Inhibition Of Pollen Tube Growth Research Articles

Phase separation of S-RNase promotes self-incompatibility in Petunia hybrida.

Self-incompatibility (SI) is an intraspecific reproductive barrier widely present in angiosperms. The SI system with the broadest occurrence in angiosperms is based on an S-RNase linked to a cluster of multiple S-locus F-box (SLF) genes found in the Solanaceae, Plantaginaceae, Rosaceae, and Rutaceae. Recent studies reveal that non-self S-RNase is degraded by the Skip Cullin F-box (SCF)SLF-mediated ubiquitin-proteasome system in a collaborative manner in Petunia, but how self-RNase functions largely remains mysterious. Here, we show that S-RNases form S-RNase condensates (SRCs) in the self-pollen tube cytoplasm through phase separation and the disruption of SRC formation breaks SI in self-incompatible Petunia hybrida. We further find that the pistil SI factors of a small asparagine-rich protein HT-B and thioredoxin h together with a reduced state of the pollen tube all promote the expansion of SRCs, which then sequester several actin-binding proteins, including the actin polymerization factor PhABRACL, the actin polymerization activity of which is reduced by S-RNase in vitro. Meanwhile, we find that S-RNase variants lacking condensation ability fail to recruit PhABRACL and are unable to induce actin foci formation required for pollen tube growth inhibition. Taken together, our results demonstrate that phase separation of S-RNase promotes SI response in P. hybrida, revealing a new mode of S-RNase action.

Self S-RNase reduces the expression of two pollen-specific COBRA genes to inhibit pollen tube growth in pear

Due to self-incompatibility (SI) prevents self-fertilization, natural or artificial cross-pollination has been conducted in many orchards to stabilize fruit yield. However, it is still puzzled which routes of self S-RNase arresting pollen tube growth. Herein, 17 COBRA genes were isolated from pear genome. Of these genes, the pollen-specifically expressed PbCOB.A.1 and PbCOB.A.2 positively mediates pollen tube growth. The promoters of PbCOB.A.1 and/or PbCOB.A.2 were bound and activated by PbABF.E.2 (an ABRE-binding factor) and PbC2H2.K16.2 (a C2H2-type zinc finger protein). Notably, the expressions of PbCOB.A.1, PbCOB.A.2, and PbC2H2.K16.2 were repressed by self S-RNase, suggesting that self S-RNase reduces the expression of PbCOB.A.1 and PbCOB.A.2 by decreasing the expression of their upstream factors, such as PbC2H2.K16.2, to arrest pollen tube growth. PbCOB.A.1 or PbCOB.A.2 accelerates the growth of pollen tubes treated by self S-RNase, but can hardly affect level of reactive oxygen species and deploymerization of actin cytoskeleton in pollen tubes and cannot physically interact with any reported proteins involved in SI. These results indicate that PbCOB.A.1 and PbCOB.A.2 may not relieve S-RNase toxicity in incompatible pollen tube. The information provides a new route to elucidate the arresting pollen tube growth during SI reaction.Graphical

VAMP726 and VAMP725 regulate vesicle secretion and pollen tube growth in Arabidopsis.

VAMP726/VAMP725 and SYP131 can form a part of a SNARE complex to mediate vesicle secretion at the pollen tube apex. Secretory vesicle fusion with the plasma membrane of the pollen tube tip is a key step in pollen tube growth. Membrane fusion was mediated by SNAREs. However, little is known about the composition and function of the SNARE complex during pollen tube tip growth. In this study, we constructed a double mutant vamp725 vamp726 via CRISPR‒Cas9. Fluorescence labeling combined with microscopic observation, luciferase complementation imaging, co-immunoprecipitation and GST pull-down were applied in the study. We show that double mutation of the R-SNAREs VAMP726 and VAMP725 significantly inhibits pollen tube growth in Arabidopsis and slows vesicle exocytosis at the apex of the pollen tube. GFP-VAMP726 andVAMP725-GFP localize mainly to secretory vesicles and the plasma membrane at the apex of the pollen tube. In addition, fluorescence recovery after photobleaching (FRAP) experiments showed that mCherry-VAMP726 colocalizes with Qa-SNARESYP131 in the central region of the pollen tube apical plasma membrane. Furthermore, we found that VAMP726 and VAMP725 can interact with the SYP131. Based on these results, we suggest that VAMP726/VAMP725 and SYP131 can form a part of a SNARE complex to mediate vesicle secretion at the pollen tube apex, and vesicle secretion may mainly occur at the central region of the pollen tube apical plasma membrane.

