Eight hydroxyproline-O-galactosyltransferases play essential roles in female reproductive development.

In angiosperms, ovules give rise to seeds upon fertilization. Thus, seed formation is dependent on both successful ovule development and tightly controlled communication between female and male gametophytes. During establishment of these interactions, cell walls play a pivotal role, especially arabinogalactan-proteins (AGPs). AGPs are highly glycosylated proteins decorated by arabinogalactan side chains, representing 90 % of the AGP molecule. AGP glycosylation is initiated by a reaction catalysed by hydroxyproline-O-galactosyltransferases (Hyp-GALTs), specifically eight of them (GALT2-9), which add the first galactose to Hyp residues. Five Hyp-GALTs (GALT2, 5, 7, 8 and 9) were previously described as essential for AGP functions in pollen and ovule development, pollen-pistil interactions, and seed morphology. In the present work, a higher order Hyp-GALT mutant (23456789) was studied, with a high degree of under-glycosylated AGPs, to gain deeper insight into the crucial roles of these eight enzymes in female reproductive tissues. Notably, the 23456789 mutant demonstrated a high quantity of unfertilized ovules, displaying abnormal callose accumulation both at the micropylar region and, sometimes, throughout the entire embryo sac. Additionally, this mutant displayed ovules with abnormal embryo sacs, had a disrupted spatiotemporal distribution of AGPs in female reproductive tissues, and showed abnormal seed and embryo development, concomitant with a reduction in AGP-GlcA levels. This study revealed that at least three more enzymes exhibit Hyp-O-GALT activity in Arabidopsis (GALT3, 4 and 6), and reinforces the crucial importance of AGP carbohydrates in carrying out the biological functions of AGPs during plant reproduction.

The transcription factor LBD10 sustains pollen tube growth and integrity by modulating reactive oxygen species homeostasis via the regulation of flavonol biosynthesis in Arabidopsis.

Reproduction in angiosperms relies on the precise growth of pollen tubes, facilitating the delivery of sperm cells to the ovule for double fertilization. LATERAL ORGAN BOUNDARIES DOMAIN10 (LBD10), a plant-specific transcription factor, plays a pivotal role in Arabidopsis pollen development. Here, we uncovered LBD10's function in sustaining pollen tube growth and integrity. The lbd10 mutant exhibited elevated levels of reactive oxygen species (ROS) and hydrogen peroxide (H2O2) in both pollen grains and tubes, leading to compromised pollen tube growth. The inhibition of ROS synthesis and scavenging of excess ROS with an antioxidant treatment each alleviated these defects in lbd10. The lbd10 mutant displayed reduced flavonol accumulation in both pollen grains and tubes. All the altered phenotypes of lbd10 were complemented by expressing LBD10 under its native promoter. Exogenous application of flavonoids recused the defects in pollen tube growth and integrity in lbd10, along with reducing the excess levels of ROS and H2O2. LBD10 directly binds the promoters of key flavonol biosynthesis genes in chromatin and promotes reporter gene expression in Arabidopsis mesophyll protoplasts. Our findings indicate that LBD10 modulates ROS homeostasis by transcriptionally activating genes crucial for flavonol biosynthesis, thereby maintaining pollen tube growth and integrity.

Tail-dependence of masting synchrony results in continent-wide seed scarcity.

Spatial synchrony may be tail-dependent, meaning it is stronger for peaks rather than troughs, or vice versa. High interannual variation in seed production in perennial plants, called masting, can be synchronized at subcontinental scales, triggering extensive resource pulses or famines. We used data from 99 populations of European beech (Fagus sylvatica) to examine whether masting synchrony differs between mast peaks and years of seed scarcity. Our results revealed that seed scarcity occurs simultaneously across the majority of the species range, extending to populations separated by distances up to 1800 km. Mast peaks were spatially synchronized at distances up to 1000 km and synchrony was geographically concentrated in northeastern Europe. Extensive synchrony in the masting lower tail means that famines caused by beech seed scarcity are amplified by their extensive spatial synchrony, with diverse consequences for food web functioning and climate change biology.

Application of genome editing in plant reproductive biology: recent advances and challenges.

