SYP72 interacts with the mechanosensitive channel MSL8 to protect pollen from hypoosmotic shock during hydration

In flowering plants, hydration of desiccated pollen grains on stigma is a prerequisite for pollen germination, during which pollen increase markedly in volume through water uptake, requiring them to survive hypoosmotic shock to maintain cellular integrity. However, the mechanisms behind the adaptation of pollen to this hypoosmotic challenge are largely unknown. Here, we identify the Qc-SNARE protein SYP72, which is specifically expressed in male gametophytes, as a critical regulator of pollen survival upon hypoosmotic shock during hydration. SYP72 interacts with the MSCS-LIKE 8 (MSL8) and is required for its localization to the plasma membrane. Intraspecies and interspecies genetic complementation experiments reveal that SYP72 paralogs and orthologs from green algae to angiosperms display conserved molecular functions and rescue the defects of Arabidopsis syp72 mutant pollen facing hypoosmotic shock following hydration. Our findings demonstrate a critical role for SYP72 in pollen resistance to hypoosmotic shock through the MSL8 cascade during pollen hydration.

Transcriptome Analysis of Triple Mutant for OsMADS62, OsMADS63, and OsMADS68 Reveals the Downstream Regulatory Mechanism for Pollen Germination in Rice (Oryza sativa)

The MADS (MCM1-AGAMOUS-DEFFICIENS-SRF) gene family has a preserved domain called MADS-box that regulates downstream gene expression as a transcriptional factor. Reports have revealed three MADS genes in rice, OsMADS62, OsMADS63, and OsMADS68, which exhibits preferential expression in mature rice pollen grains. To better understand the transcriptional regulation of pollen germination and tube growth in rice, we generated the loss-of-function homozygous mutant of these three OsMADS genes using the CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9) system in wild-type backgrounds. Results showed that the triple knockout (KO) mutant showed a complete sterile phenotype without pollen germination. Next, to determine downstream candidate genes that are transcriptionally regulated by the three OsMADS genes during pollen development, we proceeded with RNA-seq analysis by sampling the mature anther of the mutant and wild-type. Two hundred and seventy-four upregulated and 658 downregulated genes with preferential expressions in the anthers were selected. Furthermore, downregulated genes possessed cell wall modification, clathrin coat assembly, and cellular cell wall organization features. We also selected downregulated genes predicted to be directly regulated by three OsMADS genes through the analyses for promoter sequences. Thus, this study provides a molecular background for understanding pollen germination and tube growth mediated by OsMADS62, OsMADS63, and OsMADS68 with mature pollen preferred expression.

The Effect of Titanium Organic Complex on Pollination Process and Fruit Development of Apple cv. Topaz

Optimal weather conditions are necessary for the proper course of the flowering process and high activity of pollinators, which is the most important factor affecting pollination. Because weather conditions do not always favour effective pollination and because pollinators’ activities are decreasing, the application of compound stimulating pollination and fertilization may be a good perspective for increasing yield. Titanium is considered a beneficial element for plants. Preliminary studies have indicated the positive effect of titanium organic complex on pollen adhesion to the stigma and pollen germination on the stigma of tomato and cucumber. Therefore, a 2-year experiment was designed to determine the effect of titanium organic complex application on the pollination process and fruit development of apple cv. Topaz (Malus domestica Borkh.) The experiment demonstrated the positive effect of titanium organic complex on fertilization of ovules because of effective pollination, seed setting, and fruit development of apple. Application of titanium organic complex improved pollen adhesion to the stigma and pollen germination on the stigma. In addition, titanium organic complex increased the number of pollen tubes growing through the pistil style, which resulted in fertilization that was more effective, as confirmed by the higher number of seeds set in fruits. Higher numbers of seeds set in fruits positively affected their weight and size. Therefore, fruit harvested from trees to which titanium organic complex was applied were characterized by greater weight, length, and diameter compared to fruits obtained from the trees to which titanium organic complex was not applied.

