Is self-incompatibility a reproductive barrier for hybridization in a sympatric species?

PREMISE: Barriers at different reproductive stages contribute to reproductive isolation. Self-incompatibility (SI) systems that prevent self-pollination could also act to control interspecific pollination and contribute to reproductive isolation, preventing hybridization. Here we evaluated whether SI contributes to reproductive isolation among four co-occurring Opuntia species that flower at similar times and may hybridize with each other. METHODS: We assessed whether Opuntia cantabrigiensis, O. robusta, O. streptacantha, and O. tomentosa, were self-compatible and formed hybrid seeds in five manipulation treatments to achieve self-pollination, intraspecific cross-pollination, open pollination (control), interspecific crosses or apomixis, then recorded flowering phenology and synchrony. RESULTS: All species flowered in the spring with a degree of synchrony, so that two pairs of species were predisposed to interspecific pollination (O. cantabrigiensis with O. robusta, O. streptacantha with O. tomentosa). All species had distinct reproductive systems: Opuntia cantabrigiensis is self-incompatible and did not produce hybrid seeds as an interspecific pollen recipient; O. robusta is a dioecious species, which formed a low proportion of hybrid seeds; O. streptacantha and O. tomentosa are self-compatible and produced hybrid seeds. CONCLUSIONS: Opuntia cantabrigiensis had a strong pollen-pistil barrier, likely due to its self-incompatibility. Opuntia robusta, the dioecious species, is an obligate outcrosser and probably partially lost its ability to prevent interspecific pollen germination. Given that the self-compatible species can set hybrid seeds, we conclude that pollen-pistil interaction and high flowering synchrony represent weak barriers; whether reproductive isolation occurs later in their life cycle (e.g., germination or seedling survival) needs to be determined.

A Long-Lived Alpine Perennial Advances Flowering under Warmer Conditions but Not Enough to Maintain Reproductive Success.

AbstractAssessing whether phenological shifts in response to climate change confer a fitness advantage requires investigating the relationships among phenology, fitness, and environmental drivers of selection. Despite widely documented advancements in phenology with warming climate, we lack empirical estimates of how selection on phenology varies in response to continuous climate drivers or how phenological shifts in response to warming conditions affect fitness. We leverage an unusual long-term dataset with repeated, individual measurements of phenology and reproduction in a long-lived alpine plant. We analyze phenotypic plasticity in flowering phenology in relation to two climate drivers, snowmelt timing and growing degree days (GDDs). Plants flower earlier with increased GDDs and earlier snowmelt, and directional selection also favors earlier flowering under these conditions. However, reproduction still declines with warming and early snowmelt, even when flowering is early. Furthermore, the steepness of this reproductive decline increases dramatically with warming conditions, resulting in very little fruit production regardless of flowering time once GDDs exceed approximately 225 degree days or snowmelt occurs before May 15. Even though advancing phenology confers a fitness advantage relative to stasis, these shifts are insufficient to maintain reproduction under warming, highlighting limits to the potential benefits of phenological plasticity under climate change.

HD-Zip proteins modify floral structures for self-pollination in tomato.

Cleistogamy is a type of self-pollination that relies on the formation of a stigma-enclosing floral structure. We identify three homeodomain-leucine zipper IV (HD-Zip IV) genes that coordinately promote the formation of interlocking trichomes at the anther margin to unite neighboring anthers, generating a closed anther cone and cleistogamy (flower morphology necessitating strict self-pollination). These HD-Zip IV genes also control style length by regulating the transition from cell division to endoreduplication. The expression of these HD-Zip IV genes and their downstream gene, Style 2.1, was sequentially modified to shape the cleistogamy morphology during tomato evolution and domestication. Our results provide insights into the molecular basis of cleistogamy in modern tomato and suggest targets for improving fruit set and preventing pollen contamination in genetically modified crops.

Chinese cherry CpMYB44-CpSPDS2 module regulates spermidine content and florescence in tobacco.