KEY MESSAGE: This comprehensive review underscores the application of genome editing in plant reproductive biology, including recent advances and challenges associated with it. Genome editing (GE) is a powerful technology that has the potential to accelerate crop improvement by enabling efficient, precise, and rapid engineering of plant genomes. Over the last decade, this technology has rapidly evolved from the use of meganucleases (homing endonucleases), zinc-finger nucleases, transcription activator-like effector nucleases to the use of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (CRISPR/Cas), which has emerged as a popular GE tool in recent times and has been extensively used in several organisms, including plants. GE has been successfully employed in several crops to improve plant reproductive traits. Improving crop reproductive traits is essential for crop yields and securing the world's food supplies. In this review, we discuss the application of GE in various aspects of plant reproductive biology, including its potential application in haploid induction, apomixis, parthenocarpy, development of male sterile lines, and the regulation of self-incompatibility. We also discuss current challenges and future prospects of this technology for crop improvement, focusing on plant reproduction.

Evolutionary ecology of masting: mechanisms, models, and climate change.

Many perennial plants show mast seeding, characterized by synchronous and highly variable reproduction across years. We propose a general model of masting, integrating proximate factors (environmental variation, weather cues, and resource budgets) with ultimate drivers (predator satiation and pollination efficiency). This general model shows how the relationships between masting and weather shape the diverse responses of species to climate warming, ranging from no change to lower interannual variation or reproductive failure. The role of environmental prediction as a masting driver is being reassessed; future studies need to estimate prediction accuracy and the benefits acquired. Since reproduction is central to plant adaptation to climate change, understanding how masting adapts to shifting environmental conditions is now a central question.

Plant reproduction research in Latin America: Toward sustainable agriculture in a changing environment.

Food production and food security depend on the ability of crops to cope with anthropogenic climate change and successfully produce seed. To guarantee food production well into the future, contemporary plant scientists in Latin America must carry out research on how plants respond to environmental stressors such as temperature, drought, and salinity. This review shows the opportunities to apply these results locally and abroad and points to the gaps that still exist in terms of reproductive processes with the purpose to better link research with translational work in plant breeding and biotechnology. Suggestions are put forth to address these gaps creatively in the face of chronic low investment in science with a focus on applicability.

Continental-scale evaluation of downy birch pollen production: Estimating the impacts of global change.

The high prevalence of hay fever in Europe has raised concerns about the implications of climate change-induced higher temperatures on pollen production. Our study focuses on downy birch pollen production across Europe by analyzing 456 catkins during 2019-2021 in 37 International Phenological Gardens (IPG) spanning a large geographic gradient. As IPGs rely on genetically identical plants, we were able to reduce the effects of genetic variability. We studied the potential association with masting behavior and three model specifications based on mean and quantile regression to assess the impact of meteorology (e.g., temperature and precipitation) and atmospheric gases (e.g., ozone (O3) and carbon-dioxide (CO2)) on pollen and catkin production, while controlling for tree age approximated by stem circumference. The results revealed a substantial geographic variability in mean pollen production, ranging from 1.9 to 2.5 million pollen grains per catkin. Regression analyses indicated that elevated average temperatures of the previous summer corresponded to increased pollen production, while higher O3 levels led to a reduction. Additionally, catkins number was positively influenced by preceding summer's temperature and precipitation but negatively by O3 levels. The investigation of quantile effects revealed that the impacts of mean temperature and O3 levels from the previous summer varied throughout the conditional response distribution. We found that temperature predominantly affected trees characterized by a high pollen production. We therefore suggest that birches modulate their physiological processes to optimize pollen production under varying temperature regimes. In turn, O3 levels negatively affected trees with pollen production levels exceeding the conditional median. We conclude that future temperature increase might exacerbate pollen production while other factors may modify (decrease in the case of O3 and amplify for precipitation) this effect. Our comprehensive study sheds light on potential impacts of climate change on downy birch pollen production, which is crucial for birch reproduction and human health.

Pollen thermotolerance of a widespread plant, Lotus corniculatus, in response to climate warming: possible local adaptation of populations from different elevations.

One of the most vulnerable phases in the plant life cycle is sexual reproduction, which depends on effective pollen transfer, but also on the thermotolerance of pollen grains. Pollen thermotolerance is temperature-dependent and may be reduced by increasing temperature associated with global warming. A growing body of research has focused on the effect of increased temperature on pollen thermotolerance in crops to understand the possible impact of temperature extremes on yield. Yet, little is known about the effects of temperature on pollen thermotolerance of wild plant species. To fill this gap, we selected Lotus corniculatus s.l. (Fabaceae), a species common to many European habitats and conducted laboratory experiments to test its pollen thermotolerance in response to artificial increase in temperature. To test for possible local adaptation of pollen thermal tolerance, we compared data from six lowland (389-451 m a.s.l.) and six highland (841-1,030 m a.s.l.) populations. We observed pollen germination in vitro at 15 °C, 25 °C, 30 °C, and 40 °C. While lowland plants maintained a stable germination percentage across a broad temperature range (15-30 °C) and exhibited reduced germination only at extremely high temperatures (40 °C), highland plants experienced reduced germination even at 30 °C-temperatures commonly exceeded in lowlands during warm summers. This suggests that lowland populations of L. corniculatus may be locally adapted to higher temperature for pollen germination. On the other hand, pollen tube length decreased with increasing temperature in a similar way in lowland and highland plants. The overall average pollen germination percentage significantly differed between lowland and highland populations, with highland populations displaying higher germination percentage. On the other hand, the average pollen tube length was slightly smaller in highland populations. In conclusion, we found that pollen thermotolerance of L. corniculatus is reduced at high temperature and that the germination of pollen from plant populations growing at higher elevations is more sensitive to increased temperature, which suggests possible local adaptation of pollen thermotolerance.

