Pollen-stigma Interactions Research Articles

Non-transformation methods for studying signaling pathways and genes involved in Brassica rapa pollen-stigma interactions.

Self-incompatibility (SI) is a mechanism in plants that prevents self-fertilization and promotes out-crossing. SI is also widely utilized in the breeding of Brassicaceae crops. Understanding the regulatory mechanisms of SI is essential but has been greatly restrained in most Brassicaceae crops due to inefficient transformation. Here, we developed methods for studying signaling pathways and genes of pollen-stigma interactions in Brassicaceae crops lacking an efficient genetic transformation system. We pretreated excised stigmas of Brassica rapa (Brassica rapa L. ssp. Pekinensis) in vitro with chemicals to modify signaling pathways or with phosphorothioate antisense oligodeoxyribonucleotides (AS-ODNs) to modify the expression of the corresponding genes involved in pollen-stigma interactions. Using this method, we firstly determined the involvement of reactive oxygen species (ROS) in SI with the understanding that the NADPH oxidase inhibitor diphenyleneiodonium chloride (DPI), which inhibits ROS production, eliminated SI of B. rapa. We further identified the key gene for ROS production in SI and used AS-ODNs targeting BrRBOHF (Brassica rapa RESPIRATORY-BURST OXIDASE HOMOLOGF), which encodes one of the NADPH oxidases, to effectively suppress its expression, reduce stigmatic ROS, and promote the growth of self-pollen in B. rapa stigmas. Moreover, pistils treated in planta with the ROS scavenger sodium salicylate (Na-SA) disrupted SI and resulted in enlarged ovules with inbred embryos 12 days after pollination. This method will enable the functional study of signaling pathways and genes regulating SI and other pollen-stigma interactions in different Brassicaceae plants.

Morphological insights into the pollen–stigma interaction in Mimosa caesalpiniifolia Benth. (Leguminosae): exploring the lock-and-key mechanism

Morphological insights into the pollen–stigma interaction in Mimosa caesalpiniifolia Benth. (Leguminosae): exploring the lock-and-key mechanism

Knockout of stigmatic ascorbate peroxidase 1 (APX1) delays pollen rehydration and germination by mediating ROS homeostasis in Brassica napus L.

The successful interaction between pollen and stigma is a critical process for plant sexual reproduction, involving a series of intricate molecular and physiological events. After self-compatible pollination, a significant reduction in reactive oxygen species (ROS) production has been observed in stigmas, which is essential for pollen grain rehydration and subsequent pollen tube growth. Several scavenging enzymes tightly regulate ROS homeostasis. However, the potential role of these ROS-scavenging enzymes in the pollen-stigma interaction in Brassica napus remains unclear. Here, we showed that the activity of ascorbate peroxidase (APX), an enzyme that plays a crucial role in the detoxification of hydrogen peroxide (H2O2), was modulated depending on the compatibility of pollination in B. napus. We then identified stigma-expressed APX1s and generated pentuple mutants of APX1s using CRISPR/Cas9 technology. After compatible pollination, the BnaAPX1 pentuple mutants accumulated higher levels of H2O2 in the stigma, while the overexpression of BnaA09.APX1 resulted in lower levels of H2O2. Furthermore, the knockout of BnaAPX1 delayed the compatible response-mediated pollen rehydration and germination, which was consistent with the effects of a specific APX inhibitor, ρ-Aminophenol, on compatible pollination. In contrast, the overexpression of BnaA09.APX1 accelerated pollen rehydration and germination after both compatible and incompatible pollinations. However, delaying and promoting pollen rehydration and germination did not affect the seed set after compatible and incompatible pollination in APX1 pentuple mutants and overexpression lines, respectively. Our results demonstrate the fundamental role of BnaAPX1 in pollen rehydration and germination by regulating ROS homeostasis during the pollen-stigma interaction in B. napus.

Reactive oxygen species are signaling molecules that modulate plant reproduction.

