Up and down: stamen movements in Ruta graveolens (Rutaceae) enhance both outcrossing and delayed selfing

Background and Aims

Stamen movements directly determine pollen fates and mating patterns by altering positions of female and male organs. However, the implications of such movements in terms of pollination are not well understood. Recently, complex patterns of stamen movements have been identified in Loasaceae, Parnassiaceae, Rutaceae and Tropaeolaceae. In this study the stamen movements in Ruta graveolens (Rutaceae) and their impact on pollination are determined.

Methods

Pollination effects of stamen movements were studied in Ruta graveolens, in which one-by-one uplifting and falling back is followed by simultaneous movement of all stamens in some flowers. Using 30 flowers, one stamen was manipulated either to be immobilized or to be allowed to move freely towards the centre of the flower but be prevented from falling back. Pollen loads on stigmas and ovule fertilization in flowers with or without simultaneous stamen movement were determined.

Results

Pollen removal decreased dramatically (P < 0·001) when the stamen was stopped from uplifting because its anther was seldom contacted by pollinators. When a stamen stayed at the flower''s centre, pollen removal of the next freely moved anther decreased significantly (P < 0·005) because of fewer touches by pollinators and quick leaving of pollinators that were discouraged by the empty anther. Simultaneous stamen movement occurred only in flowers with low pollen load on the stigma and the remaining pollen in anthers dropped onto stigma surfaces after stamens moved to the flower''s centre.

Conclusions

In R. graveolens pollen removal is promoted through one-by-one movement of the stamen, which presents pollen in doses to pollinators by successive uplifting of the stamen and avoids interference of two consecutively dehisced anthers by falling back of the former stamen before the next one moves into the flower''s centre. Simultaneous stamen movement at the end of anthesis probably reflects an adaptation for late-acting self-pollination.     

Paternity analysis-based inference of pollen dispersal patterns, male fecundity variation, and influence of flowering tree density and general flowering magnitude in two dipterocarp species

Background and Aims

Knowledge of pollen dispersal patterns and variation of fecundity is essential to understanding plant evolutionary processes and to formulating strategies to conserve forest genetic resources. Nevertheless, the pollen dispersal pattern of dipterocarp, main canopy tree species in palaeo-tropical forest remains unclear, and flowering intensity variation in the field suggests heterogeneity of fecundity.

Methods

Pollen dispersal patterns and male fecundity variation of Shorea leprosula and Shorea parvifolia ssp. parvifolia on Peninsular Malaysian were investigated during two general flowering seasons (2001 and 2002), using a neighbourhood model modified by including terms accounting for variation in male fecundity among individual trees to express heterogeneity in flowering.

Key Results

The pollen dispersal patterns of the two dipterocarp species were affected by differences in conspecific tree flowering density, and reductions in conspecific tree flowering density led to an increased selfing rate. Active pollen dispersal and a larger number of effective paternal parents were observed for both species in the season of greater magnitude of general flowering (2002).

Conclusions

The magnitude of general flowering, male fecundity variation, and distance between pollen donors and mother trees should be taken into account when attempting to predict the effects of management practices on the self-fertilization and genetic structure of key tree species in tropical forest, and also the sustainability of possible management strategies, especially selective logging regimes.     

Stigmatic fluid aids self-pollination in Roscoea debilis (Zingiberaceae): a new delayed selfing mechanism

Background and Aims

Delayed selfing is the predominant mode of autonomous self-pollination in flowering plants. However, few delayed selfing mechanisms have been documented. This research aims to explore a new delayed selfing mechanism induced by stigmatic fluid in Roscoea debilis, a small perennial ginger.

Methods

Floral biology and flower visitors were surveyed. The capacity of autonomous selfing was evaluated by pollinator exclusion. The timing of autonomous selfing was estimated by emasculation at different flowering stages. The number of seeds produced from insect-pollination was assessed by emasculation and exposure to pollinators in the natural population. The breeding system was also tested by pollination manipulations.

Key Results

Autonomous self-pollination occurred after flowers wilted. The stigmatic fluid formed a globule on the stigma on the third day of flowering. The enlarged globule seeped into the nearby pollen grains on the fourth flowering day, thus inducing pollen germination. Pollen tubes then elongated and penetrated the stigma. Hand-selfed flowers produced as many seeds as hand-crossed flowers. There was no significant difference in seed production between pollinator-excluded flowers and hand-selfed flowers. When emasculated flowers were exposed to pollinators, they produced significantly fewer seeds than intact flowers. Visits by effective pollinators were rare.