Reproductive systems and hybridization of Lymania species (Bromeliaceae) endemic to Northeast Brazil threatened with extinction

Reproductive systems among the Bromeliaceae family are very diverse at the species level, including allogamous, autogamous, cleistogamous, agamospermic or mixed plants. Hybridization allows the creation of new genotypes with different combinations of shapes and colors to meet the ornamental plant market's demand for novelty. Knowing the reproductive strategies of the species makes it possible to obtain these new materials with ornamental potential, in addition to being essential to define better conservation strategies. Little is known about the reproductive biology of Lymania species. In addition, at least 60% of these species are on the Brazilian National List of Species Threatened with Extinction. The purpose of this study was to identify the reproductive systems, the possible interspecific hybridizations, and the possible reproductive barriers of nine Lymania species based on the germination of pollen grains and the corresponding growth of the pollen tube in the pistil by using fluorescence microscopy with ultraviolet filtering. Natural and controlled pollinations of the nine species were carried out. The reproductive strategy was defined according to the autogamy index (AI), self-incompatibility index (SII), and reproductive efficiency index (REI). All species were self-incompatible except for L. smithii, which was autogamous. Agamospermy was not observed in any of the studied species. Forty-one hybridization combinations between species were performed, resulting in a 54% success rate, with incongruity in only 12 combinations. The main incongruence phenomena identified after interspecific crosses were: poor germination of pollen grains on the stigma surface; inhibition of pollen tube growth along the style; coiling of pollen tubes along the style; pollen tubes inside the ovary but without penetration into the micropyle; and reticulate deposition of callose in the pollen tube, among others. We suggest that self-incompatibility of the species is gametophytic, preventing self-fertilization. Results showed that it is possible to perform hybridization with some species. However, morphological, genetic, and molecular studies are needed to confirm the potential of these species. Seeds generated in the different reproductive systems, as well as in the hybridizations, had germination rates above 94%. The results presented here make important contributions to the conservation of species, seed generation, and hybridization of Bromeliaceae.

Identification and analysis of CdS-RNase in Camellia drupifera: A key determinant of late-acting self-incompatibility

Plant self-incompatibility (SI) is of great significance in reproductive biology and is also the main factor affecting yield in agricultural production. Late-acting self-incompatibility (LSI) is one of the main types of plant SI, but the mechanism of LSI is not systematically analyzed. Tea oil Camellia, a strictly outcrossing plant, is a well-known woody oil cash crop. SI is one of the main reasons for the low yield of tea oil Camellia. In this study, we found that C. drupifera, one of the main species of tea oil Camellia, shows LSI, in which SI occurs in the style, and pollen tube growth in the ovary ceases at 96 h after selfing. The CdS-RNase gene of Camellia drupifera was successfully cloned, and the gene showed a length of 922 bp and encoded 238 amino acids. CdS-RNase had typical S-RNase characteristics. The functional validation results revealed that CdS-RNase was a key female factor for SI in C. drupifera, indicating that S-RNase plays an important role in the plant LSI mechanism. CdS-RNase localized to the cell membrane, was specifically expressed in the style and transferred to the ovary, and inhibited pollen tube growth in the style and ovary. There was an "enhancer factor" for the expression of CdS-RNase in selfing styles. Between 24 h and 48 h after selfing, CdS-RNase expression was significantly upregulated 28.3 times, which was more than 2.5 times higher than the outcross expression level and more than 5 times lower than the cross expression level. A total of 28 CdS-RNase proteins were identified, including 6 CdSm-RNases, and outcrossing among different CdS-RNase genotypes had a significant effect on the fruit setting rate. It may be that mismatches among CdS-RNase genotypes and self-inbreeding are the main reasons for the low yield of C. drupifera. These results provide a basis for the systematic analysis of the LSI mechanism and for solving the SI problem of tea oil Camellia.

