The few petals that arise occupy a wider zone of the flower primordium than normal. Also, a minority of petals are trumpet-shaped, thread-like or stamenoid. Studies of ptl combined with homeotic mutants have revealed that the mutant effect is specific to the second whorl, not to organs with a petal identity. We propose that the PTL gene normally promotes the induction of organ primordia in specific regions of the second floral whorl. In ptl mutants, these regions are enlarged and organ induction is variably reduced, often falling below a threshold. A dominant genetic modifier of the ptl mutant phenotype was found in the Landsberg erecta strain that significantly boosts the mean number of petals per flower, perhaps by reinforcing induction so that the threshold is now more often reached. The second major disruption in ptl mutants relates to the orientation adopted by second whorl organs from early in their development. In single mutants the full range of orientations is seen, but when B function (controlling organ identity) is also removed, most second whorl organs now face outwards rather than inwards. Orientation is unaffected in B function single mutants. Thus petals apparently perceive their orientation within the flower primordium by a mechanism requiring PTL function supported redundantly by that of B class genes.     

蝴蝶兰PhalPI基因的克隆及在花器官突变体中的表达分析

为深入研究兰科植物花器官发育的调控机理,从蝴蝶兰花瓣中克隆了一个B类MADS-box转录因子PhalPI（GenBank登录号为KY020416）。序列分析表明,该基因的cDNA全长为944 bp,含完整的开放阅读框,可编码210个氨基酸,属于BGLO/PI蛋白家族,与蝴蝶兰属的PhPI10和PeMADS6基因关系最近;表达模式分析表明,PhalPI基因在生殖器官中表达,在营养器官中不表达,在授粉后的子房中,该基因的表达水平降低。在5种花器官突变体中,PhalPI基因在萼片唇瓣化突变体的萼片和蕊柱中表达水平明显升高;在雄蕊花瓣化突变体的萼片和侧瓣中表达水平降低,在其唇瓣和蕊柱中显著升高;在侧瓣合柱化突变体的蕊柱中,PhalPI基因的表达也发生了显著升高;PhalPI基因表达的改变与花器官形态的突变相关;而在侧瓣唇瓣化和侧瓣花药化突变体中,PhalPI基因的表达水平没有变化。推测该基因在决定蝴蝶兰侧瓣和唇瓣的发育中起重要的调控作用。     

A comparison of venation pattern development in wild type and a homeotic sepaloid petal mutant of Clarkia tembloriensis (Onagraceae)

Vascular system development in sepals, petals, and sepaloid petals was compared in wild-type and crinkled petal mutant plants of Clarkia tembloriensis. Patterns of vascularization in cleared whole mounts were visualized and traced under both brightfield and polarizing illumination. Wild-type sepals exhibited a basipetal pattern of maturation, with tracheary elements maturing relatively rapidly. Mature sepals had three primary veins with numerous secondary veins. In contrast, wild-type petals exhibited an acropetal pattern of maturation, with tracheary elements maturing relatively slowly. The mature petals had only one primary vein with numerous secondary veins. Sepaloid (crinkled) petals combined characteristics of both wild-type sepals and wild-type petals. They exhibited a basipetal pattern of development and a relatively rapid maturation of the tracheary elements characteristic of wild-type sepals. Venation architecture in crinkled petal mutants showed a single primary vein with numerous secondary veins, similar to wild-type petals. The crinkled petal mutant fits the definition of a homeotic mutant in that the petal has assumed characteristics of the sepal. However, homeotic transformation from petal to sepal is incomplete since the crinkled petal still retains many of the characteristics of wild-type petals.     

CINCINNATA controls both cell differentiation and growth in petal lobes and leaves of Antirrhinum

To understand how differentiation and growth may be coordinated during development, we have studied the action of the CINCINNATA (CIN) gene of Antirrhinum. We show that in addition to affecting leaf lamina growth, CIN affects epidermal cell differentiation and growth of petal lobes. Strong alleles of cin give smaller petal lobes with flat instead of conical cells, correlating with lobe-specific expression of CIN in the wild type. Moreover, conical cells at the leaf margins are replaced by flatter cells, indicating that CIN has a role in cell differentiation of leaves as well as petals. A weak semidominant cin allele affects cell types mainly in the petal but does not affect leaf development, indicating these two effects can be separated. Expression of CIN correlates with expression of cell division markers, suggesting that CIN may influence petal growth, directly or indirectly, through effects on cell proliferation. For both leaves and petals, CIN affects growth and differentiation of the more distal and broadly extended domains (leaf lamina and petal lobe). However, while CIN promotes growth in petals, it promotes growth arrest in leaves, possibly because of different patterns of growth control in these systems.     