Conclusions

This study describes a new form of delayed autonomous self-pollination. As the predominant mechanism of sexual reproduction in R. debilis, delayed self-pollination ensures reproduction when pollinators are scarce.     

Do specialized flowers promote reproductive isolation? Realized pollination accuracy of three sympatric Pedicularis species

Background and Aims

Interest in pollinator-mediated evolutionary divergence of flower phenotype and speciation in plants has been at the core of plant evolutionary studies since Darwin. Specialized pollination is predicted to lead to reproductive isolation and promote speciation among sympatric species by promoting partitioning of (1) the species of pollinators used, (2) when pollinators are used, or (3) the sites of pollen placement. Here this last mechanism is investigated by observing the pollination accuracy of sympatric Pedicularis species (Orobanchacae).

Methods

Pollinator behaviour was observed on three species of Pedicularis (P. densispica, P. tricolor and P. dichotoma) in the Hengduan Mountains, south-west China. Using fluorescent powder and dyed pollen, the accuracy was assessed of stigma contact with, and pollen deposition on, pollinating bumble-bees, respectively.

Key Results

All three species of Pedicularis were pollinated by bumble-bees. It was found that the adaptive accuracy of female function was much higher than that of male function in all three flower species. Although peak pollen deposition corresponded to the optimal location on the pollinator (i.e. the site of stigma contact) for each species, substantial amounts of pollen were scattered over much of the bees'' bodies.

Conclusions

The Pedicularis species studied in the eastern Himalayan region did not conform with Grant''s ‘Pedicularis Model’ of mechanical reproductive isolation. The specialized flowers of this diverse group of plants seem unlikely to have increased the potential for reproductive isolation or influenced rates of speciation. It is suggested instead that the extreme species richness of the Pedicularis clade was generated in other ways and that specialized flowers and substantial pollination accuracy evolved as a response to selection generated by the diversity of co-occurring congeners.     

Reproductive success of non-rewarding Cypripedium japonicum benefits from low spatial dispersion pattern and asynchronous flowering

Background and Aims

Outcrossing animal-pollinated plants, particularly non-rewarding species, often experience pollinator limitation to reproduction. Pollinator visitation is affected by various factors, and it is hypothesized that reproduction in non-rewarding plants would benefit from low spatial flower abundance and asynchronous flowering. In order to test this hypothesis, the influence of spatial pattern and flowering phenology on male and female reproductive success (RS) was investigated in a non-rewarding orchid, Cypripedium japonicum, in central China over two flowering seasons.

Methods

The probabilities of intrafloral self-pollination and geitonogamy caused by pollinator behaviours were estimated from field observations. Pollinator limitation was evaluated by hand-pollination experiments. RS was surveyed in different spatial flower dispersal patterns and local flower densities. The effects of flowering phenological traits on RS were assessed by univariate and multivariate regression analyses.

Key Results

Hand-pollination experiments revealed that fruit production was strongly pollen limited throughout the entire reproductive season – over two seasons, 74·3 % of individuals set fruit following hand pollination, but only 5·2–7·7 % did so under natural conditions. Intrafloral self-pollination and geitonogamy within the potential clones might be rare. Both male and female fitness were substantially lower in clustered plants than in those growing singly. An increase in local conspecific flower density significantly and negatively influenced male RS, but had no effect on female RS. Phenotypic selection analysis indicated that individuals flowering earlier have the greatest probability of RS. Over 85 % of sampled flowering individuals had a flowering synchrony value >0·7; however, highly synchronous flowering was not advantageous for RS, as indicated by the negative directional selection differentials and gradients, and by the positive quadratic selection gradients.