Ultrastructural Evidence Elucidates the Mode of Action of Sulfur in Preventing Pollen Tube Development in Stigma of Citrus cv. Nadorcott and Other Horticultural Species

Citrus species have a “wet”-type stigma with abundant exudate, and their style contains numerous canals. For successful seed formation, the process includes pollen grain germination on the stigma, pollen tube development and growth through the stigma and style and, ultimately, successful ovule fertilization. However, preventing the fertilization process can be useful for many agronomic and plant-breeding purposes, such as seedless fruits or for developing new varieties. Several studies have recently shown the inhibition effect of sulfur on pollen tube development inside the Nadorcott mandarin stigma and its effective application to obtain seedless mandarins. However, when applied to the stigma, how can sulfur inhibit pollen tube growth? Moreover, does sulfur have the same effect on other species? The main objective of the present study is to clarify the mode of action of sulfur on the ultrastructure of the Nadorcott mandarin stigma and style. To fulfill this goal, untreated flowers and flowers treated with sulfur were pollinated 24 h later. The treated and untreated stigmas were analyzed and compared with several microscopy techniques. The main results showed that sulfur specifically caused an alteration to the outer layer of stigma papillary cells. This marked alteration resulted in papillary cells losing their functionality due to the deterioration and degradation of their cellular structure. Basal papillae, the stigmatic tissue and stylar canals also underwent major alteration. Sulfur also modified the quantity and uniformity distribution of the stigmatic exudate. All these alterations collectively prevented pollen tube development inside the stigma. These effects have been observed in several Citrus species and varieties, and in some other horticultural species, which suggests a generic (non species-specific) action.

PbrBZR1 interacts with PbrARI2.3 to mediate brassinosteroid-regulated pollen tube growth during self-incompatibility signaling in pear.

S-RNase-mediated self-incompatibility (SI) prevents self-fertilization and promotes outbreeding to ensure genetic diversity in many flowering plants, including pear (Pyrus sp.). Brassinosteroids (BRs) have well-documented functions in cell elongation, but their molecular mechanisms in pollen tube growth, especially in the SI response, remain elusive. Here, exogenously applied brassinolide (BL), an active BR, countered incompatible pollen tube growth inhibition during the SI response in pear. Antisense repression of BRASSINAZOLE-RESISTANT1 (PbrBZR1), a critical component of BR signaling, blocked the positive effect of BL on pollen tube elongation. Further analyses revealed that PbrBZR1 binds to the promoter of EXPANSIN-LIKE A3 (PbrEXLA3) to activate its expression. PbrEXLA3 encodes an expansin that promotes pollen tube elongation in pear. The stability of dephosphorylated PbrBZR1 was substantially reduced in incompatible pollen tubes, where it is targeted by ARIADNE2.3 (PbrARI2.3), an E3 ubiquitin ligase that is strongly expressed in pollen. Our results show that during the SI response, PbrARI2.3 accumulates and negatively regulates pollen tube growth by accelerating the degradation of PbrBZR1 via the 26S proteasome pathway. Together, our results show that an ubiquitin-mediated modification participates in BR signaling in pollen and reveal the molecular mechanism by which BRs regulate S-RNase-based SI.

Self S-RNase restricts ABF-LRX signaling cascade to arrest pollen tube during pear self-incompatibility reaction.

Gametophytic self-incompatibility (GSI) has been widely studied in flowering plants, but the study on mechanism of arresting pollen tube growth by self S-RNase in the GSI species is limited. In this study, two leucine-rich repeat extensin genes, PbLRXA2.1 and PbLRXA2.2, were identified based on transcriptome and quantitative real-time PCR analyses. The expression levels of these two LRX genes were significantly higher in the pollen grains and pollen tubes of self-compatible cultivar 'Jinzhui' (a spontaneous bud mutation of the self-incompatible cultivar 'Yali') than in 'Yali'. Both PbLRXA2.1 and PbLRXA2.2 positively mediated pollen tube growth and attenuated self S-RNase toxicity on pollen tube growth by stabilizing actin cytoskeleton and cell wall integrity. This result indicates that the abnormal expression of PbLRXA2.1 and PbLRXA2.2 involve in the breakdown of self-incompatibility in 'Jinzhui'. The PbLRXA2.1 and PbLRXA2.2 promoters were directly bound by an ABRE-binding factor PbABF.D.2. Knockdown of PbABF.D.2 expression decreased PbLRXA2.1 and PbLRXA2.2 expression and inhibited pollen tube growth. Notably, the expressions of PbLRXA2.1, PbLRXA2.2, and PbABF.D.2 were repressed by self S-RNase, suggesting that self S-RNase can arrest pollen tube growth by restricting the PbABF.D.2-PbLRXA2.1/PbLRXA2.2 signal cascade. These results provide a novel insight of the pollen tube growth arrested by self S-RNase.