The flower bud differentiation plays a crucial role in cherry yield and quality. In a preliminary study, we revealed the promotion of spermidine (Spd) in bud differentiation and quality. However, the molecular mechanism underlying Spd regulating cherry bud differentiation remains unclear. To address this research gap, we cloned CpSPDS2, a gene that encodes Spd synthase and is highly expressed in whole flowers and pistils of the Chinese cherry (cv. 'Manaohong'). Furthermore, an overexpression vector with this gene was constructed to transform tobacco plants. The findings demonstrated that transgenic lines exhibited higher Spd content, an earlier flowering time by 6 d, and more lateral buds and flowers than wild-type lines. Additionally, yeast one-hybrid assays and two-luciferase experiments confirmed that the R2R3-MYB transcription factor (CpMYB44) directly binds to and activates the CpSPDS2 promoter transcription. It is indicated that CpMYB44 promotes Spd accumulation via regulating CpSPDS2 expression, thus accelerating the flower growth. This research provides a basis for resolving the molecular mechanism of CpSPDS2 involved in cherry bud differentiation.

Flower opening dynamics, pollen-ovule ratio, stigma receptivity and stigmatic pollen germination (in-vivo) in Chaenomeles speciosa (Sweet) Nakai.

Although Chaenomeles is widely used in horticulture, traditional Chinese medicine and landscape greening, insufficient research has hindered its breeding and seed selection. This study investigated the floral phenology, floral organ characteristics, palynology, and breeding systems of Chaenomeles speciosa (Sweet) Nakai. The floral characteristics of C. speciosa were observed both visually and stereoscopically. The microstructures of the flower organs were observed using scanning electron microscopy. Pollen stainability was determined using triphenyl tetrazolium chloride staining. Stigma receptivity was determined using the benzidine-H2O2 method and the post-artificial pollination pollen germination method. The breeding system was assessed based on the outcrossing index and pollen-ovule ratio. The flowers of C. speciosa were bisexual with a flowering period from March to April. The flowering periods of single flowers ranged from 8 to 19 d, and those of single plants lasted 18-20 d. The anthers were cylindrical, with the base attached to the filament, and were split longitudinally to release pollen. The flower had five styles, with a connate base. The ovaries had five carpels and five compartments. The inverted ovules were arranged in two rows on the placental axis. The stigma of C. speciosa was dry and had many papillary protrusions. In the early flowering stage (1-2 d of flowering), the pollen exhibited high stainability (up to 84.24%), but all stainability was lost at 7 d of flowering. Storage at - 20 °C effectively delayed pollen inactivation. The stigma receptivity of C. speciosa lasted for approximately 7 days, and the breeding system was classified as outcrossing with partial self-compatibility.

Exploring the molecular regulation of vernalization-induced flowering synchrony in Arabidopsis.

Many plant populations exhibit synchronous flowering, which can be advantageous in plant reproduction. However, molecular mechanisms underlying flowering synchrony remain poorly understood. We studied the role of known vernalization-response and flower-promoting pathways in facilitating synchronized flowering in Arabidopsis thaliana. Using the vernalization-responsive Col-FRI genotype, we experimentally varied germination dates and daylength among individuals to test flowering synchrony in field and controlled environments. We assessed the activity of flowering regulation pathways by measuring gene expression across leaves produced at different time points during development and through a mutant analysis. We observed flowering synchrony across germination cohorts in both environments and discovered a previously unknown process where flower-promoting and repressing signals are differentially regulated between leaves that developed under different environmental conditions. We hypothesized this mechanism may underlie synchronization. However, our experiments demonstrated that signals originating from sources other than leaves must also play a pivotal role in synchronizing flowering time, especially in germination cohorts with prolonged growth before vernalization. Our results suggest flowering synchrony is promoted by a plant-wide integration of flowering signals across leaves and among organs. To summarize our findings, we propose a new conceptual model of vernalization-induced flowering synchrony and provide suggestions for future research in this field.

Sorghum SbGhd7 is a major regulator of floral transition and directly represses genes crucial for flowering activation.