Conclusions

These results support the hypothesis that, as a consequence of density-dependent selection, low spatio-temporal flower abundance is advantageous for attracting pollinators and for reproduction in natural populations of non-rewarding C. japonicum.Key words: Cypripedium japonicum, pollen limitation, spatial dispersal pattern, flowering synchrony, reproductive success, selection differential, selection gradient, deceptive flower     

Pollen dispersal and gene flow within and into a population of the alpine monocarpic plant Campanula thyrsoides

Background and Aims

Gene flow by seed and pollen largely shapes the genetic structure within and among plant populations. Seed dispersal is often strongly spatially restricted, making gene flow primarily dependent on pollen dispersal within and into populations. To understand distance-dependent pollination success, pollen dispersal and gene flow were studied within and into a population of the alpine monocarpic perennial Campanula thyrsoides.

Methods

A paternity analysis was performed on sampled seed families using microsatellites, genotyping 22 flowering adults and 331 germinated offspring to estimate gene flow, and pollen analogues were used to estimate pollen dispersal. The focal population was situated among 23 genetically differentiated populations on a subalpine mountain plateau (<10 km²) in central Switzerland.

Key Results

Paternity analysis assigned 110 offspring (33·2 %) to a specific pollen donor (i.e. ‘father’) in the focal population. Mean pollination distance was 17·4 m for these offspring, and the pollen dispersal curve based on positive LOD scores of all 331 offspring was strongly decreasing with distance. The paternal contribution from 20–35 offspring (6·0–10·5 %) originated outside the population, probably from nearby populations on the plateau. Multiple potential fathers were assigned to each of 186 offspring (56·2 %). The pollination distance to ‘mother’ plants was negatively affected by the mothers'' degree of spatial isolation in the population. Variability in male mating success was not related to the degree of isolation of father plants.

Conclusions

Pollen dispersal patterns within the C. thyrsoides population are affected by spatial positioning of flowering individuals and pollen dispersal may therefore contribute to the course of evolution of populations of this species. Pollen dispersal into the population was high but apparently not strong enough to prevent the previously described substantial among-population differentiation on the plateau, which may be due to the monocarpic perenniality of this species.     

The relative importance of architecture and resource competition in allocation to pollen and ovule number within inflorescences of Hosta ventricosa varies with the resource pools

Background and Aims

Allocation of resources to floral traits often declines distally within inflorescences in flowering plants. Architecture and resource competition have been proposed as underlying mechanisms. The aim of the present study is to assess the relative importance of resource competition and architectural effects in pollen and ovule production on racemes of Hosta ventricosa, an apomictic perennial herb.

Methods

Combinations of two defoliation treatments (intact and defoliated) and two fruit-set treatments (no-fruit and fruit) were created, and the roles of architecture and resource competition at each resource level were assessed.

Key Results

Pollen and ovule number per flower increased after defoliation, but pollen to ovule ratio per flower did not change. Pollen, ovules and the pollen to ovule ratio per flower declined distally on racemes at each resource level. In the intact treatment, fruit development of early flowers did not affect either pollen or ovule number of late flowers. In the defoliated treatment, fruit development of early flowers reduced both pollen and ovule numbers of late flowers due to over-compensation caused by defoliation. Late flowers on defoliated fruit racemes produced less pollen than intact fruit racemes but the same number of ovules; therefore, the reduction in pollen number was not caused by over-compensation. In addition, the fruit-set rate of early flowers during flowering was higher in intact racemes than in defoliated racemes.

Conclusions

In flowering plants, the relative importance of architecture and resource competition in allocation to pollen and ovules may vary with the resource pools or the overall resource availability of maternal plants.     

New views of tapetum ultrastructure and pollen exine development in Arabidopsis thaliana

Background and Aims

The Arabidopsis thaliana pollen cell wall is a complex structure consisting of an outer sporopollenin framework and lipid-rich coat, as well as an inner cellulosic wall. Although mutant analysis has been a useful tool to study pollen cell walls, the ultrastructure of the arabidopsis anther has proved to be challenging to preserve for electron microscopy.

Methods

In this work, high-pressure freezing/freeze substitution and transmission electron microscopy were used to examine the sequence of developmental events in the anther that lead to sporopollenin deposition to form the exine and the dramatic differentiation and death of the tapetum, which produces the pollen coat.

Key Results

Cryo-fixation revealed a new view of the interplay between sporophytic anther tissues and gametophytic microspores over the course of pollen development, especially with respect to the intact microspore/pollen wall and the continuous tapetum epithelium. These data reveal the ultrastructure of tapetosomes and elaioplasts, highly specialized tapetum organelles that accumulate pollen coat components. The tapetum and middle layer of the anther also remain intact into the tricellular pollen and late uninucleate microspore stages, respectively.

