Floral Whorls Research Articles

Krameria tomentosa oil flowers and their pollinators: Bees specialized on trichome elaiophores exploit its epithelial oil glands

Oil flowers are spread over eleven angiosperm families and exploited by bees specialized on oil gathering. This reward is produced by glandular surfaces of specific floral whorls called elaiophores. Unlike trichome elaiophores, such as those of Plantaginaceae, Krameria species bear lateral petals modified into epithelial elaiophore, convergently similar to oil-secreting Malpighiaceae calyx. Krameriaceae is an oil-offering monotypic group of eighteen species with little information on their pollination biology and interactions with oil-collecting bees. In this study, we evaluate the breeding system and describe the floral biology of Krameria tomentosa, a widespread species that occurs mainly in open vegetation habitats of northeastern Brazil. We listed its flower visitors, surveyed throughout four years, and recorded their visits by season. The pollination treatments showed that K. tomentosa is self-compatible; however, its flowers depend on pollinators to set fruits. Twenty-one Apidae visited the flowers, including the oil-bee Centris hyptidis specialized on trichome elaiophores and stingless-bees which foraged for pollen. Centris nitens and Trigona fulviventris were the most frequent visitors. K. tomentosa flowers received relatively few oil visits throughout the day and on some days no flower visitors were seen on its flowers. Nevertheless, this did not reflect on a low natural fruit set. C. hyptidis has been associated with Angelonia (Plantaginaceae) trichome elaiophores mainly due to its long forelegs and their specialized oil-collecting apparatus. Nonetheless, it exploited the K. tomentosa epithelial elaiophores in an area with no Plantaginaceae species. We presume that the association between the morphology of the oil-collecting apparatus and the elaiophore type is not enough to predict relationships in oil-systems. Additionally, we conclude that the diversity of bee visitors suggests that pollen-collecting bees also play an important role in the maintenance of K. tomentosa pollination.

Molecular characterisation of four double-flowered mutants of Silene dioica representing four centuries of variation.

Records of double-flowered Silene dioica date from the late sixteenth century and four named varieties are grown today, as previously, for their horticultural interest. Although double-flowered mutants have been characterized in several plants, their study in dioecious species is of particular interest due to influences of the homeotic mutation on the different floral whorl configurations in males and females. We have analysed four double-flowered varieties of Silene dioica: Flore Pleno and Rosea Plena date back to the seventeenth and nineteenth centuries, Thelma Kay and Firefly were recognized in the latter part of the twentieth and early twenty-first centuries. We have analysed the floral structure of the four varieties, which have distinct floral architectures. Based on Y chromosome-specific PCR analysis we show that Firefly is male and that the other three varieties are female: Random Amplification of Polymorphic DNA (RAPD) analyses suggested a common origin for the three female varieties. The double-flowered phenotype in all four varieties is caused by mutation of the C-function MADS-box transcription factor gene SDM1. We show that Firefly carries a unique 44bp insertion into SDM1, revealing an independent origin for this variety. Comparative analysis of SDM1 cDNA and genomic sequences in Flore Pleno, Rosea Plena and Thelma Kay shows that all three are caused by the same 7bp insertion within SDM1 and therefore share a common origin. The three alleles also differ by several single nucleotide polymorphisms, which represent somatic mutations accumulated over four centuries of asexual propagation.

Mutation at the tomato EXCESSIVE NUMBER OF FLORAL ORGANS (ENO) locus impairs floral meristem development, thus promoting an increased number of floral organs and fruit size

A novel tomato (Solanum lycopersicum L.) mutant affected in reproductive development, excessive number of floral organs (eno), is described in this study. The eno plants yielded flowers with a higher number of floral organs in the three innermost floral whorls and larger fruits than those found in wild-type plants. Scanning-electron microscopy study indicated that the rise in floral organ number and fruit size correlates with an increased size of floral meristem at early developmental stages. It has been reported that mutation at the FASCIATED (FAS) gene causes the development of flowers with supernumerary organs; however, complementation test and genetic mapping analyses proved that ENO is not an allele of the FAS locus. Furthermore, expression of WUSCHEL (SlWUS) and INHIBITOR OF MERISTEM ACTIVITY (IMA), the two main regulators of floral meristem activity in tomato, is altered in eno but not in fas flowers indicating that ENO could exert its function in the floral meristem independently of FAS. Interestingly, the eno mutation delayed the expression of IMA leading to a prolonged expression of SlWUS, which would explain the greater size of floral meristem. Taken together, results showed that ENO plays a significant role in the genetic pathway regulating tomato floral meristem development.