Optimization of culture medium for in vitro germination and storage conditions of Exochorda racemosa pollen.

Pollen morphology, pollen vigor, and long-term pollen storage are critical for plant cross-breeding and genetic improvement of Exochorda racemosa. We developed a protocol for viability determination and storage of E. racemosa pollen for breeding new varieties. The medium components for E. racemosa pollen germination was optimized by using an Orthogonal Assay Test Strategy (OATS). The germination rates of E. racemosa pollen were investigated after storing at different temperatures and different storage periods. The size of E. racemosa pollen was medium with three germination ditches, and the sculptural type of pollen was striate. Red ink and 2,3,5-triphenyl tetrazolium chloride (TTC) can effectively distinguish viable pollen from the unviable pollen of E. racemosa. The most suitable medium (CK2) for E. racemosa was composed of 150 g· L–1 sucrose, 100 mg·L–1 boric acid, 150 mg· L–1 Ca(NO3)2 and 50 mg· L–1 GA3. Low-temperature stress produced the greater inhibition of pollen tube growth compared with high-temperature conditions. The CK2 medium at pH 6.5 resulted in the highest pollen germination rate and most extended pollen tube length. The optimal temperature for storage of dried pollen was –80°C (P < 0.01), and the germination rate was 53.60% after storage for 390 days. Thawing in a 35°C water bath produced the best viability of E. racemosa pollen after storage at –20°C and –80°C. The short-term storage of E. racemosa fresh pollen at 4°C was better than that at –20°C and –80°C (P < 0.01). It is possible to evaluate pollen quality and store pollen grains for E. racemosa by the parameters defined in this study.

A novel hybrid seed production technology based on a unilateral cross-incompatibility gene in maize.

Hybrid seed production technology (SPT) using genic recessive male sterility is of great importance in maize breeding. Here, we report a novel SPT based on a maize unilateral cross-incompatibility gene ZmGa1F with an extremely low transgene transmission rate (TTR). Proper pollen-specific ZmGa1F expression severely inhibits pollen tube growth leading to no fertilization. The maintainer line harbors a transgene cassette in an ipe1 male sterile background containing IPE1 to restore ipe1 male fertility, ZmGa1F to prevent transgenic pollen escape, the red fluorescence protein encoding gene DsRed2 for the separation of male sterile and fertile seeds, and the herbicide-resistant gene Bar for transgenic plant selection. When the maintainer line is selfed, gametes of ipe1/transgene and ipe1/- genotypes are produced, and pollen of the ipe1/transgene genotype is not able to fertilize female gametes due to pollen tube growth inhibition by ZmGa1F. Subsequently, seeds of ipe1/ipe1 and ipe1/transgene genotypes are produced at a 1:1 ratio and could be separated easily by fluorescence-based seed sorting. Not a single seed emitting fluorescence is detected in more than 200,000 seeds examined demonstrating that the pollen-tube-inhibition (PTI)-based TTR is lower than what has been reported for similar technologies to date. This PTI-based SPT shows promising potential for future maize hybrid seed production.

Combined transcriptomic and proteomic analysis reveals multiple pathways involved in self-pollen tube development and the potential roles of FviYABBY1 in self-incompatibility in Fragaria viridis.

Fragaria viridis exhibits S-RNase-based gametophytic self-incompatibility, in which S-RNase is the major factor inhibiting pollen tube growth. However, the pathways involved in and the immediate causes of the inhibition of pollen tube growth remain unknown. Here, interactive RNA sequencing and proteome analysis revealed changes in the transcriptomic and proteomic profiles of F. viridis styles harvested at 0 and 24 h after self-pollination. A total of 2,181 differentially expressed genes and 200 differentially abundant proteins were identified during the pollen development stage of self-pollination. Differentially expressed genes and differentially abundant proteins associated with self-incompatible pollination were further mined, and multiple pathways were found to be involved. Interestingly, the expression pattern of the transcription factor FviYABBY1, which is linked to polar growth, differed from those of other genes within the same family. Specifically, FviYABBY1 expression was extremely high in pollen, and its expression trend in self-pollinated styles was consistent with that of S-RNase. Furthermore, FviYABBY1 interacted with S-RNase in a non-S haplotype way. Therefore, FviYABBY1 affects the expression of polar growth-related genes in self-pollen tubes and is positively regulated by S-RNase.