Molecular genetic understanding of flowering time regulation is crucial for sorghum development. GRAIN NUMBER, PLANT HEIGHT AND HEADING DATE 7 (SbGhd7) is one of the six classical loci conferring photoperiod sensitivity of sorghum flowering. However, its functions remain poorly studied. The molecular functions of SbGhd7 were characterized. The gene regulatory network controlled by SbGhd7 was constructed and validated. The biological roles of SbGhd7 and its major targets were studied. SbGhd7 overexpression (OE) completely prevented sorghum flowering. Additionally, we show that SbGhd7 is a major negative regulator of flowering, binding to the promoter motif TGAATG(A/T)(A/T/C) and repressing transcription of the major florigen FLOWERING LOCUS T 10 (SbFT10) and floral activators EARLY HEADING DATE (SbEhd1), FLAVIN-BINDING, KELCH REPEAT, F-BOX1 (SbFKF1) and EARLY FLOWERING 3 (SbELF3). Reinforcing the direct effect of SbGhd7, SbEhd1 OE activated the promoters of three functional florigens (SbFT1, SbFT8 and SbFT10), dramatically accelerating flowering. Our studies demonstrate that SbGhd7 is a major repressor of sorghum flowering by directly and indirectly targeting genes for flowering activation. The mechanism appears ancient. Our study extends the current model of floral transition regulation in sorghum and provides a framework for a comprehensive understanding of sorghum photoperiod response.

Development and evolution of the Asteraceae capitulum.

Asteraceae represent one of the largest and most diverse families of plants. The evolutionary success of this family has largely been contributed to their unique inflorescences, capitula that mimic solitary flowers but are typically aggregates of multiple florets. Here, we summarize the recent molecular and genetic level studies that have promoted our understanding of the development and evolution of capitula. We focus on new results on patterning of the enlarged meristem resulting in the iconic phyllotactic arrangement of florets in Fibonacci numbers of spirals. We also summarize the current understanding of the genetic networks regulating the characteristic reproductive traits in the family such as floral dimorphism and differentiation of highly specialized floral organs. So far, developmental studies in Asteraceae are still limited to a very narrow selection of model species. Along with the recent advancements in genomics and phylogenomics, Asteraceae and its relatives provide an outstanding model clade for extended evo-devo studies to exploit the morphological diversity and the underlying molecular networks and to translate this knowledge to the breeding of the key crops in the family.

Exogenous hydrogen sulphide promotes plant flowering through the Arabidopsis splicing factor AtU2AF65a.

Alternative splicing (AS) is an important regulatory mode at the post-transcriptional level, through which many flowering genes regulate floral transition by producing multiple transcripts, and splicing factors have essential roles in this process. Hydrogen sulphide (H2S) is a newly found gasotransmitter that has critical physiological roles in plants, and one of its potential modes of action is via persulfidation of target proteins at specific cysteine sites. Previously, it has been shown that both the splicing factor AtU2AF65a and H2S are involved in the regulation of plant flowering. This study found that, in Arabidopsis, the promoting effect of H2S on flowering was abolished in atu2af65a-4 mutants. Transcriptome analyses showed that when AtU2AF65a contained mutations, the regulatory function of H2S during the AS of many flowering genes (including SPA1, LUH, LUG and MAF3) was inhibited. The persulfidation assay showed that AtU2AF65a can be persulfidated by H2S, and the RNA immunoprecipitation data indicated that H2S could alter the binding affinity of AtU2AF65a to the precursor messenger RNA of the above-mentioned flowering genes. Overall, our results suggest that H2S may regulate the AS of flowering-related genes through persulfidation of splicing factor AtU2AF65a and thus lead to early flowering in plants.

Olfaction in the Anthropocene: NO3 negatively affects floral scent and nocturnal pollination.

There is growing concern about sensory pollutants affecting ecological communities. Anthropogenically enhanced oxidants [ozone (O3) and nitrate radicals (NO3)] rapidly degrade floral scents, potentially reducing pollinator attraction to flowers. However, the physiological and behavioral impacts on pollinators and plant fitness are unknown. Using a nocturnal flower-moth system, we found that atmospherically relevant concentrations of NO3 eliminate flower visitation by moths, and the reaction of NO3 with a subset of monoterpenes is what reduces the scent's attractiveness. Global atmospheric models of floral scent oxidation reveal that pollinators in certain urban areas may have a reduced ability to perceive and navigate to flowers. These results illustrate the impact of anthropogenic pollutants on an animal's olfactory ability and indicate that such pollutants may be critical regulators of global pollination.