Reactive oxygen species (ROS) can serve as signaling molecules that are essential for plant growth and development but abiotic stress can lead to ROS increases to supraoptimal levels resulting in cellular damage. To ensure efficient ROS signaling, cells have machinery to locally synthesize ROS to initiate cellular responses and to scavenge ROS to prevent it from reaching damaging levels. This review summarizes experimental evidence revealing the role of ROS during multiple stages of plant reproduction. Localized ROS synthesis controls the formation of pollen grains, pollen-stigma interactions, pollen tube growth, ovule development,and fertilization. Plants utilize ROS-producing enzymes such as respiratory burst oxidase homologsand organelle metabolic pathways to generate ROS, while the presence of scavenging mechanisms, including synthesis of antioxidant proteins and small molecules, serves to prevent its escalation to harmful levels. In this review, we summarized the function of ROS and its synthesisand scavenging mechanisms in all reproductive stages from gametophyte development until completion of fertilization. Additionally, we further address the impact of elevated temperatures induced ROS on impairing these reproductive processes and of flavonol antioxidants in maintaining ROS homeostasis to minimize temperature stress to combat the impact of global climate change on agriculture.

The Arabidopsis SNARE complex genes regulate the early stages of pollen-stigma interactions.

The VAMP721, VAMP722, SYP121, SYP122 and SNAP33 SNAREs are required in the Arabidopsis stigma for pollen hydration, further supporting a role for vesicle trafficking in the stigma's pollen responses. In the Brassicaceae, the process of accepting compatible pollen is a key step in successful reproduction and highly regulated following interactions between the pollen and the stigma. Central to this is the initiation of secretion in the stigma, which is proposed to provide resources to the pollen for hydration and germination and pollen tube growth. Previously, the eight exocyst subunit genes were shown to be required in the Arabidopsis stigma to support these pollen responses. One of the roles of the exocyst is to tether secretory vesicles at the plasma membrane for membrane fusion by the SNARE complex to enable vesicle cargo release. Here, we investigate the role of Arabidopsis SNARE genes in the stigma for pollen responses. Using a combination of different knockout and knockdown SNARE mutant lines, we show that VAMP721, VAMP722, SYP121, SYP122 and SNAP33 are involved in this process. Significant disruptions in pollen hydration were observed following pollination of wildtype pollen on the mutant SNARE stigmas. Overall, these results place the Arabidopsis SNARE complex as a contributor in the stigma for pollen responses and reaffirm the significance of secretion in the stigma to support the pollen-stigma interactions.

SD-RLK28 positively regulates pollen hydration on stigmas as a PCP-Bβ receptor in Arabidopsis thaliana.

Pollen hydration on dry stigmas is strictly regulated by pollen-stigma interactions in Brassicaceae. Although several related molecular events have been described, the molecular mechanism underlying pollen hydration remains elusive. Multiple B-class pollen coat proteins (PCP-Bs) are involved in pollen hydration. Here, by analyzing the interactions of two PCP-Bs with three Arabidopsis thaliana stigmas strongly expressing S-domain receptor kinase (SD-RLK), we determined that SD-RLK28 directly interacts with PCP-Bβ. We investigated pollen hydration, pollen germination, pollen tube growth, and stigma receptivity in the sd-rlk28 and pcp-bβ mutants. PCP-Bβ acts in the pollen to regulate pollen hydration on stigmas. Loss of SD-RLK28 had no effect on pollen viability, and sd-rlk28 plants had normal life cycles without obvious defects. However, pollen hydration on sd-rlk28 stigmas was impaired and pollen tube growth in sd-rlk28 pistils was delayed. The defect in pollen hydration on sd-rlk28 stigmas was independent of changes in reactive oxygen species levels in stigmas. These results indicate that SD-RLK28 functions in the stigma as a PCP-Bβ receptor to positively regulate pollen hydration on dry stigmas.