Conclusions

This high-quality structural information, interpreted in the context of recent functional studies, provides the groundwork for future mutant studies where tapetum and microspore ultrastructure is assessed.     

In the right place at the right time: Parnassia resolves the herkogamy dilemma by accurate repositioning of stamens and stigmas

Background and Aims

Spatial (herkogamy) and temporal (dichogamy) separation of pollen presentation and stigma receptivity have been interpreted as reducing interference between male and female functions in hermaphroditic flowers. However, spatial separation leads to a potential conflict: reduced pollination accuracy, where pollen may be placed in a location on the pollinator different from the point of stigma contact.

Methods

To understand better how herkogamous flowers resolve this conflict, a study was made of a subalpine herb, Parnassia epunctulata, the nectariferous flowers of which exhibit sequential anther dehiscence (staggered pollen presentation) and stamen movements; usually one newly dehisced anther is positioned each day over the central gynoecium, while the older stamens bend away from the central position.

Key Results

The open flowers were visited by a variety of pollinators, most of which were flies. Seed set was pollinator-dependent (bagged flowers set almost no seeds) and pollen-limited (manual pollination increased seed set over open pollination). Analyses of adaptive accuracy showed that coordinated stamen movements and style elongation (movement herkogamy) dramatically increased pollination accuracy. Specifically, dehiscing anthers and receptive stigmas were positioned accurately in the vertical and horizontal planes in relation to the opposite sexual structure and pollinator position. By contrast, the spatial correspondence between anthers and stigma was dramatically lower before the anthers dehisced and after stamens bent outwards, as well as before and after the period of stigmatic receptivity.

Conclusions

It is shown for the first time that a combination of movement herkogamy and dichogamy can maintain high pollination accuracy in flowers with generalized pollination. Staggered pollen and stigma presentation with spatial correspondence can both reduce sexual interference and improve pollination accuracy.     

Time-lapse imaging of self- and cross-pollinations in Brassica rapa

Background and Aims

Pollination is an important process in the life cycle of plants and is the first step in bringing together the male and female gametophytes for plant reproduction. While pollination has been studied for many years, accurate knowledge of the morphological aspects of this process is still far from complete. This study therefore focuses on a morphological characterization of pollination, using time-series image analysis of self- and cross-pollinations in Brassica rapa.

Methods

Time-lapse imaging of pollen behaviour during self- and cross-pollinations was recorded for 90 min, at 1 min intervals, using a stereoscopic microscope. Using time-series digital images of pollination, characteristic features of pollen behaviours during self- and cross-pollinations were studied.

Key Results

Pollen exhibited various behaviours in both self- and cross-pollinations, and these were classified into six representative patterns: germination, expansion, contraction, sudden contraction, pulsation and no change. It is noteworthy that in ‘contraction’ pollen grains shrunk within a short period of 30–50 min, and in ‘pulsation’ repeated expansion and contraction occurred with an interval of 10 min, suggesting that a dehydration system is operating in pollination. All of the six patterns were observed on an individual stigma with both self- and cross-pollinations, and the difference between self- and cross-pollinations was in the ratios of the different behaviours. With regard to water transport to and from pollen grains, this occurred in multiple steps, before, during and after hydration. Thus, pollination is regulated by a combination of multiple components of hydration, rehydration and dehydration systems.

Conclusions

Regulated hydration of pollen is a key process for both pollination and self-incompatibility, and this is achieved by a balanced complex of hydration, dehydration and nutrient supply to pollen grains from stigmatic papilla cells.     

Size-dependent gender modification in Lilium apertum (Liliaceae): does this species exhibit gender diphasy?

Background and Aims

Variation in the relative female and male reproductive success of flowering plants is widespread, despite the fundamental hermaphroditic condition of the majority of species. In many hermaphroditic populations, environmental conditions and their influence on development and size can influence the gender expression of individuals through the formation of hermaphroditic and unisexual flowers. This study investigates the hypothesis that the bulbous, animal-pollinated, perennial Lilium apertum (Liliaceae) exhibits a form of size-dependent gender modification known as gender diphasy, in which the sexual expression of individuals depends on their size, with plants often changing sex between seasons.