Screening methods for thermotolerance in pollen.

Plant reproduction is highly susceptible to temperature stress. The development of the male gametophyte in particular represents a critical element in the reproductive cycle with high sensitivity to elevated temperatures. Various methods have been used to test the effect of temperature stress on pollen performance or to determine the degree of susceptibility of given species and genotypes. The information gained informs the development of new crop varieties suited to grow under warmer conditions arising through climate change and facilitates predicting the behavior of natural populations under these conditions. The characterization of pollen performance typically employs the terms pollen viability and pollen vigor, which, however, are not necessarily used consistently across studies. Pollen viability is a nominal parameter and is often assayed relying on cellular features as proxy to infer the capability of pollen grains to germinate and complete double fertilization. Alternatively, pollen germination can be determined through in vitro growth assays, or by monitoring the ability of pollen tubes to complete different progamic steps in vivo (ability to reach an ovule, release sperm cells, lead to seed set). Pollen vigor is an ordinal parameter that describes pollen tube growth rate or the efficiency of pollen tube growth as inferred by its morphology or growth pattern. To ensure consistent and relevant terminology, this review defines these terms and summarizes the methodologies used to assess them.

Genome wide characterization and expression analysis of CrRLK1L gene family in wheat unravels their roles in development and stress-specific responses.

Catharanthus roseus receptor-like kinase 1-like (CrRLK1L) genes encode a subfamily of receptor-like kinases (RLK) that regulate diverse processes during plant growth, development, and stress responses. The first CrRLK1L was identified from the Catharanthus roseus, commonly known as Madagascar periwinkle. Subsequently, CrRLK1L gene families have been characterized in many plants. The genome of T. aestivum encodes 15 CrRLK1L genes with 43 paralogous copies, with three homeologs each, except for -2-D and -7-A, which are absent. Chromosomal localization analysis revealed a markedly uneven distribution of CrRLK1L genes across seven different chromosomes, with chromosome 4 housing the highest number of genes, while chromosome 6 lacked any CrRLK1L genes. Tissue-specific gene expression analysis revealed distinct expression patterns among the gene family members, with certain members exhibiting increased expression in reproductive tissues. Gene expression analysis in response to various abiotic and biotic stress conditions unveiled differential regulation of gene family members. Cold stress induces CrRLK1Ls -4-B and -15-A while downregulating -3-A and -7B. Drought stress upregulates -9D, contrasting with the downregulation of -7D. CrRLK1L-15-B and -15-D were highly induced in response to 1 hr of heat, and combined drought and heat stress, whereas -10-B is downregulated. Similarly, in response to NaCl stress, only CrRLK1L1 homeologs were induced. Fusarium graminearum and Claviceps purpurea inoculation induces homeologs of CrRLK1L-6 and -7. The analysis of cis-acting elements in the promoter regions identified elements crucial for plant growth and developmental processes. This comprehensive genome-wide analysis and expression study provides valuable insights into the essential functions of CrRLK1L members in wheat.

Evidence of combined flower thermal and drought vulnerabilities portends reproductive failure under hotter-drought conditions.

Despite the abundant evidence of impairments to plant performance and survival under hotter-drought conditions, little is known about the vulnerability of reproductive organs to climate extremes. Here, by conducting a comparative analysis between flowers and leaves, we investigated how variations in key morphophysiological traits related to carbon and water economics can explain the differential vulnerabilities to heat and drought among these functionally diverse organs. Due to their lower construction costs, despite having a higher water storage capacity, flowers were more prone to turgor loss (higher turgor loss point; ΨTLP) than leaves, thus evidencing a trade-off between carbon investment and drought tolerance in reproductive organs. Importantly, the higher ΨTLP of flowers also resulted in narrow turgor safety margins (TSM). Moreover, compared to leaves, the cuticle of flowers had an overall higher thermal vulnerability, which also resulted in low leakage safety margins (LSM). As a result, the combination of low TSMs and LSMs may have negative impacts on reproduction success since they strongly influenced the time to turgor loss under simulated hotter-drought conditions. Overall, our results improve the knowledge of unexplored aspects of flower structure and function and highlight likely threats to successful plant reproduction in a warmer and drier world.