Putting the brakes on pollen wall development: A conserved negative feedback loop regulates pollen exine formation in flowering plants

Pollen grains are covered by a complex multi-layered structure, the pollen wall. Exine, the outermost layer of this wall, is made from the biopolymer sporopollenin that exhibits remarkable physical and chemical stability. In different plants, exine, which shields pollen from harsh environmental stresses, has very diverse morphologies and forms different patterns (Wang and Dobritsa, 2018). It also contributes to pollination, pollen-stigma interaction, pollen hydration, and eventual release of the pollen tube. Formation of exine begins after male meiosis when four microspores are arranged in a tetrad configuration and enclosed by the callose wall (Ariizumi and Toriyama, 2011). At this stage, microspores secrete primexine, the precursor of exine that accumulates between the microspore plasma membrane and the callose wall, providing a scaffold for exine patterning (Wang et al., 2021). As the callose wall gradually gets degraded, sporopollenin is secreted by the nearby tapetal cells and deposited onto the primexine template to form mature exine. This process requires coordination of activities between microspores and the surrounding tapetum.

Self-incompatibility: a targeted, unexplored pre-fertilization barrier in flower crops of Asteraceae.

Asteraceae (synonym as Compositae) is one of the largest angiosperm families among flowering plants comprising one-tenth of all agri-horticultural species grown across various habitats except in Antarctica. These are commercially utilized as cut and loose flowers as well as pot and bedding plants in landscape gardens due to their unique floral traits. Consequently,ineffectiveseedsettingand presence of an intraspecific reproductive barrier known as self-incompatibility (SI) severelyreducestheeffectivenessof hybridization and self-fertilization bytraditionalcrossing. There have been very few detailed studies of pollen-stigma interactions in this family. Moreover, about 63% of Aster species can barely self-fertilize due to self-incompatibility (SI). The chrysanthemum (Chrysanthemum × morifolium) is one of the most economically important ornamental plants in the Asteraceae family which hugely shows incompatibility. Reasons for the low fertility and reproductive capacity of species are still indefinite or not clear. Hence, the temporal pattern of inheritance of self-incompatibility and its effect on reproductive biology needs to be investigated further to improve the breeding efficiency. This review highlights the self-incompatible (SI) system operating in important Astraceous (ornamental) crops which are adversely affected by this mechanism along with different physiological and molecular techniques involved in breaking down self-incompatibility.

Biosynthesis and transport of pollen coat precursors in angiosperms.

The pollen coat is a hydrophobic mixture on the pollen grain surface, which plays an important role in protecting male gametes from various environmental stresses and microorganism attacks, and in pollen-stigma interactions during pollination in angiosperms. An abnormal pollen coat can result in humidity-sensitive genic male sterility (HGMS), which can be used in two-line hybrid crop breeding. Despite the crucial functions of the pollen coat and the application prospect of its mutants, few studies have focused on pollen coat formation. In this Review, the morphology, composition and function of different types of pollen coat are assessed. On the basis of the ultrastructure and development process of the anther wall and exine found in rice and Arabidopsis, the genes and proteins involved in the biosynthesis of pollen coat precursors and the possible transport and regulation process are sorted. Additionally, current challenges and future perspectives, including potential strategies utilizing HGMS genes in heterosis and plant molecular breeding, are highlighted.

Ovule Transcriptome Analysis Discloses Deregulation of Genes and Pathways in Sexual and Apomictic Limonium Species (Plumbaginaceae).

The genus Limonium Mill. (sea lavenders) includes species with sexual and apomixis reproductive strategies, although the genes involved in these processes are unknown. To explore the mechanisms beyond these reproduction modes, transcriptome profiling of sexual, male sterile, and facultative apomictic species was carried out using ovules from different developmental stages. In total, 15,166 unigenes were found to be differentially expressed with apomictic vs. sexual reproduction, of which 4275 were uniquely annotated using an Arabidopsis thaliana database, with different regulations according to each stage and/or species compared. Gene ontology (GO) enrichment analysis indicated that genes related to tubulin, actin, the ubiquitin degradation process, reactive oxygen species scavenging, hormone signaling such as the ethylene signaling pathway and gibberellic acid-dependent signal, and transcription factors were found among differentially expressed genes (DEGs) between apomictic and sexual plants. We found that 24% of uniquely annotated DEGs were likely to be implicated in flower development, male sterility, pollen formation, pollen-stigma interactions, and pollen tube formation. The present study identifies candidate genes that are highly associated with distinct reproductive modes and sheds light on the molecular mechanisms of apomixis expression in Limonium sp.