Pollen, anther, stamen, and androecium mimicry.

Floral colours represent a highly diverse communication signal mainly involved in flower visitors' attraction and guidance, but also flower discrimination, filtering non-pollinators and discouraging floral antagonists. The divergent visual systems and colour preferences of flower visitors, as well as the necessity of cues for flower detection and discrimination, foster the diversity of floral colours and colour patterns. Despite the bewildering diversity of floral colour patterns, a recurrent component is a yellow UV-absorbing floral centre, and it is still not clear why this pattern is so frequent in angiosperms. The pollen, anther, stamen, and androecium mimicry (PASAM) hypothesis suggests that the system composed of the flowers possessing such yellow UV-absorbing floral reproductive structures, the flowers displaying central yellow UV-absorbing structures as floral guides, and the pollen-collecting, as well as pollen-eating, flower visitors responding to such signals constitute the world's most speciose mimicry system. In this review, we call the attention of researchers to some hypothetical PASAM systems around the globe, presenting some fascinating examples that illustrate their huge diversity. We will also present new and published data on pollen-eating and pollen-collecting pollinators' responses to PASAM structures supporting the PASAM hypothesis and will discuss how widespread these systems are around the globe. Ultimately, our goal is to promote the idea that PASAM is a plausible first approach to understanding floral colour patterns in angiosperms.

A novel type of neighbour perception elicits reproductive plasticity in an annual plant with a mixed mating system.

Plants display various forms of phenotypic plasticity in anticipation of changing conditions, many of which are influenced by information obtained from neighbouring plants. Here, we tested the hypothesis that cleistogamic Lamium amplexicaule plants can adaptively modify production of chasmogamous (CH) and cleistogamous (CL) flowers based on the perception of conspecific neighbours. The production and proportion of CH and CL flowers was examined in individual L. amplexicaule grown at varying densities or treated with root leachates from plants grown at different densities. When growing at high density or treated with root leachates from high-density pots, L. amplexicaule increased production of more expensive, potentially outcrossing CH flowers. In contrast, single plants or plants treated with root leachates from empty pots or single-source plants predominantly developed cheaper, self-pollinated CL flowers. The results demonstrate a novel root-based neighbour-perception modality that enables plants to adaptively adjust production of CH and CL flowers in response to the presence of potential reproductive partners. Further research is needed to explore the broader ecological implications of this novel interplant cueing on reproductive bet-hedging and plasticity in natural settings, as well as to identify the involved cues and their mode of operation.

Epigenetic variation in early and late flowering plants of the rubber-producing Russian dandelion Taraxacum koksaghyz provides insights into the regulation of flowering time.

The Russian dandelion (Taraxacum koksaghyz) grows in temperate zones and produces large amounts of poly(cis-1,4-isoprene) in its roots, making it an attractive alternative source of natural rubber. Most T. koksaghyz plants require vernalization to trigger flower development, whereas early flowering varieties that have lost their vernalization dependence are more suitable for breeding and domestication. To provide insight into the regulation of flowering time in T. koksaghyz, we induced epigenetic variation by in vitro cultivation and applied epigenomic and transcriptomic analysis to the resulting early flowering plants and late flowering controls, allowing us to identify differences in methylation patterns and gene expression that correlated with flowering. This led to the identification of candidate genes homologous to vernalization and photoperiodism response genes in other plants, as well as epigenetic modifications that may contribute to the control of flower development. Some of the candidate genes were homologous to known floral regulators, including those that directly or indirectly regulate the major flowering control gene FT. Our atlas of genes can be used as a starting point to investigate mechanisms that control flowering time in T. koksaghyz in greater detail and to develop new breeding varieties that are more suited to domestication.

Plasticity and not adaptation is the primary source of temperature-mediated variation in flowering phenology in North America.