'Living stones' reveal alternative petal identity programs within the core eudicots

Petals, defined as the showy laminar floral organs in the second floral whorl, have been shown to be under similar genetic control in distantly related core eudicot model organisms. On the basis of these findings, it is commonly assumed that the petal identity program regulated by B-class MADS-box gene homologs is invariant across the core eudicot clade. However, the core eudicots, which comprise >70% of angiosperm species, exhibit numerous instances of petal and sepal loss, transference of petal function between floral whorls, and recurrent petal evolution. In the face of these complex patterns of perianth evolution, the concept of a core eudicot petal identity program has not been tested. We therefore examined the petal identity program in the Caryophyllales, a core eudicot clade in which perianth differentiation into sepals and petals has evolved multiple times. Specifically, we analyzed the expression patterns of B- and C-class MADS-box homologs for evidence of a conserved petal identity program between sepal-derived and stamen-derived petaloid organs in the 'living stone' family Aizoaceae. We found that neither sepal-derived nor stamen-derived petaloid organs exhibit gene expression patterns consistent with the core eudicot petal identity program. B-class gene homologs are not expressed during the development of sepal-derived petals and are not implicated in petal identity in stamen-derived petals, as their transient expression coincides with early expression of the C-class homolog. We therefore provide evidence for petal development that is independent of B-class genes and suggest that different genetic control of petal identity has evolved within this lineage of core eudicots. These findings call for a more comprehensive understanding of perianth variation and its genetic causes within the core eudicots--an endeavor that will have broader implications for the interpretation of perianth evolution across angiosperms.     

Are Petals Sterile Stamens or Bracts? The Origin and Evolution of Petals in the Core Eudicots

BACKGROUND: The aim of this paper is to discuss the controversial origins of petals from tepals or stamens and the links between the morphological expression of petals and floral organ identity genes in the core eudicots. SCOPE: I challenge the widely held classical view that petals are morphologically derived from stamens in the core eudicots, and sepals from tepals or bracts. Morphological data suggest that tepal-derived petals have evolved independently in the major lineages of the core eudicots (i.e. asterids, Santalales and rosids) from Berberidopsis-like prototypes, and that staminodial petals have arisen only in few isolated cases where petals had been previously lost (Caryophyllales, Rosales). The clear correlation between continuous changes in petal morphology, and a scenario that indicates numerous duplications to have taken place in genes controlling floral organ development, can only be fully understood within a phylogenetic context. B-gene expression plays a fundamental role in the evolution of the petals by controlling petaloidy, but it does not clarify petal homology. CONCLUSIONS: An increased synorganization of the flower in the core eudicots linked with the establishment of floral whorls restricts the petaloid gene expression to the second whorl, reducing the similarities of petals with tepals from which they were originally derived. An increased flower size linked with secondary polyandry or polycarpelly may lead to a breakdown of the restricted gene expression and a reversal to ancestral characteristics of perianth development. An altered 'sliding boundary' hypothesis is proposed for the core eudicots to explain shifts in petaloidy of the perianth and the event of staminodial petals. The repetitive changes of function in the perianth of the core eudicots are linked with shifts in petaloidy to the outer perianth whorl, or losses of petal or sepal whorls that can be secondarily compensated for by the inclusion of bracts in the flower. The origin and evolution of petals appears to be as complex on a molecular basis as it is from a morphological point of view.     

THE MISSING PETAL CHARACTER IN OENOTHERA AND ITS RELATION TO THE CRUCIATE CHARACTER

Two characters are known in Oenothera which show inconstancy of behavior resulting in phenotypic mosaicism. These are absence of petals mp and cruciate petals cr. The latter character has been studied intensively by Oehlkers and Renner. The former is discussed here for the first time. The missing petal character exhibits mosaicism in essentially all cases. Flowers with four, three, two, one, or no petals may appear on the same plant on the same day. Where petals are present, they occupy normal positions and are usually normal in size and shape; where absent, no primordia are produced. It is suggested that the cr character is not based on a highly mutable locus (Oehlkers) or on one in which gene conversion occurs (Renner), but is the result of a mutant gene at a locus basic to the development of sepals which is. capable, under certain conditions, of functioning not only in the sepals, but also in cells of petal primordia, thereby suppressing genes for petal development. The sepaloid tissue which it produces in the petal is much more complex than, the petaloid tissue which it suppresses. The mp locus is basic to the initiation of petals; mp is a mutant gene with reduced potency. Whether it is able to function depends upon the cellular environment in which it finds itself. In both cases mosaicism is the result, not of frequently recurring alteration in genic structure, but of regulation of gene action based on variations in the cellular milieu.     

RABBIT EARS is a second-whorl repressor of AGAMOUS that maintains spatial boundaries in Arabidopsis flowers

The RABBIT EARS (RBE) gene has been identified as a regulator of petal development in Arabidopsis thaliana. We find that second-whorl petals in rbe mutants can be replaced with staminoid organs, stamens or filaments and that some rbe flowers have increased numbers of sepals and exhibit fusion of sepals. We show that these rbe defects are due to AGAMOUS (AG) misexpression in the second whorl. Consistent with its role in maintaining the spatial boundary of AG expression, rbe enhanced the second-whorl defects present in ap2-1, lug-1 and clf-2 mutants. In the development of second-whorl organs, RBE acts in the same pathway and downstream of UNUSUAL FLORAL ORGANS (UFO). Enhanced first-whorl organ fusion in ap2-2 rbe-3, ant-4 rbe-3 and cuc2-1 rbe-3 double mutants supports an additional role for RBE in organ separation. RBE thus acts to maintain two different types of spatial boundaries in young flowers: boundaries between organ primordia within a whorl and boundaries of homeotic gene expression between whorls.