Methods

Variation in floral traits was examined in relation to their size using marked individuals in natural populations, and also under glasshouse conditions. Measurements were taken of the height, flower number, floral sex expression, flower size, flower biomass and pollen production of individuals over consecutive years between 2009 and 2012 in seven populations in south-west China.

Key Results

Flowers of L. apertum are either perfect (hermaphroditic) or staminate (male) and, in any given season, plants exhibit one of three sex phenotypes: only hermaphrodite flowers, a mixture of hermaphroditic and male flowers, or only male flowers. Transitions between each of these sex phenotypes were observed over consecutive years and were commonly size-dependent, particularly transitions from small plants bearing only male flowers to those that were taller with hermaphroditic flowers. Hermaphroditic flowers were significantly larger, heavier and produced more pollen than male flowers.

Conclusions

The results for L. apertum are consistent with the ‘size advantage hypothesis’ developed for animal species with sex change. The theory predicts that when individuals are small they should exhibit the sex for which the costs of reproduction are less, and this usually involves the male phase. L. apertum provides an example of gender diphasy, a rare sexual system in flowering plants.     

Effects of floral display size on male and female reproductive success in Mimulus ringens

Background and Aims

The number of flowers blooming simultaneously on a plant may have profound consequences for reproductive success. Large floral displays often attract more pollinator visits, increasing outcross pollen receipt. However, pollinators frequently probe more flowers in sequence on large displays, potentially increasing self-pollination and reducing pollen export per flower. To better understand how floral display size influences male and female fitness, we manipulated display phenotypes and then used paternity analysis to quantify siring success and selfing rates.

Methods

To facilitate unambiguous assignment of paternity, we established four replicate (cloned) arrays of Mimulus ringens, each consisting of genets with unique combinations of homozygous marker genotypes. In each array, we trimmed displays to two, four, eight or 16 flowers. When fruits ripened, we counted the number of seeds per fruit and assigned paternity to 1935 progeny.

Key Results

Siring success per flower declined sharply with increasing display size, while female success per flower did not vary with display. The rate of self-fertilization increased for large floral displays, but siring losses due to geitonogamous pollen discounting were much greater than siring gains through increased self-fertilization. As display size increased, each additional seed sired through geitonogamous self-pollination was associated with a loss of 9·7 seeds sired through outcrossing.

Conclusions

Although total fitness increased with floral display size, the marginal return on each additional flower declined steadily as display size increased. Therefore, a plant could maximize fitness by producing small displays over a long flowering period, rather than large displays over a brief flowering period.     

Genotypic differences in pod wall and seed growth relate to invertase activities and assimilate transport pathways in asparagus bean

Background and Aims

Coordination of sugar transport and metabolism between developing seeds and their enclosing fruit tissues is little understood. In this study the physiological mechanism is examined using two genotypes of asparagus bean (Vigna unguiculata ssp. sesquipedialis) differing in pod wall and seed growth rates. Pod growth dominates over seed growth in genotype ‘Zhijiang 121’ but not in ‘Zhijiang 282’ in which a ‘bulging pod’ phenotype is apparent from 8 d post-anthesis (dpa) onward.

Methods

Seed and pod wall growth rates and degree of pod-bulging were measured in the two genotypes together with assays of activities of sucrose-degrading enzymes and sugar content in pod wall and seed and evaluation of cellular pathways of phloem unloading in seed coat using a symplasmic fluorescent dye, 5(6)-carboxyfluorescein (CF).

Key Results

Activities of cell wall, cytoplasmic and vacuolar invertases (CWIN, CIN and VIN) were significantly smaller in pod walls of ‘282’ than in ‘121’ at 10 dpa onwards. Low INV activities were associated with weak pod wall growth of ‘282’. In seed coats, CF was confined within the vasculature in ‘282’ but moved beyond the vasculature in ‘121’, indicating apoplasmic and symplasmic phloem unloading, respectively. Higher CWIN activity in ‘282’ seed coats at 6–8 dpa correlated with high hexose concentration in embryos and enhanced early seed growth. However, CWIN activity in ‘282’ decreased significantly compared with ‘121’ from 10 dpa onwards, coinciding with earlier commencement of nuclei endoreduplication in their embryos.