Phenology varies widely over space and time because of its sensitivity to climate. However, whether phenological variation is primarily generated by rapid organismal responses (plasticity) or local adaptation remains unresolved. Here we used 1,038,027 herbarium specimens representing 1,605 species from the continental United States to measure flowering-time sensitivity to temperature over time (Stime) and space (Sspace). By comparing these estimates, we inferred how adaptation and plasticity historically influenced phenology along temperature gradients and how their contributions vary among species with different phenology and native climates and among ecoregions differing in species composition. Parameters Sspace and Stime were positively correlated (r = 0.87), of similar magnitude and more frequently consistent with plasticity than adaptation. Apparent plasticity and adaptation generated earlier flowering in spring, limited responsiveness in late summer and delayed flowering in autumn in response to temperature increases. Nonetheless, ecoregions differed in the relative contributions of adaptation and plasticity, from consistently greater importance of plasticity (for example, southeastern United States plains) to their nearly equal importance throughout the season (for example, Western Sierra Madre Piedmont). Our results support the hypothesis that plasticity is the primary driver of flowering-time variation along temperature gradients, with local adaptation having a widespread but comparatively limited role.

Phenotypic and transcriptome profiling of spikes reveals the regulation of light regimens on spike growth and fertile floret number in wheat.

The spike growth phase is critical for the establishment of fertile floret (grain) numbers in wheat (Triticum aestivum L.). Then, how to shorten the spike growth phase and increase grain number synergistically? Here, we showed high-resolution analyses of floret primordia (FP) number, morphology and spike transcriptomes during the spike growth phase under three light regimens. The development of all FP in a spike could be divided into four distinct stages: differentiation (Stage I), differentiation and morphology development concurrently (Stage II), morphology development (Stage III), and polarization (Stage IV). Compared to the short photoperiod, the long photoperiod shortened spike growth and stimulated early flowering by shortening Stage III; however, this reduced assimilate accumulation, resulting in fertile floret loss. Interestingly, long photoperiod supplemented with red light shortened the time required to complete Stages I-II, then raised assimilates supply in the spike and promoted anther development before polarization initiation, thereby increasing fertile FP number during Stage III, and finally maintained fertile FP development during Stage IV until they became fertile florets via a predicted dynamic gene network. Our findings proposed a light regimen, critical stages and candidate regulators that achieved a shorter spike growth phase and a higher fertile floret number in wheat.

Floral-promoting GmFT homologs trigger photoperiodic after-effects: An important mechanism for early-maturing soybean varieties to regulate reproductive development and adapt to high latitudes.

Soybean (Glycine max) is a typical short-day plant, but has been widely cultivated in high-latitude long-day (LD) regions because of the development of early-maturing genotypes which are photoperiod-insensitive. However, some early-maturing varieties exhibit significant responses to maturity under different daylengths but not for flowering, depicting an evident photoperiodic after-effect, a poorly understood mechanism. In this study, we investigated the postflowering responses of 11 early-maturing soybean varieties to various preflowering photoperiodic treatments. We confirmed that preflowering SD conditions greatly promoted maturity and other postflowering developmental stages. Soybean homologs of FLOWERING LOCUS T (FT), including GmFT2a, GmFT3a, GmFT3b and GmFT5a, were highly accumulated in leaves under preflowering SD treatment. More importantly, they maintained a high expression level after flowering even under LD conditions. E1 RNAi and GmFT2a overexpression lines showed extremely early maturity regardless of preflowering SD and LD treatments due to constitutively high levels of floral-promoting GmFT homolog expression throughout their life cycle. Collectively, our data indicate that high and stable expression of floral-promoting GmFT homologs play key roles in the maintenance of photoperiodic induction to promote postflowering reproductive development, which confers early-maturing varieties with appropriate vegetative growth and shortened reproductive growth periods for adaptation to high latitudes.

early maturity 7 promotes early flowering by controlling the light input into the circadian clock in barley.