Two euAGAMOUS genes control C-function in Medicago truncatula.

C-function MADS-box transcription factors belong to the AGAMOUS (AG) lineage and specify both stamen and carpel identity and floral meristem determinacy. In core eudicots, the AG lineage is further divided into two branches, the euAG and PLE lineages. Functional analyses across flowering plants strongly support the idea that duplicated AG lineage genes have different degrees of subfunctionalization of the C-function. The legume Medicago truncatula contains three C-lineage genes in its genome: two euAG genes (MtAGa and MtAGb) and one PLENA-like gene (MtSHP). This species is therefore a good experimental system to study the effects of gene duplication within the AG subfamily. We have studied the respective functions of each euAG genes in M. truncatula employing expression analyses and reverse genetic approaches. Our results show that the M. truncatula euAG- and PLENA-like genes are an example of subfunctionalization as a result of a change in expression pattern. MtAGa and MtAGb are the only genes showing a full C-function activity, concomitant with their ancestral expression profile, early in the floral meristem, and in the third and fourth floral whorls during floral development. In contrast, MtSHP expression appears late during floral development suggesting it does not contribute significantly to the C-function. Furthermore, the redundant MtAGa and MtAGb paralogs have been retained which provides the overall dosage required to specify the C-function in M. truncatula.

Floral primordia-targeted ACS (1-aminocyclopropane-1-carboxylate synthase) expression in transgenic Cucumis melo implicates fine tuning of ethylene production mediating unisexual flower development.

Floral primordia-targeted expression of the ethylene biosynthetic gene, ACS , in melon suggests that differential timing and ethylene response thresholds combine to promote carpels, inhibit stamens, and prevent asexual bud formation. Typical angiosperm flowers produce both male and female reproductive organs. However, numerous species have evolved unisexuality. Melons (Cucumis melo L.) can produce varying combinations of male, female or bisexual flowers. Regardless of final sex, floral development begins with sequential initiation of all four floral whorls; unisexuality results from carpel or stamen primordia arrest regulated by the G and A loci, respectively. Ethylene, which promotes femaleness, is a key factor regulating sex expression. We sought to further understand the location, timing, level, and relationship to sex gene expression required for ethylene to promote carpel development or inhibit stamen development. Andromonoecious melons (GGaa) were transformed with the ethylene biosynthetic enzyme gene, ACS (1-aminocyclopropane-1-carboxylate synthase), targeted for expression in stamen and petal, or carpel and nectary, primordia using Arabidopsis APETALA3 (AP3) or CRABSCLAW (CRC) promoters, respectively. CRC::ACS plants did not exhibit altered sex phenotype. AP3::ACS melons showed increased femaleness manifested by gain of a bisexual-only phase not seen in wild type, decreased male buds and flowers, and loss of the initial male-only phase. In extreme cases, plants became phenotypically hermaphrodite, rather than andromonoecious. A reduced portion of buds progressed beyond initial whorl formation. Both the ACS transgene and exogenous ethylene reduced the expression of the native carpel-suppressing gene, G, while elevating expression of the stamen-suppressing gene, A. These results show ethylene-mediated regulation of key sex expression genes and suggest a mechanism by which temporally regulated ethylene production and differential ethylene response thresholds can promote carpels, inhibit stamens, and prevent the formation of asexual buds.

Floral development in Scutellaria pinnatifida (Lamiaceae): the ontogenetic basis for sepal reduction.