Comprehensive Analysis of Arabinogalactan Protein-Encoding Genes Reveals the Involvement of Three BrFLA Genes in Pollen Germination in Brassica rapa

Arabinogalactan proteins (AGPs) are a superfamily of hydroxyproline-rich glycoproteins that are massively glycosylated, widely implicated in plant growth and development. No comprehensive analysis of the AGP gene family has been performed in Chinese cabbage (Brassica rapa ssp. chinensis). Here, we identified a total of 293 putative AGP-encoding genes in B. rapa, including 25 classical AGPs, three lysine-rich AGPs, 30 AG-peptides, 36 fasciclin-like AGPs (FLAs), 59 phytocyanin-like AGPs, 33 xylogen-like AGPs, 102 other chimeric AGPs, two non-classical AGPs and three AGP/extensin hybrids. Their protein structures, phylogenetic relationships, chromosomal location and gene duplication status were comprehensively analyzed. Based on RNA sequencing data, we found that 73 AGP genes were differentially expressed in the floral buds of the sterile and fertile plants at least at one developmental stage in B. rapa, suggesting a potential role of AGPs in male reproductive development. We further characterized BrFLA2, BrFLA28 and BrFLA32, three FLA members especially expressed in anthers, pollen grains and pollen tubes. BrFLA2, BrFLA28 and BrFLA32 are indispensable for the proper timing of pollen germination under high relative humidity. Our study greatly extends the repertoire of AGPs in B. rapa and reveals a role for three members of the FLA subfamily in pollen germination.

Intra-Plant Variability for Heat Tolerance Related Attributes in Upland Cotton

Abiotic stress, particularly heat stress, affects various parts of the cotton plant and ultimately impacts the seed cotton yield. Different portions of a single cotton plant of a cultivar exhibit variable responses to stress during reproductive and vegetative phases. To test this hypothesis, physiological and morphological traits related to heat stress were observed for two flowering positions in 13 genotypes of upland cotton. These genotypes were sown in field conditions in triplicate following a randomized complete block design. Data were collected for pollen germination, pollen viability, cell membrane thermostability, chlorophyll content, boll weight, and boll retention for both the top and bottom branches of each genotype. The collected data were analyzed for the identification of variability within and between genotypes for these two flowering positions. Tukey’s test was applied to estimate the significance of differences between genotypes and positions within each genotype. Results showed that the two positions within the same plant statistically varied from each other. The bottom branches of the genotypes performed significantly better for all traits measured except boll weight. The genotype AA-933 performed best for pollen germination and boll retention, while CYTO-608 exhibited maximum pollen viability in both the bottom and top flower positions compared with other genotypes. Overall, MNH-1016 and CIM-602 showed better cell membrane thermostability and chlorophyll content, respectively. This intra-plant variability can be further exploited in breeding programs to enhance the stress tolerance capabilities of the resulting varieties.

Two Carbohydrate-Based Natural Extracts Stimulate in vitro Pollen Germination and Pollen Tube Growth of Tomato Under Cold Temperatures

To date, it is widely accepted by the scientific community that many agricultural regions will experience more extreme temperature fluctuations. These stresses will undoubtedly impact crop production, particularly fruit and seed yields. In fact, pollination is considered as one of the most temperature-sensitive phases of plant development and until now, except for the time-consuming and costly processes of genetic breeding, there is no immediate alternative to address this issue. In this work, we used a multidisciplinary approach using physiological, biochemical, and molecular techniques for studying the effects of two carbohydrate-based natural activators on in vitro tomato pollen germination and pollen tube growth cultured in vitro under cold conditions. Under mild and strong cold temperatures, these two carbohydrate-based compounds significantly enhanced pollen germination and pollen tube growth. The two biostimulants did not induce significant changes in the classical molecular markers implicated in pollen tube growth. Neither the number of callose plugs nor the CALLOSE SYNTHASE genes expression were significantly different between the control and the biostimulated pollen tubes when pollens were cultivated under cold conditions. PECTIN METHYLESTERASE (PME) activities were also similar but a basic PME isoform was not produced or inactive in pollen grown at 8°C. Nevertheless, NADPH oxidase (RBOH) gene expression was correlated with a higher number of viable pollen tubes in biostimulated pollen tubes compared to the control. Our results showed that the two carbohydrate-based products were able to reduce in vitro the effect of cold temperatures on tomato pollen tube growth and at least for one of them to modulate reactive oxygen species production.