Conclusions

The study shows genotypic differences between ‘bulging pod’ and ‘non-bulging’ phenotypes of asparagus bean in sucrose metabolism in relation to the pathway of phloem unloading in developing seed coats, and to pod and seed growth. Low INV activity in pod wall corresponds to its shortened and weak growth period; by contrast, the apoplasmic path in the seed coat is associated with high CWIN activity and strong early seed growth.     

Floral closure induced by pollination in gynodioecious Cyananthus delavayi (Campanulaceae): effects of pollen load and type, floral morph and fitness consequences

Background and aims

Pollination-induced floral changes, which have been widely documented in flowering plants, have been assumed to enhance the plant''s reproductive success. However, our understanding of the causes and consequences of these changes is still limited. Using an alpine gynodioecious species, Cyananthus delavayi, we investigated the factors affecting floral closure and estimated the fitness consequences of floral closure.

Methods

The timings of floral closure and fertilization were determined. The effects of pollen load, pollen type (cross- or self-pollen) and floral morph (female or perfect flower) on the occurrence of floral closure were examined. Ovule fertilization and seed production were examined to investigate the causes and consequences of floral closure. Flowers were manipulated to prevent closing to detect potential benefits for female fitness.

Key Results

Floral closure, which could be induced by a very low pollen load, occurred within 4–7 h after pollination, immediately following fertilization. The proportion of closed flowers was influenced by pollen load and floral morph, but not by pollen type. Floral closure was more likely to occur in flowers with a higher proportion of fertilized ovules, but there was no significant difference in seed production between closed and open flowers. Those flowers in which closure was induced by natural pollination had low fruit set and seed production. Additionally, seed production was not influenced by closing-prevented manipulation when sufficient pollen deposition was received.

Conclusions

The occurrence of floral closure may be determined by the proportion of fertilized ovules, but this response can be too sensitive to ensure sufficient pollen deposition and can, to some extent, lead to a cost in female fitness. These results implied that the control of floral receptivity by the recipient flowers does not lead to an optimal fitness gain in C. delavayi.     

A genetic resource for early-morning flowering trait of wild rice Oryza officinalis to mitigate high temperature-induced spikelet sterility at anthesis

Background and Aims

High temperatures over 32–36 °C at anthesis induce spikelet sterility in rice. The use of a germplasm with an early-morning flowering (EMF) trait has been hypothesized as a way of avoiding this problem. In this study, the effect of the EMF trait on avoiding high temperature-induced sterility at anthesis by flowering at a cooler temperature in the early morning was evaluated.

Methods

The EMF trait was introgressed from wild rice (Oryza officinalis) into the rice cultivar ‘Koshihikari’ (O. sativa). First, spikelets of the EMF line and Koshihikari were subjected to rising temperatures during the daytime in the greenhouse to test for differences in spikelet sterility. Secondly, spikelets of both plants were exposed to 26, 34 and 38 °C at anthesis and to 38 °C beginning at least 1 h after flowering, in the growth chambers at 70 % relative humidity, to test for differences in tolerance to high temperatures.

Key Results

Spikelets of the EMF line started and completed flowering a few hours earlier than Koshihikari. In a greenhouse experiment, spikelets of Koshihikari opened after the air temperature reached 35 °C, but those of the EMF line could open at cooler temperatures. Under these conditions, spikelet sterility significantly increased in Koshihikari, but did not in the EMF line. The number of sterile spikelets increased as their flowering time was delayed in Koshihikari. Furthermore, the chamber experiments revealed that 60 % of the spikelets from both lines were sterile when exposed to 38 °C at anthesis, indicating that tolerance of high temperature was similar in both genotypes.

Conclusions

Reduced sterility in the EMF line subjected to rising temperatures at anthesis in the greenhouse was attributed to an earlier flowering time compared with Koshihikari. The EMF trait of wild rice is effective in mitigating anticipated yield loss due to global warming by escaping high-temperature stress at anthesis during the daytime.     

Profiling and functional classification of esterases in olive (Olea europaea) pollen during germination

Background and Aims

A pollen grain contains a number of esterases, many of which are released upon contact with the stigma surface. However, the identity and function of most of these esterases remain unknown. In this work, esterases from olive pollen during its germination were identifided and functionally characterized.