The Scutellaria is a Labiatae genus (subfamily Scutellarioideae) with a highly specialised floral structure. The genus is characterised by a peculiar two-lobed calyx with a projecting appendage, named the scutellum. Here, we present a detailed analysis of floral development, using epi-illumination light microscopy, to clarify open questions about its floral organisation. Floral whorls appeared in an acropetal sequence, with a marked temporal overlap of petal and stamen appearance. Organ appearance in each whorl proceeded unidirectionally from the abaxial to the adaxial side. Significant developmental features included the formation of the scutellum, reduction of sepal lobes and formation of a three-lobed nectary disc. Our study revealed that both loss of organ initiation and fusion of primordia are responsible for the reduction in sepal members in Scutellaria. The nectary structure was markedly different from most other studied Lamiaceae.

Novel functional roles for PERIANTHIA and SEUSS during floral organ identity specification, floral meristem termination, and gynoecial development.

The gynoecium is the female reproductive structure of angiosperm flowers. In Arabidopsis thaliana the gynoecium is composed of two carpels that are fused into a tube-like structure. As the gynoecial primordium arises from the floral meristem, a specialized meristematic structure, the carpel margin meristem (CMM), develops from portions of the medial gynoecial domain. The CMM is critical for reproductive competence because it gives rise to the ovules, the precursors of the seeds. Here we report a functional role for the transcription factor PERIANTHIA (PAN) in the development of the gynoecial medial domain and the formation of ovule primordia. This function of PAN is revealed in pan aintegumenta (ant) as well as seuss (seu) pan double mutants that form reduced numbers of ovules. Previously, PAN was identified as a regulator of perianth organ number and as a direct activator of AGAMOUS (AG) expression in floral whorl four. However, the seu pan double mutants display enhanced ectopic AG expression in developing sepals and the partial transformation of sepals to petals indicating a novel role for PAN in the repression of AG in floral whorl one. These results indicate that PAN functions as an activator or repressor of AG expression in a whorl-specific fashion. The seu pan double mutants also display enhanced floral indeterminacy, resulting in the formation of “fifth whorl” structures and disruption of WUSCHEL (WUS) expression patterns revealing a novel role for SEU in floral meristem termination.

Expression of paralogous SEP-, FUL-, AG- and STK-like MADS-box genes in wild-type and peloric Phalaenopsis flowers

The diverse flowers of Orchidaceae are the result of several major morphological transitions, among them the most studied is the differentiation of the inner median tepal into the labellum, a perianth organ key in pollinator attraction. Type A peloria lacking stamens and with ectopic labella in place of inner lateral tepals are useful for testing models on the genes specifying these organs by comparing their patterns of expression between wild-type and peloric flowers. Previous studies focused on DEFICIENS- and GLOBOSA-like MADS-box genes because of their conserved role in perianth and stamen development. The “orchid code” model summarizes this work and shows in Orchidaceae there are four paralogous lineages of DEFICIENS/AP3-like genes differentially expressed in each floral whorl. Experimental tests of this model showed the conserved, higher expression of genes from two specific DEF-like gene lineages is associated with labellum development. The present study tests whether eight MADS-box candidate SEP-, FUL-, AG-, and STK-like genes have been specifically duplicated in the Orchidaceae and are also differentially expressed in association with the distinct flower organs of Phalaenopsis hyb. “Athens.” The gene trees indicate orchid-specific duplications. In a way analogous to what is observed in labellum-specific DEF-like genes, a two-fold increase in the expression of SEP3-like gene PhaMADS7 was measured in the labellum-like inner lateral tepals of peloric flowers. The overlap between SEP3-like and DEF-like genes suggests both are associated with labellum specification and similar positional cues determine their domains of expression. In contrast, the uniform messenger levels of FUL-like genes suggest they are involved in the development of all organs and their expression in the ovary suggests cell differentiation starts before pollination. As previously reported AG-like and STK-like genes are exclusively expressed in gynostemium and ovary, however no evidence for transcriptional divergence was found in the stage investigated. Gene expression suggests a developmental regulatory system based on the combined activity of duplicate MADS-box genes. We discuss its feasibility based on documented protein interactions and patterns of expression.

