<Emphasis Type="Italic">DORNRÖSCHEN-LIKE</Emphasis>, an AP2 gene,is necessary for stamen emergence in Arabidopsis

To identify novel genes in petal and stamen development, a genetic screen was carried out for enhancers of the unusual B class mutant pistillata-5 (pi-5). In pi-5 flowers, second whorl organs develop as sepals rather than petals, but third whorl stamens are normal. One pi-5 enhancer, dornröschen-like-2 (drnl-2), results in third whorl positions developing as filamentous organs. In addition to enhancing the pi-5 phenotype, drnl-2 mutants also exhibit a phenotype in a wild-type PI background. Although stamen primordia are morphologically visible during early stages of flower development, they fail to enlarge in drnl-2 mutants. DRNL, which encodes a single AP2 domain protein, is expressed in a dynamic pattern in the embryo, seedling, and flower. Analysis of both the drnl-2 mutant phenotype and the DRNL expression pattern in flowers suggests that DRNL plays a critical role in stamen emergence in Arabidopsis.     

Inflorescence and floral development in <Emphasis Type="Italic">Trochodendron aralioides</Emphasis> (Trochodendraceae)

In the early development of Trochodendron aralioides (Trochodendraceae) inflorescences lateral flowers are initiated after the appearance of the floral pherophylls (subtending bracts). The terminal flower is preceded by metaxyphylls and is initiated earlier than the uppermost lateral flowers of the botryoid inflorescence. Small scales (interpreted as rudimentary perianth organs) precede the stamens. These scales are more distinct in the terminal flower than in the lateral flowers. In the radially symmetrical terminal flower, small scales (or metaxyphylls) and stamens are initiated in a spiral during early development. At anthesis, stamen phyllotaxis appears irregular or approximately whorled as a result of the rapid elongation and irregular slight curvature of the stamen filaments which distorts the originally regular pattern. Finally, the numerous carpels arise simultaneously in a single whorl. It takes about 9 months for flowers to develop and the 2-year reproductive cycle of T. aralioides is typical of many trees. The floral development of T. aralioides is compared with that of other basal eudicots. The bottle-shaped, unicellular stigmatic papillae and long, decurrent stigma of basally united carpels are similar to those of the Buxales¸ suggesting a close relationship.     

Functional Conservation of an <Emphasis Type="Italic">AGAMOUS</Emphasis> Orthologous Gene Controlling Reproductive Organ Development in the Gymnosperm Species <Emphasis Type="Italic">Taxus chinensis</Emphasis> var. <Emphasis Type="Italic">mairei</Emphasis>

Arabidopsis AGAMOUS (AG) has roles in specifying reproductive organ (stamens and carpels) identity, floral meristem determinacy, and repression of A-function. To investigate possible roles of AG orthologous genes in gymnosperm species and evolution of C function, we isolated and identified AG orthologous gene TcAG from Taxus chinensis var. mairei (family Taxaceae, order Coniferales), a member of the last divergant lineage from higher Conifer that sisters to Gnetales. Sequence alignment and phylogenetic analysis grouped TcAG into the gymnosperm AG lineage. TcAG was expressed in both developing male and female cones, but there was no expression in juvenile leaves. Ectopic expression of TcAG in an Arabidopsis ag mutant produced flowers with the third whorl petaloid stamen and fourth whorl normal carpel, but failed to convert first whorl sepals into carpeloid organs and second whorl petals into stamenoid organs. A 35S::TcAG transgenic Arabidopsis ag mutant had very early flowering, and produced a misshapen inflorescence with a shortened floral axis. Our results suggest that establishment of the complete C-function occurred gradually during AG lineage evolution even in gymnosperms.     