Breeding for variation in photoperiod response is crucial to adapt crop plants to various environments. Plants measure changes in day length by the circadian clock, an endogenous timekeeper that allows plants to anticipate changes in diurnal and seasonal light-dark cycles. Here, we describe the early maturity 7 (eam7) locus in barley (Hordeum vulgare), which interacts with PHOTOPERIOD 1 (Ppd-H1) to cause early flowering under non-inductive short days. We identify LIGHT-REGULATED WD 1 (LWD1) as a putative candidate to underlie the eam7 locus in barley as supported by genetic mapping and CRISPR-Cas9-generated lwd1 mutants. Mutations in eam7 cause a significant phase advance and a misregulation of core clock and clock output genes under diurnal conditions. Early flowering was linked to an upregulation of Ppd-H1 during the night and consequent induction of the florigen FLOWERING LOCUS T1 under short days. We propose that EAM7 controls photoperiodic flowering in barley by controlling the light input into the clock and diurnal expression patterns of the major photoperiod response gene Ppd-H1.

Rhythmic histone acetylation acts in concert with day-night oscillation of the floral volatile metabolic network.

The biosynthesis of specialized metabolites is strictly regulated by environmental inputs such as the day-night cycle, but the underlying mechanisms remain elusive. In Petunia hybrida cv. Mitchell flowers, the biosynthesis and emission of volatile compounds display a diurnal pattern with a peak in the evening to attract nocturnal pollinators. Using petunia flowers as a model system, we found that chromatin level regulation, especially histone acetylation, plays an essential role in mediating the day-night oscillation of the biosynthetic gene network of specialized metabolites. By performing time-course chromatin immunoprecipitation assays for histone modifications, we uncovered that a specific group of genes involved in the regulation, biosynthesis, and emission of floral volatile compounds, which displays the greatest magnitude in day-night oscillating gene expression, is associated with highly dynamic histone acetylation marks H3K9ac and H3K27ac. Specifically, the strongest oscillating genes featured a drastic removal of histone acetylation marks at night, potentially to shut down the biosynthesis of floral volatile compounds during the morning when they are not needed. Inhibiting daytime histone acetylation led to a compromised evening induction of these genes. Overall, our study suggested an active role of chromatin modification in the diurnal oscillation of specialized metabolic network.

FLZ13 interacts with FLC and ABI5 to negatively regulate flowering time in Arabidopsis.

The transition from vegetative to reproductive growth, known as flowering, is a critical developmental process in flowering plants to ensure reproductive success. This process is strictly controlled by various internal and external cues; however, the underlying molecular regulatory mechanisms need to be further characterized. Here, we report a plant-specific protein, FCS-LIKE ZINC FINGER PROTEIN 13 (FLZ13), which functions as a hitherto unknown negative modulator of flowering time in Arabidopsis thaliana. Biochemical analysis showed that FLZ13 directly interacts with FLOWERING LOCUS C (FLC), a major flowering repressor, and that FLZ13 largely depends on FLC to repress the transcription of two core flowering integrators: FLOWERING LOCUS T and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1. In addition, FLZ13 works together with ABSCISIC ACID INSENSITIVE 5 to activate FLC expression to delay flowering. Taken together, our findings suggest that FLZ13 is an important component of the gene regulatory network for flowering time control in plants.

Flower production decreases with warmer and more humid atmospheric conditions in a western Amazonian forest.

Climate models predict that everwet western Amazonian forests will face warmer and wetter atmospheric conditions, and increased cloud cover. It remains unclear how these changes will impact plant reproductive performance, such as flowering, which plays a central role in sustaining food webs and forest regeneration. Warmer and wetter nights may cause reduced flower production, via increased dark respiration rates or alteration in the reliability of flowering cue-based processes. Additionally, more persistent cloud cover should reduce the amounts of solar irradiance, which could limit flower production. We tested whether interannual variation in flower production has changed in response to fluctuations in irradiance, rainfall, temperature, and relative humidity over 18 yrs in an everwet forest in Ecuador. Analyses of 184 plant species showed that flower production declined as nighttime temperature and relative humidity increased, suggesting that warmer nights and greater atmospheric water saturation negatively impacted reproduction. Species varied in their flowering responses to climatic variables but this variation was not explained by life form or phylogeny. Our results shed light on how plant communities will respond to climatic changes in this everwet region, in which the impacts of these changes have been poorly studied compared with more seasonal Neotropical areas.