Comprehensive Analysis of BrHMPs Reveals Potential Roles in Abiotic Stress Tolerance and Pollen–Stigma Interaction in Brassica rapa

Heavy metal-associated proteins (HMPs) participate in heavy metal detoxification. Although HMPs have been identified in several plants, no studies to date have identified the HMPs in Brassica rapa (B. rapa). Here, we identified 85 potential HMPs in B. rapa by bioinformatic methods. The promoters of the identified genes contain many elements associated with stress responses, including response to abscisic acid, low-temperature, and methyl jasmonate. The expression levels of BrHMP14, BrHMP16, BrHMP32, BrHMP41, and BrHMP42 were upregulated under Cu2+, Cd2+, Zn2+, and Pb2+ stresses. BrHMP06, BrHMP30, and BrHMP41 were also significantly upregulated after drought treatment. The transcripts of BrHMP06 and BrHMP11 increased mostly under cold stress. After applying salt stress, the expression of BrHMP02, BrHMP16, and BrHMP78 was induced. We observed increased BrHMP36 expression during the self-incompatibility (SI) response and decreased expression in the compatible pollination (CP) response during pollen-stigma interactions. These changes in expression suggest functions for these genes in HMPs include participating in heavy metal transport, detoxification, and response to abiotic stresses, with the potential for functions in sexual reproduction. We found potential co-functional partners of these key players by protein-protein interaction (PPI) analysis and found that some of the predicted protein partners are known to be involved in corresponding stress responses. Finally, phosphorylation investigation revealed many phosphorylation sites in BrHMPs, suggesting post-translational modification may occur during the BrHMP-mediated stress response. This comprehensive analysis provides important clues for the study of the molecular mechanisms of BrHMP genes in B. rapa, especially for abiotic stress and pollen-stigma interactions.

The triticale mature pollen and stigma proteomes – assembling the proteins for a productive encounter

Triticeae crops are major contributors to global food production and ensuring their capacity to reproduce and generate seeds is critical. However, despite their importance our knowledge of the proteins underlying Triticeae reproduction is severely lacking and this is not only true of pollen and stigma development, but also of their pivotal interaction. When the pollen grain and stigma are brought together they have each accumulated the proteins required for their intended meeting and accordingly studying their mature proteomes is bound to reveal proteins involved in their diverse and complex interactions. Using triticale as a Triticeae representative, gel-free shotgun proteomics was used to identify 11,533 and 2977 mature stigma and pollen proteins respectively. These datasets, by far the largest to date, provide unprecedented insights into the proteins participating in Triticeae pollen and stigma development and interactions. The study of the Triticeae stigma has been particularly neglected. To begin filling this knowledge gap, a developmental iTRAQ analysis was performed revealing 647 proteins displaying differential abundance as the stigma matures in preparation for pollination. An in-depth comparison to an equivalent Brassicaceae analysis divulged both conservation and diversification in the makeup and function of proteins involved in the pollen and stigma encounter. SignificanceSuccessful pollination brings together the mature pollen and stigma thus initiating an intricate series of molecular processes vital to crop reproduction. In the Triticeae crops (e.g. wheat, barley, rye, triticale) there persists a vast deficit in our knowledge of the proteins involved which needs to be addressed if we are to face the many upcoming challenges to crop production such as those associated with climate change. At maturity, both the pollen and stigma have acquired the protein complement necessary for their forthcoming encounter and investigating their proteomes will inevitably provide unprecedented insights into the proteins enabling their interactions. By combining the analysis of the most comprehensive Triticeae pollen and stigma global proteome datasets to date with developmental iTRAQ investigations, proteins implicated in the different phases of pollen-stigma interaction enabling pollen adhesion, recognition, hydration, germination and tube growth, as well as those underlying stigma development were revealed. Extensive comparisons between equivalent Triticeae and Brassiceae datasets highlighted both the conservation of biological processes in line with the shared goal of activating the pollen grain and promoting pollen tube invasion of the pistil to effect fertilization, as well as the significant distinctions in their proteomes consistent with the considerable differences in their biochemistry, physiology and morphology.

