Tricellular Stage Research Articles

Ultrastructural study of pollen and tapetum development in Hydrocleys nymphoides, Alisma plantago-aquatica, and Sagittaria montevidensis (Alismataceae).

The Alismataceae family, widely distributed across tropical temperate swamps and wetlands, includes 15 genera post-merger with Limnocharitaceae. In Argentina, six genera are represented across three clades. Embryological characters, notably the male gametophyte and anther, are crucial in taxonomy due to their stability against environmental changes. This study aims to analyze the ultrastructure of the tapetum and pollen grain development in three economically and ecologically important species representing each clade: Sagittaria montevidensis (Clade A), Hydrocleys nymphoides (Clade B), and Alisma plantago-aquatica (Clade C). Anthers at different developmental stages were processed according to classic techniques for their observation with bright-field and transmission electron microscopy. The three studied species within the Alismataceae family exhibit similar reproductive characteristics. Seven stages of pollen grain development were identified. The microsporogenesis is successive with a regular meiosis. The ultrastructure of the tapetal cells shows similarities to other species with plasmodial tapetum. During the microspore tetrad stage, there is tapetal hyperactivity and an increase in secretion processes. In the free microspore stage, the tapetal cells lose their walls and increase the amount of rough endoplasmic reticulum forming a network of cisternae that extend into evaginations. Later cells completely invade the anther locule and fuse to form a tapetal plasmodium. No peritapetal membrane with orbicules was observed. Pollen is released at the tricellular stage. The pollen grain wall presents an ectexine with a basal layer, columellae, and tectum with supratectal spines while an endexine is not observed in any of the three species. This research enhances the understanding of tapetal cell interactions with developing pollen grains and contributes to the knowledge of the ultrastructure of plasmodial tapetum. Moreover, these findings highlight evolutionary reproductive patterns in Alismataceae, suggesting the plasmodial tapetum as a synapomorphy for the order.

The transcribed centromeric gene OsMRPL15 is essential for pollen development in rice.

Centromeres consist of highly repetitive sequences that are challenging to map, clone, and sequence. Active genes exist in centromeric regions, but their biological functions are difficult to explore owing to extreme suppression of recombination in these regions. In this study, we used the CRISPR/Cas9 system to knock out the transcribed gene Mitochondrial Ribosomal Protein L15 (OsMRPL15), located in the centromeric region of rice (Oryza sativa) chromosome 8, resulting in gametophyte sterility. Osmrpl15 pollen was completely sterile, with abnormalities appearing at the tricellular stage including the absence of starch granules and disrupted mitochondrial structure. Loss of OsMRPL15 caused abnormal accumulation of mitoribosomal proteins and large subunit rRNA in pollen mitochondria. Moreover, the biosynthesis of several proteins in mitochondria was defective, and expression of mitochondrial genes was upregulated at the mRNA level. Osmrpl15 pollen contained smaller amounts of intermediates related to starch metabolism than wild-type pollen, while biosynthesis of several amino acids was upregulated, possibly to compensate for defective mitochondrial protein biosynthesis and initiate consumption of carbohydrates necessary for starch biosynthesis. These results provide further insight into how defects in mitoribosome development cause gametophyte male sterility.

The DC1 Domain Protein BINUCLEATE POLLEN is Required for POLLEN Development in Arabidopsis thaliana.

The development of the male gametophyte is a tightly regulated process that requires the precise control of cell division and gene expression. A relevant aspect to understand the events underlying pollen development regulation constitutes the identification and characterization of the genes required for this process. In this work, we showed that the DC1 domain protein BINUCLEATE POLLEN (BNP) is essential for pollen development and germination. Pollen grains carrying a defective BNP alleles failed to complete mitosis II and exhibited impaired pollen germination. By yeast two-hybrid analysis and bimolecular fluorescence complementation assays, we identified a set of BNP-interacting proteins. Among confirmed interactors, we found the NAC family transcriptional regulators Vascular Plant One-Zinc Finger 1 (VOZ1) and VOZ2. VOZ1 localization changes during pollen development, moving to the vegetative nucleus at the tricellular stage. We observed that this relocalization requires BNP; in the absence of BNP in pollen from bnp/BNP plants, VOZ1 nuclear localization is impaired. As the voz1voz2 double mutants showed the same developmental defect observed in bnp pollen grains, we propose that BNP requirement to complete microgametogenesis could be linked to its interaction with VOZ1/2 proteins. BNP could have the role of a scaffold protein, recruiting VOZ1/2 to the endosomal system into assemblies that are required for their further translocation to the nucleus, where they act as transcriptional regulators.