Nitric Oxide Participates in Aluminum-Stress-Induced Pollen Tube Growth Inhibition in Tea (Camelliasinensis) by Regulating CsALMTs.

Nitric oxide (NO), as a signal molecule, is involved in the mediation of heavy-metal-stress-induced physiological responses in plants. In this study, we investigated the effect of NO on Camellia sinensis pollen tubes exposed to aluminum (Al) stress. Exogenous application of the NO donor decreased the pollen germination rate and pollen tube length and increased the malondialdehyde (MDA) content and antioxidant enzyme activities under Al stress. Simultaneously, the NO donor effectively increased NO content in pollen tube of C. sinensis under Al stress and could aggravate the damage of Al3+ to C. sinensis pollen tubes by promoting the uptake of Al3+. In addition, application of the NO-specific scavenger significantly alleviated stress damage in C. sinensis pollen tube under Al stress. Moreover, 18 CsALMT members from a key Al-transporting gene family were identified, which could be divided into four subclasses. Pearson correlation analysis showed the expression level of CsALMT8 showed significant positive correlation with the Al3+ concentration gradient and NO levels, but a significant negative correlation with pollen germination rate and pollen tube length. The expression level of CsALMT5 was negatively correlated with the Al3+ concentration gradient and NO level, and positively correlated with pollen germination rate and pollen tube length. The expression level of CsALMT17 showed a significant negative correlation with Al3+ concentration and NO content in pollen tubes, but significant positive correlation with pollen germination rate and pollen tube length. In conclusion, a complex signal network regulated by NO-mediated CsALMTs revealed that CsALMT8 was regulated by environmental Al3+ and NO to assist Al3+ entry into pollen tubes; CsALMT5 might be influenced by the Al3+ signal, stimulate malate efflux in vacuoles and chelate with Al3+ to detoxify Al in C. sinensis pollen tube.

Cellulose accumulation mediated by PbrCSLD5, a cellulose synthase-like protein, results in cessation of pollen tube growth in Pyrus bretschneideri.

Cellulose, a key component of the cell wall, plays an important role in maintaining the growth of pollen tubes. However, the molecular mechanism of cellulose participating in the cessation of pear pollen tube growth remains unclear. Here, we reported that at 15 h post-cultured (HPC), the slow-growth pear pollen tubes showed thickened cell walls and cellulose accumulation in the inner wall. Transcriptome data and quantitative real-time PCR analysis showed that PbrCSLD5, a cellulose synthesis-like gene, was highly expressed in the 15 HPC pear pollen tubes. Knockdown of PbrCSLD5 caused a decrease in cellulose content in pear pollen tubes. Moreover, PbrCSLD5 overexpression in Arabidopsis resulted in the accumulation of cellulose and disruption of normal pollen tube growth. Transcription factor PbrMADS52 was found to bind to the promoter of PbrCSLD5 and enhanced its expression. Our results suggested that the PbrMADS52-PbrCSLD5 signaling pathway led to increased cellulose content in the pear pollen tube cell wall, thereby inhibiting pollen tube growth. These results provided new insights into the regulation of pollen tube growth.

Agronomic treatments to avoid seed presence in ‘Nadorcott’ mandarin I. Effect on in vivo pollen tube growth

Fresh market demands high quality fruit and, therefore, citrus growers and researchers are constantly looking for solutions to avoid seed presence. Current horticultural techniques have low effectiveness or high cost. The objective of this study is to evaluate the inhibition effect of seven products on the in vivo pollen tube growth in Nadorcott mandarin, which is a high-value seedy variety. To achieve this main objective, three inorganic fertilizers (ammonium nitrate, potassium nitrate, sulfur), and four saccharides (saccharose, methyl cellulose, callose, chitosan) were applied to Nadorcott stigmas 24 h before and after hand pollination. Pollen tubes were counted 1, 3, and 5 days after treatment in longitudinal blue violet autofluorescence-stained sections. Of the seven evaluated products, only sulfur had a strong inhibitory effect. Elemental sulfur (S8) inhibited pollen tube growth by 94-100%. This strong effect was observed regardless of sulfur being applied 24 h before or after pollination, and on fixed flowers 1, 3 or 5 days after applications. Saccharose treatment seemed to have the opposite effect: stimulated pollen tube growth, but the difference with the positive control was small and non-significant. The sulfur effect could be useful for designing agronomic applications capable of preventing seed presence in Nadorcott mandarin.