FLORAL DEVELOPMENT OF HYDROCHARIS MORSUS-RANAE (HYDROCHARITACEAE)

In both male and female flowers of H. morsus-ranae the primordia of the floral appendages appear in an acropetal succession consisting of alternating trimerous whorls. In the male flower a whorl of sepals is followed by a whorl of petals, three whorls of stamens, and a whorl of filamentous staminodes. The mature androecial arrangement therefore consists of two antisepalous stamen whorls, an antipetalous whorl of stamens, and antipetalous staminodes. Shortly before anthesis, basal meristematic upgrowth between filaments of adjacent whorls produces paired stamens, joining Whorls 1 and 3, and Whorl 2 with the staminodial whorl. A central domelike structure develops between the closely appressed filaments of the inner stamen and staminodial whorl, giving the structure a lobed appearance. After petal inception in the female flower a whorl of antisepalous staminodes develop, each of which may bifurcate to form a pair of staminodes. During staminode development a girdling primordium arises by upgrowth at the periphery of the floral apex. The girdling primordium rapidly forms six gynoecial primordia, which then go on to produce six free styles with bifid stigmas. Intercalary meristem activity, below the point of floral appendage attachment, leads to the production of a syncarpous inferior ovary with six parietal placentae. The styles and carpels remain open along their ventral sutures. During the final stages of female floral development, several hundred ovules develop along the carpel walls, and three nectaries develop dorsally and basally on the three antipetalous styles.     

Ovule,fruit and seed development in Abolboda (Xyridaceae,Poales): implications for taxonomy and phylogeny

Xyridaceae belongs to the xyrid clade of Poales, but the phylogenetic position of the xyrid families is only weakly supported. Xyridaceae is divided into two subfamilies and five genera, the relationships of which remain unclear. The development of the ovule, fruit and seed of Abolboda spp. was studied to identify characteristics of taxonomic and phylogenetic value. All of the studied species share anatropous, tenuinucellate and bitegmic ovules with a micropyle formed by the inner and outer integuments, megagametophyte development of the Polygonum type, seeds with a tanniferous hypostase, a helobial and starchy endosperm and an undifferentiated embryo, seed coat derived from both integuments with a tanniferous tegmen and a micropylar operculum, and fruits with a parenchymatous endocarp and mesocarp and a sclerenchymatous exocarp. Most of the ovule and seed characteristics described for Abolboda are also present in Xyris and may represent a pattern for the family. Abolboda is distinguished by the ovule type, endosperm formation and the number of layers in the seed coat, in agreement with its classification in Abolbodoideae. The following characteristics link Xyridaceae to Eriocaulaceae and Mayacaceae, supporting the xyrid clade: tenuinucellate, bitegmic ovules; seeds with a tanniferous hypostase, a starchy endosperm and an undifferentiated embryo; and a seed coat with a tanniferous tegmen. A micropylar operculum in the seeds of Abolboda is described for the first time here and may represent a synapomorphy for the xyrids. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175 , 144–154.     

Floral meristem size and organ number correlation in <Emphasis Type="Italic">Eucryphia</Emphasis> (Cunoniaceae)

We present a comparative flower ontogenetic study in five species of the genus Eucryphia with the aim of testing whether differences in the organ number observed can be explained by changes in the meristematic size of floral meristem and floral organs. Species native to Oceania, viz. E. milliganii, E. lucida and E. moorei, have the smallest gynoecia with ca. 6 carpels, while the Chilean E. glutinosa and E. cordifolia present more than ten carpels. E. milliganii has the smallest flower with the lowest stamen number (ca. 50), while the other species produce around 200 stamens and more. Standardized measurements of meristematic sectors were taken in 49 developing flowers that were classified into three well-defined ontogenetic stages. Sizes of meristems varied significantly among species within each developmental stage as revealed by ANOVA analyses. Significant regressions between organ number and corresponding meristem size were consistent with the premise that a larger meristem size prior to organ initiation could be determining for a higher organ number. Flower organogenesis in Eucryphia also involves relevant meristem expansion while the organs are initiated, which results in a particular androecium patterning with a chaotic stamen arrangement. Meristem expansion also appears to be slower but more extensive in species with larger initial meristematic size, suggesting that flower phenotype can be determined in ontogeny by this heterochronic interplay of space and time.     