Hexose transporter SWEET5 confers galactose sensitivity to Arabidopsis pollen germination via a galactokinase.

Galactose is an abundant and essential sugar used for the biosynthesis of many macromolecules in different organisms, including plants. Galactose metabolism is tightly and finely controlled, since excess galactose and its derivatives are inhibitory to plant growth. In Arabidopsis (Arabidopsis thaliana), root growth and pollen germination are strongly inhibited by excess galactose. However, the mechanism of galactose-induced inhibition during pollen germination remains obscure. In this study, we characterized a plasma membrane-localized transporter, Arabidopsis Sugars Will Eventually be Exported Transporter 5, that transports glucose and galactose. SWEET5 protein levels started to accumulate at the tricellular stage of pollen development and peaked in mature pollen, before rapidly declining after pollen germinated. SWEET5 levels are responsible for the dosage-dependent sensitivity to galactose, and galactokinase is essential for these inhibitory effects during pollen germination. However, sugar measurement results indicate that galactose flux dynamics and sugar metabolism, rather than the steady-state galactose level, may explain phenotypic differences between sweet5 and Col-0 in galactose inhibition of pollen germination.

The Arabidopsis AGC kinases NDR2/4/5 interact with MOB1A/1B and play important roles in pollen development and germination.

Pollen formation and pollen tube growth are essential for the delivery of male gametes into the female embryo sac for double fertilization. Little is known about the mechanisms that regulate the late developmental process of pollen formation and pollen germination. In this study, we characterized a group of Arabidopsis AGC kinase proteins, NDR2/4/5, involved in pollen development and pollen germination. The NDR2/4/5 genes are mainly expressed in pollen grains at the late developmental stages and in pollen tubes. They function redundantly in pollen formation and pollen germination. At the tricellular stages, the ndr2 ndr4 ndr5 mutant pollen grains exhibit an abnormal accumulation of callose, precocious germination and burst in anthers, leading to a drastic reduction in fertilization and a reduced seed set. NDR2/4/5 proteins can interact with another group of proteins (MOB1A/1B) homologous to the MOB proteins from the Hippo signaling pathway in yeast and animals. The Arabidopsis mob1a mob1b mutant pollen grains also have a phenotype similar to that of ndr2 ndr4 ndr5 pollen grains. These results provide new evidence demonstrating that the Hippo signaling components are conserved in plants and play important roles in sexual plant reproduction.

Ultrastructural characterization of microlipophagy induced by the interaction of vacuoles and lipid bodies around generative and sperm cells in Arabidopsis pollen

During pollen maturation, various organelles change their distribution and function during development as male gametophytes. We analyzed the behavior of lipid bodies and vacuoles involved in lipophagy in Arabidopsis pollen using serial section SEM and conventional TEM. At the bicellular pollen stage, lipid bodies in the vegetative cells lined up at the surface of the generative cell. Vacuoles then tightly attached, drew in, and degraded the lipid bodies and eventually occupied the space of the lipid bodies. Degradation of lipid began before transfer of the entire contents of the lipid body. At the tricellular stage, vacuoles instead of lipid bodies surrounded the sperm cells. The degradation of lipid bodies is morphologically considered microautophagy. The atg2-1 Arabidopsis mutant is deficient in one autophagy-related gene (ATG). In this mutant, the assembly of vacuoles around sperm cells was sparser than that in wild-type pollen. The deficiency of ATG2 likely prevents or slows lipid degradation, although it does not prevent contact between organelles. These results demonstrate the involvement of microlipophagy in the pollen development of Arabidopsis.

The Canonical E2Fs Are Required for Germline Development in Arabidopsis.

A number of cell fate determinations, including cell division, cell differentiation, and programmed cell death, intensely occur during plant germline development. How these cell fate determinations are regulated remains largely unclear. The transcription factor E2F is a core cell cycle regulator. Here we show that the Arabidopsis canonical E2Fs, including E2Fa, E2Fb, and E2Fc, play a redundant role in plant germline development. The e2fa e2fb e2fc (e2fabc) triple mutant is sterile, although its vegetative development appears normal. On the one hand, the e2fabc microspores undergo cell death during pollen mitosis. Microspores start to die at the bicellular stage. By the tricellular stage, the majority of the e2fabc microspores are degenerated. On the other hand, a wild type ovule often has one megaspore mother cell (MMC), whereas the majority of e2fabc ovules have two to three MMCs. The subsequent female gametogenesis of e2fabc mutant is aborted and the vacuole is severely impaired in the embryo sac. Analysis of transmission efficiency showed that the canonical E2Fs from both male and female gametophyte are essential for plant gametogenesis. Our study reveals that the canonical E2Fs are required for plant germline development, especially the pollen mitosis and the archesporial cell (AC)-MMC transition.