Effects of selfing on the evolution of sexual reproduction.

Plants exhibit diverse breeding systems, with populations capable of outcrossing, selfing, and/or asexual reproduction. However, interactions between the three reproductive pathways remain not fully clear. Sexual reproduction introduces segregation and recombination, but incurs several costs. Selfing can affect the relative costs and benefits of sexual vs. asexual reproduction. Building population genetic models, I explore how selfing affects the evolution of a sexual reproduction rate modifier via (a) indirect selection due to segregation, (b) indirect selection from changes in recombination rates, and (c) selection from the cost of meiosis and mate limitation. The dominant selective force mediating the evolution of sex is found to vary with the rate of sexual reproduction and selfing, but selective force (a) and (c) are generally stronger than selective force (b). A modifier enhancing sexual reproduction tends to be favored by indirect selection generated by partially recessive, small-effect deleterious mutations, while hindered by highly recessive lethal mutations. Overall, evolution toward higher sexual reproduction is hindered at low sexual reproduction rates and intermediate selfing rates, but favored under high selfing rates. The results suggest that asexual reproduction may precede the evolution of selfing and offer insights into the evolution of mechanisms reducing geitonogamy in partially clonal populations.

Effective population size mediates the impact of pollination services on pollen limitation.

Inadequate pollen receipt limits flowering plant reproduction worldwide. Ecological causes of pollen limitation (PL), like pollinator scarcity and low plant abundance, have been a primary research focus. The genetic diversity of plant populations could impact both quantity and quality components of PL in concert with ecological factors, yet empirical examples are lacking. We evaluated joint effects of ecological factors (flower abundance, pollinator visitation) and genetic effective population size (NE) on PL across 13 populations of the common herb Argentina anserina. We used a histological approach with 5504 styles from 1137 flowers to separate quantity and quality components of PL, and link these to reproductive output. NE and pollinator visitation interacted to shape PL, but NE had stronger direct effects. Effectively smaller populations experienced stronger quantity PL, and controlled crosses in a pollinator-free environment revealed that stigmatic pollen quantity was an intrinsic population-level attribute that increased with NE. Pollinator visitation enhanced pollen quality, but only in effectively larger populations. Quantity and quality PL negatively impacted fruit and seed set, respectively. Results highlight that PL is dictated by plant population genetic diversity in addition to commonly evaluated ecological factors. Efforts to support pollinators will more strongly enhance plant reproduction in genetically diverse populations.

Evolution of flowering time due to variation in the onset of pollen dispersal among individuals.

The evolution of flowering time is often attributed to variations in pollinator rates over time. This study proposes that flowering time can evolve through siring success variation among individuals caused by differential pollen dispersal timing (a result of flowering time variation). By building quantitative genetic models, I show that flowering time evolves to be earlier when the pollen removal rate is low and pollen deposition rate is high, and the fertilization ability of removed pollen declines slowly. Using evolutionary game theory, I show that the evolutionarily stable variance of flowering time is large when the pollen removal rate is either low or high, the pollen deposition rate is moderate, and the fertilization ability of removed pollen declines rapidly. Investigation of the coevolution of flower longevity and flowering time shows that under constant pollination rates, late flowering will be correlated with long-lived flowers due to nonrandom mating, which suggests that the observed correlation between late flowering and short-lived flowers is caused by other factors, such as declining pollination rates during late-stage flowering. I discuss how altered pollination rates under climate change will influence flowering time evolution and the importance of distinguishing between pollen removal and deposition rates.

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.

Pollinator-mediated effects of landscape-scale land use on grassland plant community composition and ecosystem functioning - seven hypotheses.