PbrRALF2-elicited reactive oxygen species signaling is mediated by the PbrCrRLK1L13-PbrMPK18 module in pear pollen tubes

Rapid alkalinization factors (RALFs) are cysteine-rich peptides that play important roles in a variety of biological processes, such as cell elongation and immune signaling. Recent studies in Arabidopsis have shown that RALFs regulate pollen tube growth via plasma membrane receptor-like kinases (RLKs). However, the downstream signal transduction mechanisms of RLKs in pollen tubes are unknown. Here, we identified PbrRALF2, a pear (Pyrus bretschneideri) pollen RALF peptide that inhibits pollen tube growth. We found that PbrRALF2 interacts with a malectin-like domain-containing RLK, PbrCrRLK1L13. The relative affinity between PbrRALF2 and PbrCrRLK1L13 was at the submicromolar level, which is consistent with the values of ligand–receptor kinase pairs and the physiological concentration for PbrRALF2-mediated inhibition of pollen tube growth. After binding to its extracellular domain, PbrRALF2 activated the phosphorylation of PbrCrRLK1L13 in a dose-dependent manner. We further showed that the MAP kinase PbrMPK18 is a downstream target of PbrCrRLK1L13 that mediates PbrRALF2-elicited reactive oxygen species (ROS) production. The excessive accumulation of ROS inhibits pollen tube growth. We show that MPK acts as a mediator for CrRLK1L to stimulate ROS production, which might represent a general mechanism by which RALF and CrRLK1L function in signaling pathways.

Identification and expression analysis of the PbrMLO gene family in pear, and functional verification of PbrMLO23

Mildew resistance locus O (MLO) is a plant-specific gene family that plays an important role in the growth and development of plants and their interactions with the environment. However, the available information on this gene family in pear is limited. Here, 24 PbrMLO genes were identified and divided into five subfamilies (I, II, III, IV and V). Whole-genome duplication (WGD) and dispersed duplication contributed to the expansion of the PbrMLO family. In addition, gene expression analysis revealed that PbrMLO genes were distributed in various pear tissues, suggesting their diverse functions. We selected PbrMLO23 for further functional analysis. Expression profile analysis by qRT-PCR showed that PbrMLO23 was highly expressed in pollen. Subcellular localization analysis showed that PbrMLO23 was located on the plasma membrane. When the expression level of PbrMLO23 was knocked down by using antisense oligonucleotides, pollen tube lengths increased, indicating that PbrMLO23 plays a functional role in inhibiting pollen tube growth. In summary, these results provide evolutionary insight into PbrMLO and its functional characteristics and lay a foundation for further analysis of the functions of PbrMLO members in pear.

Identification of pollen and pistil polygalacturonases in Nicotiana tabacum and their function in interspecific stigma compatibility

The pollen and pistil polygalacturonases in Nicotiana tabacum were identified and found to regulate pollen tube growth and interspecific compatibility. Polygalacturonase (PG) is one of the enzymes catalyzing the hydrolysis of pectin. This process plays important roles in the pollen and pistil. In this research, the pollen and pistil PGs in Nicotiana tabacum (NtPGs) were identified, and their expression, localization and the potential function in the pollen and interspecific stigma incompatibility were explored. The results showed that 118 NtPGs were retrieved from the genome of N. tabacum. The phylogenetic tree and RT-qPCR analysis led to the identification of 10 pollen PGs; among them, two, seven and one showed specifically higher expression levels in the early development of anthers, during pollen maturation and in mature anthers, respectively, indicating their function difference. Immunofluorescence analysis showed that PGs were located in the cytoplasm of (1) mature pollen and (2) in vitro grown pollen tubes, as well as in the wall of in vivo grown pollen tubes. Four NtPGs in clade A were identified as the pistil PGs, and the pistil PGs were not found in clade E. Significantly higher PGs expression was recorded after incompatible pollination in comparison with the compatible stigma, indicating a potential function of PGs in regulating stigma incompatibility. The influence of PGs on pollen tube growth was explored in vitro and partly in vivo, showing that high PGs activity inhibited pollen tube growth. The application of PGs on the otherwise compatible stigma resulted in pollen tube growth inhibition or failure of germination. These results further supported that increased PGs expression in incompatible stigma might be partially responsible for the interspecific stigma incompatibility in Nicotiana.