Pollen Coat Proteomes of Arabidopsis thaliana, Arabidopsis lyrata, and Brassica oleracea Reveal Remarkable Diversity of Small Cysteine-Rich Proteins at the Pollen-Stigma Interface

The pollen coat is the outermost domain of the pollen grain and is largely derived from the anther tapetum, which is a secretory tissue that degenerates late in pollen development. By being localised at the interface of the pollen-stigma interaction, the pollen coat plays a central role in mediating early pollination events, including molecular recognition. Amongst species of the Brassicaceae, a growing body of data has revealed that the pollen coat carries a range of proteins, with a number of small cysteine-rich proteins (CRPs) being identified as important regulators of the pollen-stigma interaction. By utilising a state-of-the-art liquid chromatography/tandem mass spectrometry (LC-MS/MS) approach, rich pollen coat proteomic profiles were obtained for Arabidopsis thaliana, Arabidopsis lyrata, and Brassica oleracea, which greatly extended previous datasets. All three proteomes revealed a strikingly large number of small CRPs that were not previously reported as pollen coat components. The profiling also uncovered a wide range of other protein families, many of which were enriched in the pollen coat proteomes and had functions associated with signal transduction, cell walls, lipid metabolism and defence. These proteomes provide an excellent source of molecular targets for future investigations into the pollen-stigma interaction and its potential evolutionary links to plant-pathogen interactions.

Cytoskeleton Remodeling in Arabidopsis Stigmatic Cells Following Pollination.

In plants, the first interaction that occurs between the male gametophytes (pollen grains) and the stigmatic epidermis of the female organ is crucial for successful reproduction. The stigma consists of a dome of flask-shaped cells specialized in pollen capture. In these stigmatic cells, the cytoskeleton network (cortical microtubules and actin microfilaments) actively responds to pollen contact and undergoes dynamic remodeling required for successful pollen acceptance to occur. Here, we have designed several microscopy mountings to monitor stigmatic cytoskeleton dynamics. These designs are based on the constraints linked to the tightly regulated pollen-stigma interaction and depend upon the experimental goal, either a static view or live-cell imaging.

Eco-Geography and Phenology Are the Major Drivers of Reproductive Isolation in the Royal Irises, a Species Complex in the Course of Speciation

The continuous nature of speciation implies that different species are found at different stages of divergence, from no- to complete reproductive isolation. This process and its underlying mechanisms are best viewed in incipient species. Moreover, the species complex can offer unique insight into how reproductive isolation (RI) has evolved. The royal irises (Iris section Oncocyclus) are a young group of species in the course of speciation, providing an ideal system for speciation study. We quantified pre- and post-zygotic reproductive barriers between the eight Israeli species of this complex and estimated the total RI among them. We tested for both pre-pollination and post-pollination reproductive barriers. Pre-pollination barriers, i.e., eco-geographic divergence and phenological differentiation were the major contributors to RI among the Iris species. On the other hand, post-pollination barriers, namely pollen-stigma interactions, fruit set, and seed viability had negligible contributions to total RI. The strength of RI was not uniform across the species complex, suggesting that species may have diverged at different rates. Overall, this study in a young, recently diverged group of species provides insight into the first steps of speciation, suggesting a crucial role of the pre-zygotic barriers.