The search for Pleiades in trait constellations: functional integration and phenotypic selection in the complex flowers of Morrenia brachystephana (Apocynaceae)

Pollinator-mediated natural selection on single traits, such as corolla tube or spur length, has been well documented. However, flower phenotypes are usually complex, and selection is expected to act on several traits that functionally interact rather than on a single isolated trait. Despite the fact that selection on complex phenotypes is expectedly widespread, multivariate selection modelling on such phenotypes still remains under-explored in plants. Species of the subfamily Asclepiadoideae (Apocynaceae) provide an opportunity to study such complex flower contrivances integrated by fine-scaled organs from disparate developmental origin. We studied the correlation structure among linear floral traits (i) by testing a priori morphological, functional or developmental hypotheses among traits and (ii) by exploring the organization of flower covariation, considering alternative expectations of modular organization or whole flower integration through conditional dependence analysis (CDA) and integration matrices. The phenotypic selection approach was applied to determine whether floral traits involved in the functioning of the pollination mechanism were affected by natural selection. Floral integration was low, suggesting that flowers are organized in more than just one correlation pleiad; our hypothetical functional correlation matrix was significantly correlated with the empirical matrix, and the CDA revealed three putative modules. Analyses of phenotypic selection showed significant linear and correlational gradients, lending support to expectations of functional interactions between floral traits. Significant correlational selection gradients found involved traits of different floral whorls, providing evidence for the existence of functional integration across developmental domains.

Deciphering the Physalis floridana Double-Layered-Lantern1 Mutant Provides Insights into Functional Divergence of the GLOBOSA Duplicates within the Solanaceae

Physalis spp. develop the "Chinese lantern" trait, also known as inflated calyx syndrome, that is a morphological novelty. Here, we identified the double-layered-lantern1 (doll1) mutant, a recessive and monofactorial mutation, in Physalis floridana; its corolla and androecium were transformed into the calyx and gynoecium, respectively. Two GLOBOSA-like MADS-box paralogous genes PFGLO1 and PFGLO2 were found in Physalis floridana, while the mutated phenotype was cosegregated with a large deletion harboring PFGLO1 and was complemented by the PFGLO1 genomic locus in transgenic plants, and severe PFGLO1 knockdowns phenocopied doll1. Thus, DOLL1 encodes the PFGLO1 protein and plays a primary role in determining corolla and androecium identity. However, specific PFGLO2 silencing showed no homeotic variation but rather affected pollen maturation. The two genes featured identical floral expression domains, but the encoding proteins shared 67% identity in sequences. PFGLO1 was localized in the nucleus when expressed in combination with a DEFICIENS homolog from Physalis floridana, whereas PFGLO2 was imported to the nucleus on its own. The two proteins were further found to have evolved different interacting partners and regulatory patterns, supporting the hypothesis that PFGLO2 is functionally separated from organ identity. Such a divergent pattern of duplicated GLO genes is unusual within the Solanaceae. Moreover, the phenotypes of the PFGLO1PFGLO2 double silencing mutants suggested that PFGLO2, through genetically interacting with PFGLO1, also exerts a role in the control of organ number and tip development of the second floral whorl. Our results, therefore, shed new light on the functional evolution of the duplicated GLO genes.

The tomato NAC transcription factor SlNAM2 is involved in flower-boundary morphogenesis

Being composed of several whorls of distinct floral organs, the flower is one of the most complex organs in the plant. As such, the formation and maintenance of boundaries that separate the meristem from the floral organ primordium and adjacent organs are critical for its normal development. In Arabidopsis, the miR164-regulated NAM genes play key roles in floral-boundary specification. By contrast, much less is known about floral-boundary establishment in the model crop tomato. It was found that the miR164-regulated NAM gene GOBLET is expressed in the floral meristem–organ boundaries and its loss-of-function mutant produces flowers with fused organs, indicating its requirement for tomato floral-boundary formation. It was found here that sly-miR164 targets the transcripts of three additional uncharacterized NAM genes in developing flowers. It is shown that, after floral-boundary initiation, the NAM gene Solyc03g115850 (SlNAM2) is expressed as stripes that mark the boundaries between sepals and between different floral whorls. Furthermore, ectopic accumulation of SlNAM2-encoding transcripts caused various growth-suppression and extraorgan phenotypes typically observed in plants over-expressing known boundary genes. Flower-specific silencing of sly-miR164-targeted NAM genes (AP1>>MIR164) caused defects in the separation of sepals and floral whorls indicating abnormal boundary specification. However, supplementing these NAM-deficient flowers with miR164-resistant SlNAM2 suppressed their fusion phenotypes and completely restored floral boundaries. Together, our results strongly suggest that SlNAM2 participates in the establishment of tomato flower whorl and sepal boundaries.