Isolation and characterization of the C-class MADS-box gene from the distylous pseudo-cereal <Emphasis Type="Italic">Fagopyrum esculentum</Emphasis>

In the model species Arabidopsis thaliana, the floral homeotic C-class gene AGAMOUS (AG) specifies reproductive organ (stamen and carpels) identity and floral meristem determinacy. Gene function analyses in other core eudicots species reveal functional conservation, subfunctionalization and function switch of the C-lineage in this clade. To identify the possible roles of AG-like genes in regulating floral development in distylous species with dimorphic flowers (pin and thrum) and the C function evolution, we isolated and identified an AG ortholog from Fagopyrum esculentum (buckwheat, Family Polygonaceae), an early diverging species of core eudicots preceding the rosids-asterids split. Protein sequence alignment and phylogenetic analysis grouped FaesAG into the euAG lineage. Expression analysis suggested that FaesAG expressed exclusively in developing stamens and gynoecium of pin and thrum flowers. Moreover, FaesAG expression reached a high level in both pin and thrum flowers at the time when the stamens were undergoing rapidly increased in size and microspore mother cells were in meiosis. FaesAG was able to substitute for the endogenous AG gene in specifying stamen and carpel identity and in an Arabidopsis ag-1 mutant. Ectopic expression of FaesAG led to very early flowering, and produced a misshapen inflorescence and abnormal flowers in which sepals had converted into carpels and petals were converted to stamens. Our results confirmed establishment of the complete C-function of the AG orthologous gene preceding the rosids-asterids split, despite the distinct floral traits present in early- and late-diverging lineages of core eudicot angiosperms.     

The floral organogenesis of <Emphasis Type="Italic">Koelreuteria bipinnata</Emphasis> and its variety <Emphasis Type="Italic">K. bipinnata</Emphasis> var. <Emphasis Type="Italic">integrifolia</Emphasis> (Sapindaceae): evidence of floral constraints on the evolution of monosymmetry

The floral organogenesis and anatomy of Koelreuteria bipinnata and its variety K. bipinnata var. integrifolia (Sapindaceae) has been investigated to clarify the identity of the two taxa in relation to other species of Koelreuteria, and to understand the shift to monosymmetry in the genus. Although the floral development is highly similar, we found a number of striking differences. Flowers arise in thyrses, with lateral branches forming cincinni of 5–9 flowers. Sepals initiate in a spiral sequence. Five petals arise unidirectionally alternating to the sepals. The last formed petal and one stamen between sepals 3 and 5 are strongly delayed, appearing as a common primordium, while this petal is completely suppressed in var. integrifoliola. Eight stamens initiate sequentially, differ in size and partly precede the development of petals. The gynoecium develops as a triangular primordium on which three carpellary lobes become demarcated simultaneously. Placentation is axile. Septal slits occurring within the style are interpreted as a deep-reaching non-nectariferous extension of the stigma. The massive, oblique disk with crenate apex develops in an extrastaminal position, but is interrupted on the radius of the lost petal. Floral developmental evidence supports variety K. bipinnata var. integrifolia rather than being synonym of K. bipinnata. Floral development is compared with K. paniculata and is discussed in the context of floral evolution of Sapindaceae. Our study demonstrates the importance of developmental shifts on floral evolution. The triangular gynoecium has a strong spatial impact in obliquely reorganizing the symmetry of the flower. It is demonstrated that spatial constraints of calyx and ovary are responsible for the reduction in one of the petals, two stamens and a shift in symmetry of the flower.     

Functional analysis of an <Emphasis Type="Italic">APETALA1</Emphasis>-like MADS box gene from <Emphasis Type="Italic">Eustoma grandiflorum</Emphasis> in regulating floral transition and formation

An Eustoma grandiflorum APETALA1 (EgAP1) gene showing high homology to the SQUA subfamily of MADS-box genes was isolated and characterized. EgAP1, containing a conserved euAP1 motif at the C-terminus, showed high sequence identity to Antirrhinum majus SQUAMOSA in the SQUA subfamily. EgAP1 mRNA was detected in the leaf and expressed significantly higher in young flower buds than in mature flower buds. In flowers, EgAP1 mRNA was strongly detected in sepal, weakly detected in petal and was absent in stamen and carpel. Transgenic Arabidopsis plants ectopically expressing EgAP1 flowered early and produced terminal flowers. In addition, the conversion of petals into stamen-like structures was also observed in 35S::EgAP1 flowers. 35S::EgAP1 was able to complement the ap1 flower defects by restoring the defect for sepal formation and significantly increasing second whorl petal production in Arabidopsis ap1 mutant plants. These results revealed that EgAP1 is the APETALA1 homolog in E. grandiflorum and that the function of EgAP1 is involved in floral induction and flower formation.     