Interactions of airborne graphene oxides with the sexual reproduction of a model plant: When production impurities matter

The increasing use of graphene-related materials (GRMs) in everyday-life products raises concerns for their possible release into the environment and consequent impact on organisms. GRMs have widely varying effects on plants and, according to recent evidences, graphene oxide (GO) has the potential to interfere with the sexual reproduction owing to its acidic properties and production residues. Here, stigmas of the model plant Cucurbita pepo (summer squash) were subjected to simulated dry depositions of GO and GO purified from production residues (PGO). Stigmas were then hand-pollinated and GRM deposition was checked by ESEM and confocal microscopy. Analysis of stigma integrity, pH homeostasis and pollen-stigma interactions did not reveal negative effects. Fruit and seed production were not affected, but GO depositions of 22.1 ± 7.2 ng mm−2 affected the normal development of seeds, decreasing seed dimensions, seed germination and germination speed. The elemental analysis revealed that GO has significant quantities of production residues, such as strong acids and oxidants, while PGO has only traces, which justifies the differences observed in the effects caused by the two materials. Our results show that GO depositions of up to 11.1 ± 3.6 ng mm−2, which fall within the variation range of total dry particulate matter depositions reported in the literature, are safe for reproduction of C. pepo. This is the first “safety” limit ever recorded for depositions of “out-of-the-box” GO concerning the reproduction of a seed plant. If confirmed for wind-pollinated species, it might be considered for policymaking of GRMs emissions in the air.

The ZmMYB84-ZmPKSB regulatory module controls male fertility through modulating anther cuticle-pollen exine trade-off in maize anthers.

Anther cuticle and pollen exine are two crucial lipid layers that ensure normal pollen development and pollen-stigma interaction for successful fertilization and seed production in plants. Their formation processes share certain common pathways of lipid biosynthesis and transport across four anther wall layers. However, molecular mechanism underlying a trade-off of lipid-metabolic products to promote the proper formation of the two lipid layers remains elusive. Here, we identified and characterized a maize male-sterility mutant pksb, which displayed denser anther cuticle but thinner pollen exine as well as delayed tapetal degeneration compared with its wild type. Based on map-based cloning and CRISPR/Cas9 mutagenesis, we found that the causal gene (ZmPKSB) of pksb mutant encoded an endoplasmic reticulum (ER)-localized polyketide synthase (PKS) with catalytic activities to malonyl-CoA and midchain-fatty acyl-CoA to generate triketide and tetraketide α-pyrone. A conserved catalytic triad (C171, H320 and N353) was essential for its enzymatic activity. ZmPKSB was specifically expressed in maize anthers from stages S8b to S9-10 with its peak at S9 and was directly activated by a transcription factor ZmMYB84. Moreover, loss function of ZmMYB84 resulted in denser anther cuticle but thinner pollen exine similar to the pksb mutant. The ZmMYB84-ZmPKSB regulatory module controlled a trade-off between anther cuticle and pollen exine formation by altering expression of a series of genes related to biosynthesis and transport of sporopollenin, cutin and wax. These findings provide new insights into the fine-tuning regulation of lipid-metabolic balance to precisely promote anther cuticle and pollen exine formation in plants.

Proteomic Analysis of Pollen–Stigma Interaction between Vitis rotundifolia and Vitis vinifera

Muscadine grape (Vitis rotundifolia) is highly resistant to many diseases and insects that attack european grape (Vitis vinifera). However, distant hybridization incompatibility between V. rotundifolia (female) and V. vinifera (male) impedes the utilization of V. rotundifolia in grape breeding. This study used fourth-dimension label-free protein quantitation to detect the key genes and pathways in the V. rotundifolia stigma after self-pollination (V. rotundifolia × V. rotundifolia) and cross-pollination (V. rotundifolia × V. vinifera). A histological analysis showed that pollen tube growth in the stigma of V. rotundifolia was arrested 8 hours after cross-pollination, but not after self-pollination. A proteomic analysis identified 32 differentially expressed proteins (DEPs) in the stigma of V. rotundifolia between self-pollination and cross-pollination. A heatmap analysis grouped these DEPs into four clusters. The top gene ontology terms were ATPase-coupled transmembrane transporter activity, extracellular region, DNA replication, and cellular carbohydrate biosynthetic process. A Kyoto Encyclopedia of Genes and Genomes analysis revealed that these DEPs participated in DNA replication and starch and sucrose metabolism pathways. The downregulated A5AY88, D7TJ35, D7SU26, F6HJI1, and F6GUE7 may have a role in cross incompatibility. This study revealed the cross incompatibility of grapes at histological and proteomic levels.