A monographic study of the Oliniaceae

Morphological variation within the Oliniaceae is discussed with reference to diagnostic characters used to delimit species and infraspecific taxa. The Oliniaceae is hereby considered to comprise a single genus, Olinia Thunb. with ten species assigned to two sections: sect. Olinia, and sect. Rochetiana. The two sections are distinguishable on the basis of the pattern of leaf venation, shape of leaves and petals. Section Olinia comprises O. capensis, O emarginata, O. micrantha, O. ventosa, O. radiata and O. vanguerioides; whereas section Rochetiana includes O. huillensis, O. rochetiana, O. ruandensis and O. usambarensis. O. huillensis Welw. shows infraspecific variation categorised as O. huillensis subsp. huillensis, O. huillensis subsp. burttdavii and O. huillensis subsp. discolor occurring mainly in southern Africa. Floral features provide useful and reliable distinguishing characters that consistently separate taxa at specific and infraspecific ranks. In particular, the following features are taxonomically useful: flowers which are organised into either tightly compact inflorescences or loose and open inflorescences, floral whorls comprising either four or five units, and variation in the shape and length of petal lobes. A full taxonomic account of Olinia over its total geographic distribution is provided with an identification key, descriptions, illustrations and distribution maps for all recognised taxa.

Novel members of the AGAMOUS LIKE 6 subfamily of MIKCC-type MADS-box genes in soybean

BackgroundThe classical (C) MIKC-type MADS-box transcription factors comprise one gene family that plays diverse roles in the flowering process ranging from floral initiation to the development of floral organs. Despite their importance in regulating developmental processes that impact crop yield, they remain largely unexplored in the major legume oilseed crop, soybean.ResultsWe identified 57 MIKCc-type transcription factors from soybean and determined the in silico gene expression profiles of the soybean MIKCc-type genes across different tissues. Our study implicates three MIKCc-type transcription factors as novel members of the AGAMOUS LIKE 6 (AGL6) subfamily of the MIKCC-type MADS-box genes, and we named this sister clade PsMADS3. While similar genes were identified in other legume species, poplar and grape, no such gene is represented in Arabidopsis thaliana or rice. RT-PCR analysis on these three soybean PsMADS3 genes during early floral initiation processes revealed their temporal expression similar to that of APETALA1, a gene known to function as a floral meristem identity gene. However, RNA in situ hybridisation showed that their spatial expression patterns are markedly different from those of APETALA1.ConclusionLegume flower development system differs from that in the model plant, Arabidopsis. There is an overlap in the initiation of different floral whorls in soybean, and inflorescent meristems can revert to leaf production depending on the environmental conditions. MIKCC-type MADS-box genes have been shown to play key regulatory roles in different stages of flower development. We identified members of the PsMADS3 sub-clade in legumes that show differential spatial expression during floral initiation, indicating their potential novel roles in the floral initiation process. The results from this study will contribute to a better understanding of legume-specific floral developmental processes.