Morphogenesis and Gene Mapping of <Emphasis Type="Italic">deformed interior floral organ 1</Emphasis> (<Emphasis Type="Italic">difo1</Emphasis>), a Novel Mutant Associated with Floral Organ Development in Rice

Floral organ identity and specific number directly affect anthesis habits, fertilization and grain yield. Here, we identified a deformed interior floral organ 1 (difo1) mutant from selfing progenies of indica cv. Zhonghui8015 (Zh8015) after ⁶⁰Co γ-ray treatment. Compared with the Zh8015 spikelet, the interior floral organs of the difo1 mutant present various numbers of stamens and stigmas, with no typical filament and no mature pollen grains. Most difo1 flowers exhibited an increased number of stigmas that were attached to the stamens and an intumescent ovule-like cell mass in addition to the ovary. Transverse sections of spikelets and scanning electron microscopy analysis revealed an indeterminate number of interior floral organs and abnormal early spikelet development for the difo1 mutant. Instead of the linear-shaped surface of wild-type stamens, difo1 displayed a glossy stamen surface resulting in immature stamens and complete sterility. In addition, the difo1 mutant exhibited delayed anthesis, rapid anthesis and non-extended stamens compared with wild type. Genetic analysis and gene mapping revealed that difo1 was controlled by a single recessive gene, which was fine-mapped to a 54-kb interval on the short arm of chromosome 4 between markers S22 and RM16439 harboring nine ORFs. Sequence analysis revealed that the mutant carried a single nucleotide deletion in its promoter region, which likely corresponded to the phenotype, in a C₂H₂-type zinc finger protein gene (LOC_Os04g08600). Moreover, qRT-PCR analysis showed a significantly down-regulated expression pattern for DIFO1 and many floral organ identity genes in the interior floral organs of difo1. DIFO1 is therefore an important floral organ development gene in rice, particularly with regard to interior organ meristem identity and floret primordium differentiation.     

Floral development and vasculature in Eriocaulon (Eriocaulaceae) provide insights into the evolution of Poales

Background and AimsFloral developmental studies are crucial for understanding the evolution of floral structures and sexual systems in angiosperms. Within the monocot order Poales, both subfamilies of Eriocaulaceae have unisexual flowers bearing unusual nectaries. Few previous studies have investigated floral development in subfamily Eriocauloideae, which includes the large, diverse and widespread genus Eriocaulon. To understand floral variation and the evolution of the androecium, gynoecium and floral nectaries of Eriocaulaceae, we analysed floral development and vasculature in Eriocaulon and compared it with that of subfamily Paepalanthoideae and the related family Xyridaceae in a phylogenetic context.MethodsThirteen species of Eriocaulon were studied. Developmental analysis was carried out using scanning electron microscopy, and vasculature analysis was carried out using light microscopy. Fresh material was also analysed using scanning electron microscopy with a cryo function. Character evolution was reconstructed over well-resolved phylogenies.Key ResultsPerianth reductions can occur due to delayed development that can also result in loss of the vascular bundles of the median sepals. Nectariferous petal glands cease development and remain vestigial in some species. In staminate flowers, the inner stamens can emerge before the outer ones, and carpels are transformed into nectariferous carpellodes. In pistillate flowers, stamens are reduced to staminodes and the gynoecium has dorsal stigmas.ConclusionsFloral morphology is highly diverse in Eriocaulon, as a result of fusion, reduction or loss of perianth parts. The nectariferous carpellodes of staminate flowers originated first in the ancestor of Eriocaulaceae; petal glands and nectariferous branches of pistillate flowers originated independently in Eriocaulaceae through transfer of function. We present a hypothesis of floral evolution for the family, illustrating a shift from bisexuality to unisexuality and the evolution of nectaries in a complex monocot family, which can contribute to future studies on reproductive biology and floral evolution in other groups.     