Is airborne graphene oxide a possible hazard for the sexual reproduction of wind-pollinated plants?

Products containing graphene-related materials (GRMs) are becoming increasingly common, allowing GRM nanoparticles (NPs) to enter the environment during their life cycle. Thanks to their lightness and bidimensional geometry, GRM NPs can be easily dispersed in the air and travel very long distances. The flowers of wind-pollinated plants may be exposed to airborne GRMs, being apt to intercept pollen from the air and, inevitably, other airborne particles. Here, stigmas of four wind-pollinated plants (Corylus avellana, common hazel; Juglans regia, walnut; Quercus ilex, holm oak; Zea mays, maize) were exposed to airborne graphene oxide (GO) and GO purified from production residues (PGO) at a concentration of 3.7 ng m−3. Subsequently, the stigmas were pollinated and the adhesion of GOs and their effects on stigma integrity and pollen-stigma interaction were examined. The effect of GO NPs in presence of liquid water on the stigma of C. avellana was also investigated. GOs NPs were intercepted by all species, but their effect varied among them. GO reduced pollen adhesion in J. regia and Q. ilex, whereas pollen germination was unaffected in all four species. The presence of a film of water neither completely removed GO NPs from the stigma, nor it enhanced the toxic effect of GO acidity. PGO never affected pollen-stigma interaction, indicating that the phytotoxic substances used for the production of GO, still in traces in commercial GO, are the main cause of GO toxicity. These results reconfirm the need to verify GRMs effects also on key biological processes beside single model organisms.

Stigmatic Transcriptome Analysis of Self-Incompatible and Compatible Pollination in Corylus heterophylla Fisch. × Corylus avellana L.

Self-incompatibility (SI) protects plants from inbreeding depression due to self-pollination and promotes the outcrossing process to maintain a high degree of heterozygosity during evolution. Corylus is an important woody oil and nut species that shows sporophytic SI (SSI). Yet the molecular mechanism of SI in Corylus remains largely unknown. Here we conducted self- (“Dawei” × “Dawei”) and cross-pollination (“Dawei” × “Liaozhen No. 7”) experiments and then performed an RNA-Seq analysis to investigate the mechanism of pollen–stigma interactions and identify those genes that may be responsible for SSI in Corylus. We uncovered 19,163 up- and 13,314 downregulated genes from the comparison of different pollination treatments. These differentially expressed genes (DEGs) were significantly enriched in plant–pathogen interaction, plant hormone signal transduction, and MAPK signaling pathway–plant. We found many notable genes potentially involved in pollen–stigma interactions and SSI mechanisms, including genes encoding receptor-like protein kinases (RLK), calcium-related genes, disease-resistance genes, and WRKY transcription factors. Four upregulated and five downregulated DEGs were consistently identified in those comparison groups involving self-incompatible pollination, suggesting they had important roles in pollen–pistil interactions. We further identified the S-locus region of the Corylus heterophylla genome based on molecular marker location. This predicted S-locus contains 38 genes, of which 8 share the same functional annotation as the S-locus genes of Corylus avellana: two PIX7 homologous genes (EVM0002129 and EVM0025536), three MIK2 homologous genes (EVM0002422, EVM0005666, and EVM0009820), one aldose 1-epimerase (EVM0002095), one 3-dehydroquinate synthase II (EVM0021283), and one At3g28850 homologous gene (EVM0016149). By characterizing the pistil process during the early postpollination phase via transcriptomic analysis, this study provides new knowledge and lays the foundation for subsequent analyses of pollen-pistil interactions.