Ectopic Expression of a Self-Incompatibility Module Triggers Growth Arrest and Cell Death in Vegetative Cells.

Self-incompatibility (SI) is used by many angiosperms to reject self-pollen and avoid inbreeding. In field poppy (Papaver rhoeas), SI recognition and rejection of self-pollen is facilitated by a female S-determinant, PrsS, and a male S-determinant, PrpS PrsS belongs to the cysteine-rich peptide family, whose members activate diverse signaling networks involved in plant growth, defense, and reproduction. PrsS and PrpS are tightly regulated and expressed solely in pistil and pollen cells, respectively. Interaction of cognate PrsS and PrpS triggers pollen tube growth inhibition and programmed cell death (PCD) of self-pollen. We previously demonstrated functional intergeneric transfer of PrpS and PrsS to Arabidopsis (Arabidopsis thaliana) pollen and pistil. Here, we show that PrpS and PrsS, when expressed ectopically, act as a bipartite module to trigger a self-recognition:self-destruct response in Arabidopsis independently of its reproductive context in vegetative cells. The addition of recombinant PrsS to seedling roots expressing the cognate PrpS resulted in hallmark features of the P rhoeas SI response, including S-specific growth inhibition and PCD of root cells. Moreover, inducible expression of PrsS in PrpS-expressing seedlings resulted in rapid death of the entire seedling. This demonstrates that, besides specifying SI, the bipartite PrpS-PrsS module can trigger growth arrest and cell death in vegetative cells. Heterologous, ectopic expression of a plant bipartite signaling module in plants has not been shown previously and, by extrapolation, our findings suggest that cysteine-rich peptides diversified for a variety of specialized functions, including the regulation of growth and PCD.

Screening Thermal Shock as an Apple Blossom Thinning Method. I. Stigmatic Receptivity, Pollen Tube Growth, and Leaf Injury in Response to Temperature and Timing of Thermal Shock

The use of short-duration applications of thermal energy (thermal shock; TS) as an apple blossom thinning strategy was investigated. Effects of TS temperature and timing on stigmatic receptivity, pollen tube growth in vivo, and visible leaf injury were evaluated in multiple experiments on ‘Crimson Gala’. TS treatments were applied to blossoms and spur leaves using a variable temperature heat gun. TS temperatures ≥86 °C had a strong inhibitory effect on pollen tube growth on the stigmatic surface and in the style. TS temperatures &gt;79 °C reduced average pollen tube length to less than the average style length. Timing of TS treatment (0 or 24 hours after pollination) was not an influential factor, indicating that effective TS temperatures reduced pollen tube growth up to 24 hours after the pollination event. The onset of thermal injury to vegetative tissues occurred at similar TS temperatures that inhibited pollen tube growth in vivo. Excessive leaf injury (&gt;33%) was observed at 95 °C, suggesting relatively narrow differences in thermal sensitivity between reproductive and vegetative tissues. Inconsistent TS temperatures and/or responses were observed in some experiments. Ambient air temperature may have influenced heat gun output temperatures and/or plant susceptibility. While results suggest some promise, additional work is required to validate and further develop this concept.

Germination potential and pollen tube growth in Galanthus nivalis L.

The present research studies the pollen biology of snowdrop (Galanthus nivalis L.), from the point of view of pollen’s germinating potential and length growth of pollen tubes at two temperatures: 18 °C and 4 °C. The germination was conducted “in vitro” on nutritive mediums containing different concentrations of sucrose (0, 5, 10, 15, 20, 25, 40, 50, 70, 100%). The pollen germination was optimum (more than 90 %) on nutritive mediums containing 15 % and 20 % sucrose, at both studied temperatures, after 24 h. In parallel with germination potential, the adaptation of male gametophytes to the two temperatures was analyzed. The longest pollen tubes were formed on 10–25% sucrose mediums at 18 °C and on 10–15% sucrose mediums at 4 °C, and they were maintained also after 120 h since pollen inoculation. For these concentrations, the pollen tube length was similar at both temperatures. In conclusion, 18 °C is suitable for long tubes formation using a largest range of sucrose concentration than 4 °C. In fact, in most plant species, low temperatures not only inhibit pollen tube growth, but also induce flowers abortion. Present results bring new evidences that the snowdrop’s male gametophyte is genetically settled to have a normal development at low temperatures. The optimal germination rate and pollen tube growth at 4 °C highlight the vernal character of the pollen of G. nivalis.