Isolation of 2-phenylethanol biosynthesis related genes and their relationship with 2-phenylethanol accumulation in <Emphasis Type="Italic">Rosa rugosa</Emphasis>

Rosa rugosa is a famous traditional flower in China used not only as a landscape plant but also as a time-honored and valuable aromatic plant. The natural aromatic substance 2-phenylethanol is the major indispensable ingredient of rose flower and rose essential oil. This study adopted R. rugosa ‘Tanghong’ as a test material to isolate 2-phenylethanol biosynthesis related genes RrAADC and RrPAR. The temporal and spatial expression patterns of the two genes in different flower developmental stages and floral organ parts were measured, and the relationship of 2-phenylethanol accumulation to RrAADC and RrPAR expression in R. rugosa was determined. The content of 2-phenylethanol in R. rugosa gradually increased with the degree of flower opening and peaked at the withering stage. The expression level of RrAADC gradually decreased with the degree of flower opening. Meanwhile, the expression level of RrPAR gradually increased from the budding stage to the half opening stage, rapidly decreased at the full opening stage, and slightly increased again toward the withering stage. The content of 2-phenylethanol was the highest in the petals, followed by that in the stamens and pistils. However, this compound was not detected in other parts of the fully opened flower. The expression level of RrAADC peaked in the stamens but was relatively low in the other parts. The expression level of RrPAR was the highest in the stamens and calyxes, followed by that in the pistils and receptacles, and the lowest in the petals and stalks. These results suggest that RrAADC and RrPAR coordinately regulate the biological synthesis of 2-phenylethanol in R. rugosa.     

Unusual alternations of floral organs in <Emphasis Type="Italic">Paeonia</Emphasis>: Structure and possible mechanism of formation

Morphological analysis of flowers was carried out in Paeonia L. cultivars. Some unusual alternations of floral organs were described: sepal-(petal-stamen) × n-carpel, where 2 ≤ n ≤ 4 (appearance of an additional zone of petal and stamen formation in the medial flower part). The identity of floral organs was not affected in the flowers with this unusual alternation. It was shown on the basis of mathematical simulation of the genes responsible for flower development that these alternations may be determined by increased pool of stem cells, which may lead to delayed termination of flower development.     

Reproductive biology of Abolboda pulchella and A. poarchon (Xyridaceae: Poales)

Background and Aims

Abolboda (Xyridaceae) belongs to the Poales, a predominantly wind-pollinated order whose phylogeny has been widely studied in recent years. The reproductive biology of Abolboda pulchella and A. poarchon was studied to determine the main pollination system of these species, providing the first experimental data on reproduction in the Xyridaceae.

Methods

A field study was conducted, including observations on the morphology and biology of the flowers, insect visits and pollinator behaviour. Experimental pollination treatments were performed to assess agamospermy, spontaneous self-pollination and self-compatibility. Pollination success was determined by pollen tube growth, and reproductive success was assessed by fruit- and seed-set.

Key Results

Abolboda pulchella and A. poarchon were pollinated by Apidae, Megachilidae and Halictidae bees. The floral resources were pollen and nectar that was produced by stylar appendages, an uncommom nectary type for monocotyledons. The species were self-compatible, and pollen tube growth from self-pollen was similar to that of cross-pollen. However, herkogamy prevented spontaneous selfing, rendering the plants dependent on the pollinator''s activity. There was no production of seeds by agamospermy.

Conclusions

Melittophily is the main pollination system of these two Abolboda species. Nectar production was first recorded here for Xyridaceae, and along with self-compatibility, herkogamy and bee pollination, is an informative characteristic that can be used in future phylogenetic analyses of the family as well as Poales.     