Methods

The esterolytic capacity of olive (Olea europaea) pollen was examined using in vitro and in-gel enzymatic assays with different enzyme substrates. The functional analysis of pollen esterases was achieved by inhibition assays by using specific inhibitors. The cellular localization of esterase activities was performed using histochemical methods.

Key Results

Olive pollen showed high levels of non-specific esterase activity, which remained steady after hydration and germination. Up to 20 esterolytic bands were identified on polyacrylamide gels. All the inhibitors decreased pollen germinability, but only diisopropyl fluorophosphate (DIFP) hampered pollen tube growth. Non-specific esterase activity is localized on the surface of oil bodies (OBs) and small vesicles, in the pollen intine and in the callose layer of the pollen tube wall. Acetylcholinesterase (AChE) activity was mostly observed in the apertures, exine and pollen coat, and attached to the pollen tube wall surface and to small cytoplasmic vesicles.

Conclusions

In this work, for the first time a systematic functional characterization of esterase enzymes in pollen from a plant species with wet stigma has been carried out. Olive pollen esterases belong to four different functional groups: carboxylesterases, acetylesterases, AChEs and lipases. The cellular localization of esterase activity indicates that the intine is a putative storage site for esterolytic enzymes in olive pollen. Based on inhibition assays and cellular localization of enzymatic activities, it can be concluded that these enzymes are likely to be involved in pollen germination, and pollen tube growth and penetration of the stigma.     

Dichogamy correlates with outcrossing rate and defines the selfing syndrome in the mixed-mating genus Collinsia

Background and Aims

How and why plants evolve to become selfing is a long-standing evolutionary puzzle. The transition from outcrossing to highly selfing is less well understood in self-compatible (SC) mixed-mating (MM) species where potentially subtle interactions between floral phenotypes and the environment are at play. We examined floral morphological and developmental traits across an entire SC MM genus, Collinsia, to determine which, if any, predict potential autonomous selfing ability when pollinators are absent (AS) and actual selfing rates in the wild, s_m, and to best define the selfing syndrome for this clade.

Methods

Using polymorphic microsatellite markers, we obtained 30 population-level estimates of s_m across 19 Collinsia taxa. Species grand means for the timing of herkogamy (stigma–anther contact) and dichogamy (stigmatic receptivity, SR), AS, floral size, longevity and their genetic correlations were quantified for 22 taxa.

Key Results

Species fell into discrete selfing and outcrossing groups based on floral traits. Loss of dichogamy defines Collinsia''s selfing syndrome. Floral size, longevity and herkogamy also differ significantly between these groups. Most taxa have high AS rates (>80 %), but AS is uncorrelated with any measured trait. In contrast, s_m is significantly correlated only with SR. High variance in s_m was observed in the two groups.

Conclusions

Collinsia species exhibit clear morphological and developmental traits diagnostic of ‘selfing’ or ‘outcrossing’ groups. However, many species in both the ‘selfing’ and the ‘outcrossing’ groups were MM, pointing to the critical influence of the pollination environment, the timing of AS and outcross pollen prepotency on s_m. Flower size is a poor predictor of Collinsia species'' field selfing rates and this result may apply to many SC species. Assessment of the variation in the pollination environment, which can increase selfing rates in more ‘outcrossing’ species but can also decrease selfing rates in more ‘selfing’ species, is critical to understanding mating system evolution of SC MM taxa.     

Molecular control of seasonal flowering in rice,arabidopsis and temperate cereals

Background

Rice (Oryza sativa) and Arabidopsis thaliana have been widely used as model systems to understand how plants control flowering time in response to photoperiod and cold exposure. Extensive research has resulted in the isolation of several regulatory genes involved in flowering and for them to be organized into a molecular network responsive to environmental cues. When plants are exposed to favourable conditions, the network activates expression of florigenic proteins that are transported to the shoot apical meristem where they drive developmental reprogramming of a population of meristematic cells. Several regulatory factors are evolutionarily conserved between rice and arabidopsis. However, other pathways have evolved independently and confer specific characteristics to flowering responses.

Scope

This review summarizes recent knowledge on the molecular mechanisms regulating daylength perception and flowering time control in arabidopsis and rice. Similarities and differences are discussed between the regulatory networks of the two species and they are compared with the regulatory networks of temperate cereals, which are evolutionarily more similar to rice but have evolved in regions where exposure to low temperatures is crucial to confer competence to flower. Finally, the role of flowering time genes in expansion of rice cultivation to Northern latitudes is discussed.