The evolution of plant form

‘The evolution of plant form’ is volume 45 of Wiley-Blackwell's ‘Annual Plant Reviews’, and it provides an interdisciplinary analysis of this field, ranging from paleobotany to genomics. The first half of the book addresses morphological innovations in plant evolution by clade in an up-to-date phylogenetic context. An introductory chapter presents an overview of the characteristics of each major land plant group mapped onto a recent phylogenetic framework. It includes a detailed discussion of a selection of fossils in relation to characteristics not shown by any living groups, although the affinities allocated differ from treatments of other authors (e.g. Boyce, 2008; Harrison and Langdale 2008). The following five chapters present accounts of the morphology of each major land plant group, including bryophytes, lycophytes, ferns, gymnosperms and angiosperms. The chapter on bryophytes provides a good level of detail about liverworts, mosses and hornworts in the context of recent group-specific phylogenies. This is accompanied by some interesting speculation about character transitions within the bryophytes, and between the bryophytes and the vascular plants, with hypotheses about how they could be tested. The chapter on lycophytes presents an informative overview of morphological distinctions between the three major extant lycophyte lineages, whilst the one on ferns discusses the labile modularity of fern architecture in an explicit homology framework. Particular attention is paid to the evolution of embryos, shoots, leaves, and sporangium bearing structures. The chapter on gymnosperms provides an overview of the main organ systems in gymnosperms and discusses divergences in leaf and seed form between groups. The clonal analyses of Korn (2001, 2002) would have added to the discussion of shoot and leaf development here. The chapter on angiosperms addresses hypotheses relating to the origin of flowers from reproductive shoot systems, the innovation of the carpel, and the elaboration of floral symmetry in an integrated morphological and molecular context. The second half of the book moves into a consideration of genetic mechanisms which could have underpinned the radiation of land plant phyla. Chapter 7 presents an accessible and up-to-date explanation of QTL approaches which assesses the genetic basis of morphological diversity. It goes on to discuss genetic signatures of selection and how they can be statistically evaluated. A set of minimum requirements for determining a causal link between genes and character evolution is clearly identified, and some case studies from flowering plants are discussed in more detail. Chapter 8 gives an overview of plant genome evolution in relation to the generation of genetic diversity. It considers genome size within the land plants in relation to gene density and factors that influence genome evolution. The attributes of genomes sampled across the plant tree of life are discussed in relation to the amplification of gene families that may have been important in plant evolution. Chapter 9 presents three specific areas where evo-devo studies using reverse genetics have recently made an impact, namely the evolution of reproductive shoots, floral whorls and leaves. The discussion is informative and accessible. Chapter 10 considers the interplay between environment and development and its significance in plant evolution, and a final chapter pulls together the main themes addressed in the book, pointing out some areas for further future research. Strengths of this book are that the progression of topics covered by chapter is clear and logical, and the information covered is up-to-date and well integrated. I particularly liked the way that a discussion of fossil taxa is intercalated with considerations of living taxa throughout the book. I found some of the hypotheses of evolution interesting and innovative. In my view the book is more firmly rooted in a morphological and phylogenetic context than a developmental and genetic context, although the latter contributions were also clear and accessible. I thought that recent advances in developmental genetics could have allowed the authors to postulate specific testable hypotheses about some of the particular character changes distinguishing groups. In particular I felt that a consideration of mechanisms underpinning the innovation of the embryo, a vegetative sporophyte shoot, and branching would have added something, and these areas have been covered by recent reviews in the developmental literature (e.g. Harrison et al., 2010; Bosca et al., 2011; Domagalska and Leyser, 2011). Overall I felt that this book will be a great first port of call for those interested in morphological evolution and it will be useful at all levels from undergraduate onwards. It will be particularly useful for molecular and developmental biologists wishing to move into a comparative approach.

The corona of the daffodil Narcissus bulbocodium shares stamen‐like identity and is distinct from the orthodox floral whorls

The structural homology of the daffodil corona has remained a source of debate throughout the history of botany. Over the years it has been separately referred to as a modified petal stipule, stamen and tepal. Here we provide insights from anatomy and molecular studies to clarify the early developmental stages and position of corona initiation in Narcissus bulbocodium. We demonstrate that the corona initiates as six separate anlagen from hypanthial tissue between the stamens and perianth. Scanning electron microscope images and serial sections demonstrate that corona initiation occurs late in development, after the other floral whorls are fully developed. To define more precisely the identity of the floral structures, daffodil orthologues of the ABC floral organ identity genes were isolated and expression patterns were examined in perianth, stamens, carpel, hypanthial tube and corona tissue. Coupled with in situ hybridisation experiments, these analyses showed that the expression pattern of the C-class gene NbAGAMOUS in the corona is more similar to that of the stamens than that of the tepals. In combination, our results demonstrate that the corona of the daffodil N. bulbocodium exhibits stamen-like identity, develops independently from the orthodox floral whorls and is best interpreted as a late elaboration of the region between the petals and stamens associated with epigyny and the hypanthium.