A Dual-Successive-Screen Model at Pollen/Stigma and Pollen Tube/Ovary Explaining Paradoxical Self-Incompatibility Diagnosis in the Olive Tree—An Interpretative Update of the Literature

The ‘pollen test’ and ‘fruit set test’ following controlled crossing combinations of parents are the most commonly used methods for pollination incompatibility studies in Olea europaea L. Self-incompatibility (SI), with diagnoses based on the pollen test and pollen germination, indicating self-compatibility, is not always followed by fruit set in this species. To solve this dispute, we have reconciled all observations into a new model. Mismatches between field and laboratory data and between methods are resolved by the dual-successive-screen model (DSSM) supposing two different loci for the expression of the two SI mechanisms. Pollen/stigma is controlled by diallelic SI, or DSI, inferring two G1 and G2 compatibility/incompatibility (C/I) groups for varieties, whereas pollen tubes in ovaries are controlled by poly-allelic SI or PASI with twenty C/I groups. To explain the selfing of varieties, we have suggested that some determinants in the pollen tube and stigma are unstable and degrade (DS-D for degradation of S-determinant) after three to five days, enabling some pollen tubes to avoid being rejected, hence reaching ovules. DSI and PASI in the DSSM and DS-D mechanisms, plus the andromonoecy of the olive tree, complexify SI studies. Inferences from DSSM and DS-D mechanisms in olive orchard practice are detailed.

Fast Drying Combined With Rapid Cooling Can Enhance The Survival of The Desiccation Sensitive Wheat Pollen After Ultra-Low Temperature Storage

Long-term storage of pollen is important for the fertilization of spatially or temporally isolated female parents, especially during hybrid breeding. Wheat pollen is dehydration-sensitive and rapidly loses viability after shedding. To preserve wheat pollen, we hypothesized that fast-(flash)-drying and fast cooling (150°C min-1) compared to slow-(air)-drying and slow cooling (1°C min-1) would increase the rate of intracellular water content (WC) removal, decrease intracellular ice crystal formation, and increase viability after exposure to ultra-low temperatures. High correlations were found between pollen WC and viability analyzed by impedance flow cytometry (IFC viability: r=0.92, P<0.001) and pollen germination (r=0.94, P<0.001). After 10 min of air-drying, 66% WC was lost and pollen germination was at 12.2±12.3%. After 10 min of flash-drying, WC of pollen reduced by 74%. IFC viability decreased from 90.2±6.7 to 39.4±17.9%, and pollen germination dropped from 33.7±16.9 to 1.9±3.9%. After 12 min of flash-drying, WCs decreased to <0.34 mg H2O mg-1 DW, ice crystal formation was completely prevented (ΔH=0 J mg-1 DW), and pollen germination reached 1.2±1.0%. After slow and fast cooling, flash-dried pollen (WC 0.91±0.11 mg H2O mg-1 DW) showed less ice crystal formation during cryomicroscopic-video-recordings and had IFC viability of 4.5±7.0% (slow) and 6.1±8.8% (fast), respectively, compared to air-dried pollen which lost all viability. Generally, fast-(flash)-drying and increased cooling rates may enable the survival of wheat pollen likely due to (1) a fast rate of intracellular WC loss that reduces deleterious biochemical changes associated with the drying process and (2) a delay and reduction in intracellular ice crystal formation.