Conclusions

Understanding the mechanisms involved in photoperiodic flowering and comparing the regulatory networks of dicots and monocots has revealed how plants respond to environmental cues and adapt to seasonal changes. The molecular architecture of such regulation shows striking similarities across diverse species. However, integration of specific pathways on a basal scheme is essential for adaptation to different environments. Artificial manipulation of flowering time by means of natural genetic resources is essential for expanding the cultivation of cereals across different environments.     

Histological and Micro-CT Evidence of Stigmatic Rostellum Receptivity Promoting Auto-Pollination in the Madagascan Orchid Bulbophyllum bicoloratum

Background

The rostellum, a projecting part of the gynostemium in orchid flowers, separates the anther(s) from the stigma and thus commonly prevents auto-pollination. Nonetheless, as a modified (usually distal) portion of the median stigma lobe, the rostellum has been frequently invoked of having re-gained a stigmatic function in rare cases of orchid auto-pollination. Here it is shown that a newly discovered selfing variant of Madagascan Bulbophyllum bicoloratum has evolved a modified rostellum allowing the penetration of pollen tubes from in situ pollinia.

Methods

Gynostemium micro-morphology and anatomy of selfing and outcrossing variants of B . bicoloratum was studied by using light and scanning electron microscopy and histological sections. Pollen tube growth in the selfing variant was further observed via X-ray computed microtomography (micro-CT), providing 3D reconstructions of floral tissues at a micron scale.

Findings

Selfing variants possess a suberect (‘displaced’) rostellum rather than the conventional, erect type. Very early in anthesis, the pollinia of selfers are released from the anther and slide down onto the suberect rostellum, where pollen tube growth preferentially occurs through the non-vascularized, i.e. rear (adaxial) and (semi-) lateral parts. This penetrated tissue is comprised of a thin layer of elongate and loosely arranged cells, embedded in stigmatic exudates, as also observed in the stigmatic cavity of both selfing and outcrossing variants.

Conclusions

Our results provide the first solid evidence of a stigmatic function for the rostellum in orchid flowers, thereby demonstrating for the first time the feasibility of the micro-CT technique for accurately visualizing pollen tube growth in flowering plants. Rostellum receptivity in B . bicoloratum probably uniquely evolved as an adaptation for reproductive assurance from an outcrossing ancestor possessing an erect (non-receptive) rostellum. These findings open up new avenues in the investigation of an organ that apparently re-gained its ‘primordial function’ of being penetrated by pollen tubes.     

Function and evolution of sterile sex organs in cryptically dioecious Petasites tricholobus (Asteraceae)

Background and Aims

Why are sterile anthers and carpels retained in some flowering plants, given their likely costs? To address this question, a cryptically dioecious species, Petasites tricholobus, in which male and female plants each have two floret types that appear pistillate and hermaphroditic, was studied. The aim was to understand the function of sterile hermaphroditic florets in females. In addition, the first examination of functions of sterile female structures in male plants was conducted in the hermaphroditic florets on males of this species. These female structures are exceptionally large in this species despite being sterile.

Methods

Differences in floret morphology between the sex morphs were documented and the possible functions of sterile sex organs investigated using manipulative experiments. Tests were carried out to find out if sterile female structures in male florets attract pollinators and if they aid in pollen dispersal, also to find out if the presence and quantity of sterile hermaphroditic florets in females increase pollinator attraction and reproductive success. To investigate what floret types provide nectar, all types of florets were examined under a scanning electron microscope to search for nectaries.

Key Results

The sterile female structures in male florets did not increase pollinator visits but were essential to secondary pollen presentation, which significantly enhanced pollen dispersal. Sterile pistillate florets on male plants did not contribute to floral display and disappeared in nearly half of the male plants. The sterile hermaphroditic florets on female plants attracted pollinators by producing nectar and enhanced seed production.

Conclusions

The presence of female structures in male florets and hermaphroditic florets on female plants is adaptive despite being sterile, and may be evolutionarily stable. However, the pistillate florets on male plants appear non-adaptive and are presumably in decline. Differential fates of the sterile sex organs in the species are determined by both the historical constraints and the ecological functions.