Genetic analysis of ectopic growth suppression during planar growth of integuments mediated by the Arabidopsis AGC protein kinase UNICORN

BackgroundThe coordination of growth within a tissue layer is of critical importance for tissue morphogenesis. For example, cells within the epidermis undergo stereotypic cell divisions that are oriented along the plane of the layer (planar growth), thereby propagating the layered epidermal structure. Little is known about the developmental control that regulates such planar growth in plants. Recent evidence suggested that the Arabidopsis AGC VIII protein kinase UNICORN (UCN) maintains planar growth by suppressing the formation of ectopic multicellular protrusions in several floral tissues including integuments. In the current model UCN controls this process during integument development by directly interacting with the ABERRANT TESTA SHAPE (ATS) protein, a member of the KANADI (KAN) family of transcription factors, thereby repressing its activity. Here we report on the further characterization of the UCN mechanism.ResultsPhenotypic analysis of flowers of ucn-1 plants impaired in floral homeotic gene activity revealed that any of the four floral whorls could produce organs carrying ucn-1 protrusions. The ectopic outgrowths of ucn integuments did not accumulate detectable signals of the auxin and cytokinin reporters DR5rev::GFP and ARR5::GUS, respectively. Furthermore, wild-type and ucn-1 seedlings showed similarly strong callus formation upon in vitro culture on callus-inducing medium. We also show that ovules of ucn-1 plants carrying the dominant ats allele sk21-D exhibited more pronounced protrusion formation. Finally ovules of ucn-1 ett-1 double mutants and ucn-1 ett-1 arf4-1 triple mutants displayed an additive phenotype.ConclusionsThese data deepen the molecular insight into the UCN-mediated control of planar growth during integument development. The presented evidence indicates that UCN downstream signaling does not involve the control of auxin or cytokinin homeostasis. The results also reveal that UCN interacts with ATS independently of an ATS/ETT complex required for integument initiation and they further emphasize the necessity to balance UCN and ATS proteins during maintenance of planar growth in integuments.

Comparative Transcript Profiling of a Male Sterile Cybrid Pummelo and Its Fertile Type Revealed Altered Gene Expression Related to Flower Development

Male sterile and seedless characters are highly desired for citrus cultivar improvement. In our breeding program, a male sterile cybrid pummelo, which could be considered as a variant of male fertile pummelo, was produced by protoplast fusion. Herein, ecotopic stamen primordia initiation and development were detected in this male sterile cybrid pummelo. Histological studies revealed that the cybrid showed reduced petal development in size and width, and retarded stamen primordia development. Additionally, disorganized cell proliferation was also detected in stamen-like structures (fused to petals and/or carpel). To gain new insight into the underlying mechanism, we compared, by RNA-Seq analysis, the nuclear gene expression profiles of floral buds of the cybrid with that of fertile pummelo. Gene expression profiles which identified a large number of differentially expressed genes (DEGs) between the two lines were captured at both petal primordia and stamen primordia distinguishable stages. For example, nuclear genes involved in nucleic acid binding and response to hormone synthesis and metabolism, genes required for floral bud identification and expressed in particular floral whorls. Furthermore, in accordance with flower morphology of the cybrid, expression of PISTILLATA (PI) was reduced in stamen-like structures, even though it was restricted to correct floral whorls. Down-regulated expression of APETALA3 (AP3) coincided with that of PI. These finding indicated that, due to their whorl specific effects in flower development, citrus class-B MADS-box genes likely constituted ‘perfect targets’ for CMS retrograde signaling, and that dysfunctional mitochondria seemed to cause male sterile phenotype in the cybrid pummelo.