Expression pattern of ABA metabolic and signalling genes during floral development and fruit set in sweet cherry

Abscisic acid plays a crucial role in the regulation of fruit development and ripening, however, its role in the floral development and the fruit set is still unclear. In the present study, the ABA accumulation and the expression patterns of genes related to ABA metabolism and signalling in sweet cherry were investigated. The results showed that ABA accumulation increased and peaked at stage V in ovary, at stage VI in stamen, and in young fruit it peaked at 7 days after full bloom. The expression pattern of ABA synthetase PaNCED1 was consistent with the changes of ABA accumulation. Among four ABA degradation enzymes PaCYP707As, PaCYP707A4 was highly expressed in ovary, PaCYP707A1 was mainly in stamen, and PaCYP707A2 was in young fruit, and their expressions were reversed to the trend of PaNCED1. With regard to ABA signalling genes, among three ABA receptors PaPYLs, PaPYL2 and PaPYL3 were high expression genes in ovary and in young fruit with similar expression patterns, while PaPYL3 was the high expression gene in stamen. Within six PaPP2Cs, PaPP2C1/2/3 were highly expressed in ovary and young fruit, while PaPP2C3/4 were mainly in stamen. The six PaSnRK2s showed different expression patterns: PaSnRK2.1/2.2/2.4 were highly expressed in ovary and young fruit, while PaSnRK2.1/2.3 were highly expressed in stamen. In situ hybridization results showed that PaPYL3, PaPP2C3 and PaSnRK2.4 were expressed in seed, pulp and fruit peel during fruit set. In conclusion, ABA and its signaling may play an important role in the regulation of floral development and fruit set.     

Comparative floral anatomy of <Emphasis Type="Italic">Rhynchospora consanguinea</Emphasis> and <Emphasis Type="Italic">Rhynchospora pubera</Emphasis> (Cyperoideae,Cyperaceae)

Rhynchospora is one of the most species-rich genera of Cyperaceae and one of the few with entomophilous species. Considering the few anatomical studies of the genus, especially of the reproductive structures, this study comparatively analysed the anatomy of flowers of two representative species of Rhynchospora, R. consanguinea and R. pubera, including the floral vasculature and the anatomy of the style base, which is persistent with the fruit. Both species have congested inflorescences with light-coloured bracts and bisexual flowers, and phenolic idioblasts in the anthers and gynoecium, characteristics that suggest insect pollination. In R. consanguinea, the bisexual and the most proximal male flower has perianths, a new character state reported for the genus. The floral vasculature pattern is similar in both species, but differs from that previously described for Rhynchospora. In both species, there are two vascular bundles in the rachilla, which split into three receptacular bundles, the latter forming a vascular plexus where the bundles of stamens, gynoecium and ovule are connected. No lateral carpellary trace was observed, and the presence of the abaxial receptacular bundle was interpreted as a vestige of the tricarpellate ancestral condition. In the fruit of both species, the thickened style base (stylopodium) has a parenchyma with idioblasts containing phenolic compounds and idioblasts with helical or reticulate cell wall thickenings. The stylopodium is a homologous structure in the species of Rhynchospora and to other genera of Cyperoideae and evolved several times in the subfamily.     

Floral structure and development in Nartheciaceae (Dioscoreales), with special reference to ovary position and septal nectaries

We present a comparative study of the floral structure and development of Nartheciaceae, a small dioscorealean family consisting of five genera (Aletris, Lophiola, Metanarthecium, Narthecium, and Nietneria). A noticeable diversity existed in nine floral characters. Analyses of their respective character states in the light of a phylogenetic context revealed that the flowers of Nartheciaceae, whose plesiomorphies occur in Aletris and Metanarthecium, have evolved toward in all or part of Lophiola, Narthecium, and Nietneria: (1) loss of a perianth tube; (2) stamen insertion at the perianth base; (3) congenital carpel fusion; (4) loss of the septal nectaries; (5) unilocular style; (6) unfused lateral carpellary margins in the style; (7) flower with the median outer tepal on the abaxial side; (8) flower with moniliform hairs; and (9) flower with weak monosymmetry. We further found that, as the flowers developed, the ovary shifted its position from inferior to superior. As a whole, their structure changes suggest that the Nartheciaceae flowers have evolved in close association with pollination and seed dispersal. By considering inferior ovaries and the presence of septal nectaries as plesiomorphies of Nartheciaceae, we discussed evolution of the ovary position and septal nectaries in all the monocots.