Environmental change is disrupting mutualisms between organisms worldwide. Reported declines in insect populations and changes in pollinator community compositions in response to land use and other environmental drivers have put the spotlight on the need to conserve pollinators. While this is often motivated by their role in supporting crop yields, the role of pollinators for reproduction and resulting taxonomic and functional assembly in wild plant communities has received less attention. Recent findings suggest that observed and experimental gradients in pollinator availability can affect plant community composition, but we know little about when such shifts are to be expected, or the impact they have on ecosystem functioning. Correlations between plant traits related to pollination and plant traits related to other important ecosystem functions, such as productivity, nitrogen uptake or palatability to herbivores, lead us to expect non-random shifts in ecosystem functioning in response to changes in pollinator communities. At the same time, ecological and evolutionary processes may counteract these effects of pollinator declines, limiting changes in plant community composition, and in ecosystem functioning. Despite calls to investigate community- and ecosystem-level impacts of reduced pollination, the study of pollinator effects on plants has largely been confined to impacts on plant individuals or single-species populations. With this review we aim to break new ground by bringing together aspects of landscape ecology, ecological and evolutionary plant-insect interactions, and biodiversity-ecosystem functioning research, to generate new ideas and hypotheses about the ecosystem-level consequences of pollinator declines in response to land-use change, using grasslands as a focal system. Based on an integrated set of seven hypotheses, we call for more research investigating the putative pollinator-mediated links between landscape-scale land use and ecosystem functioning. In particular, future research should use combinations of experimental and observational approaches to assess the effects of changes in pollinator communities over multiple years and across species on plant communities and on trait distributions both within and among species.

The impact of heat stress in plant reproduction.

The increment in global temperature reduces crop productivity, which in turn threatens food security. Currently, most of our food supply is produced by plants and the human population is estimated to reach 9 billion by 2050. Gaining insights into how plants navigate heat stress in their reproductive phase is essential for effectively overseeing the future of agricultural productivity. The reproductive success of numerous plant species can be jeopardized by just one exceptionally hot day. While the effects of heat stress on seedlings germination and root development have been extensively investigated, studies on reproduction are limited. The intricate processes of gamete development and fertilization unfold within a brief timeframe, largely concealed within the flower. Nonetheless, heat stress is known to have important effects on reproduction. Considering that heat stress typically affects both male and female reproductive structures concurrently, it remains crucial to identify cultivars with thermotolerance. In such cultivars, ovules and pollen can successfully undergo development despite the challenges posed by heat stress, enabling the completion of the fertilization process and resulting in a robust seed yield. Hereby, we review the current understanding of the molecular mechanisms underlying plant resistance to abiotic heat stress, focusing on the reproductive process in the model systems of Arabidopsis and Oryza sativa.

SPA, a Stigma-style-transmitting tract Physical microenvironment Assay for investigating mechano-signaling in pollen tubes.

Accurate sensing and responding to physical microenvironment are crucial for cell function and survival, but the underlying molecular mechanisms remain elusive. Pollen tube (PT) provides a perfect single-cell model for studying mechanobiology since it's naturally subjected to complex mechanical instructions from the pistil during invasive growth. Recent reports have revealed discrepant PT behaviors between in vivo and flat, two-dimensional in vitro cultures. Here, we established the Stigma-style-transmitting tract (TT) Physical microenvironment Assay (SPA) to recapitulate pressure changes in the pistil. This biomimetic assay has enabled us to swiftly identify highly redundant genes, GEF8/9/11/12/13, as new regulators for maintaining PTs integrity during style-to-TT emergence. In contrast to normal growth on solid medium, SPA successfully phenocopied gef8/9/11/12/13 PT in vivo growth-arrest deficiency. Our results suggest the existence of distinct signaling pathways regulating in vivo and in vitro PT integrity maintenance, underscoring the necessity of faithfully mimicking the physical microenvironment for studying plant cell biology.

Plant Hormone and Fatty Acid Screening of Nicotiana tabacum and Lilium longiflorum Stigma Exudates.

Pollen germination in vivo on wet stigmas is assisted by the receptive fluid-stigma exudate. Its exact composition is still unknown because only some components have been studied. For the first time, hormonal screening was carried out, and the fatty acid (FA) composition of lipid-rich (Nicotiana tabacum) and sugar-rich (Lilium longiflorum) exudates was studied. Screening of exudate for the presence of plant hormones using HPLC-MS revealed abscisic acid (ABA) in tobacco stigma exudate at the two stages of development, at pre-maturity and in mature stigmas awaiting pollination, increasing at the fertile stage. To assess physiological significance of ABA on stigma, we tested the effect of this hormone in vitro. ABA concentration found in the exudate strongly stimulated the germination of tobacco pollen, a lower concentration had a weaker effect, increasing the concentration did not increase the effect. GC-MS analysis showed that both types of exudate are characterized by a predominance of saturated FAs. The lipids of tobacco stigma exudate contain significantly more myristic, oleic, and linoleic acids, resulting in a higher unsaturation index relative to lily stigma exudate lipids. The latter, in turn, contain more 14-hexadecenoic and arachidic acids. Both exudates were found to contain significant amounts of squalene. The possible involvement of saturated FAs, ABA, and squalene in various exudate functions, as well as their potential relationship on the stigma, is discussed.