NO APICAL MERISTEM (MtNAM) regulates floral organ identity and lateral organ separation in Medicago truncatula

• The CUP-SHAPED COTYLEDON (CUC)/NO APICAL MERISTEM (NAM) family of genes control boundary formation and lateral organ separation, which is critical for proper leaf and flower patterning. However, most downstream targets of CUC/NAM genes remain unclear. • In a forward screen of the tobacco retrotransposon1 (Tnt1) insertion population in Medicago truncatula, we isolated a weak allele of the no-apical-meristem mutant mtnam-2. Meanwhile, we regenerated a mature plant from the null allele mtnam-1. These materials allowed us to extensively characterize the function of MtNAM and its downstream genes. • MtNAM is highly expressed in vegetative shoot buds and inflorescence apices, specifically at boundaries between the shoot apical meristem and leaf/flower primordia. Mature plants of the regenerated null allele and the weak allele display remarkable floral phenotypes: floral whorls and organ numbers are reduced and the floral organ identity is compromised. Microarray and quantitative RT-PCR analyses revealed that all classes of floral homeotic genes are down-regulated in mtnam mutants. Mutations in MtNAM also lead to fused cotyledons and leaflets of the compound leaf as well as a defective shoot apical meristem. • Our results revealed that MtNAM shares the role of CUC/NAM family genes in lateral organ separation and compound leaf development, and is also required for floral organ identity and development.

Evolution of petaloid sepals independent of shifts in B-class MADS box gene expression

Attractive petals are an integral component of animal-pollinated flowers and in many flowering plant species are restricted to the second floral whorl. Interestingly, multiple times during angiosperm evolution, petaloid characteristics have expanded to adjacent floral whorls or to extra-floral organs. Here, we investigate developmental characteristics of petaloid sepals in Rhodochiton atrosanguineum, a close relative of the model species Antirrhinum majus (snapdragon). We undertook this in two ways, first using scanning electron microscopy we investigate the micromorphology of petals and sepals, followed by expression studies of genes usually responsible for the formation of petaloid structures. From our data, we conclude that R. atrosanguineum petaloid sepals lack micromorphological characteristics of petals and that petaloid sepals did not evolve through regulatory evolution of B-class MADS box genes, which have been shown to specify second whorl petal identity in a number of model flowering plant species including snapdragon. These data, in conjunction with other studies, suggests multiple convergent pathways for the evolution of showy sepals.

Isolation and Characterization of an AGAMOUS-Like Gene from Magnolia wufengensis (Magnoliaceae)

The floral homeotic C function gene AGAMOUS (AG) plays crucial roles in Arabidopsis development by specifying stamen and carpel identity, repressing A-class genes, as well as regulating floral meristem determination. Although the function of AG homologs from other core eudicots appears highly conserved, the role of AG orthologs in the design of floral architecture in basal angiosperm remains unknown. We isolated and identified an AG ortholog from Magnolia wufengensis, a woody basal angiosperm belonging to the Magnoliaceae. Sequence and phylogenetic analyses revealed that it is a clade member of the euAG lineage, and hence, the gene is referred to as MAwuAG (M. wufengensis AGAMOUS). Moreover, two highly conserved motifs specific to C proteins, AG motifs I and II, are found in the C-terminal regions of the MAwuAG protein, but the N-terminal extensions that usually appear in euAG lineage members from eudicots were not found in MAwuAG. The cDNA has the first in-frame ATG immediately preceding the MADS domain. A semi-quantitative PCR analysis showed that the expression of MAwuAG was restricted to reproductive organs of stamens and carpels. The transgenic Arabidopsis containing 35S::MAwuAG displayed extremely early flowering, bigger stamens and carpels, and homeotic conversion of petals into staminoid organs, but ectopic expression of MAwuAG in the first whorls failed to convert the sepals into carpeloid structures that are usually observed in the overexpression transgenic Arabidopsis of AG orthologs from other core eudicots. In addition, the phenotype of the transgenic 35S::MAwuAG Arabidopsis revealed that the abscission of the outer three floral whorls (sepals, petals, and stamens) was inhibited.