The Rice Eukaryotic Translation Initiation Factor 3 Subunit f (OseIF3f) Is Involved in Microgametogenesis.

Microgametogenesis is the post-meiotic pollen developmental phase when unicellular microspores develop into mature tricellular pollen. In rice, microgametogenesis can influence grain yields to a great degree because pollen abortion occurs more easily during microgametogenesis than during other stages of pollen development. However, our knowledge of the genes involved in microgametogenesis in rice remains limited. Due to the dependence of pollen development on the regulatory mechanisms of protein expression, we identified the encoding gene of the eukaryotic translation initiation factor 3, subunit f in Oryza sativa (OseIF3f). Immunoprecipitation combined with mass spectrometry confirmed that OseIF3f was a subunit of rice eIF3, which consisted of at least 12 subunits including eIF3a, eIF3b, eIF3c, eIF3d, eIF3e, eIF3f, eIF3g, eIF3h, eIF3i, eIF3k, eIF3l, and eIF3m. OseIF3f showed high mRNA levels in immature florets and is highly abundant in developing anthers. Subcellular localization analysis showed that OseIF3f was localized to the cytosol and the endoplasmic reticulum in rice root cells. We further analyzed the biological function of OseIF3f using the double-stranded RNA-mediated interference (RNAi) approach. The OseIF3f-RNAi lines grew normally at the vegetative stage but displayed a large reduction in seed production and pollen viability, which is associated with the down-regulation of OseIF3f. Further cytological observations of pollen development revealed that the OseIF3f-RNAi lines showed no obvious abnormalities at the male meiotic stage and the unicellular microspore stage. However, compared to the wild-type, OseIF3f-RNAi lines contained a higher percentage of arrested unicellular pollen at the bicellular stage and a higher percentage of arrested unicellular and bicellular pollen, and aborted pollen at the tricellular stage. These results indicate that OseIF3f plays a role in microgametogenesis.

Evaluation of rice promoters conferring pollen-specific expression in a heterologous system, Arabidopsis

Promoters can direct gene expression specifically to targeted tissues or cells. Effective with both crop species and model plant systems, these tools can help researchers overcome the practical obstacles associated with transgenic protocols. Here, we identified promoters that allow one to target the manipulation of gene expression during pollen development. Utilizing published transcriptomic databases for rice, we investigated the promoter activity of selected genes in Arabidopsis. From various microarray datasets, including those for anthers and pollen grains at different developmental stages, we selected nine candidate genes that showed high levels of expression in the late stages of rice pollen development. We named these Oryza sativa late pollen-specific genes. Their promoter regions contained various cis-acting elements that could be responsible for anther-/pollen-specific expression. Promoter::GUS-GFP reporters were constructed and introduced into Arabidopsis plants. Histochemical GUS staining revealed that six of the nine rice promoters conferred strong GUS expression that was restricted to the anthers in Arabidopsis. Further analysis showed that although the GUS signals were not detected at the unicellular stage, they strengthened in the bicellular or tricellular stages, peaking at the mature pollen stage. This paralleled their transcriptomic profiles in rice. Based on our results, we proposed that these six rice promoters, which are active in the late stages of pollen formation in the dicot Arabidopsis, can aid molecular breeders in generating new varieties of a monocot plant, rice.

Two linked genes on rice chromosome 2 for F1 pollen sterility in a hybrid between Oryza sativa and O. glumaepatula.

Hybrid incompatibility plays an important role in establishment of post-zygotic reproductive isolation. To unveil genetic basis of hybrid incompatibilities between diverged species of genus Oryza AA genome species, we conducted genetic dissection of hybrid sterility loci, S22(t), which had been identified in backcross progeny derived from Oryza sativa ssp. japonica (recurrent parent) and South American wild rice O. glumaepatula near the end of the short arm of chromosome 2. The S22(t) region was found to be composed of two loci, designated S22A and S22B, that independently induce F1 pollen sterility. Pollen grains containing either of the sterile alleles (S22A-glums or S22B-glums) were sterile if produced on a heterozygous plant. No transmission of the S22A-glums allele via pollen was observed, whereas a low frequency of transmission of S22B-glums was observed. Cytological analysis showed that the sterile pollen grains caused by S22A could reach the bicellular or tricellular stage, and the nearly-sterile pollen grains caused by S22B could reach the tricellular stage. Our genetic analysis showed repulsion linkage effect is possible to induce strong reproductive barrier by high pollen sterility based on recombination value and transmission ratio of hybrid sterility gene to the progeny was influenced by frequency of competitors on fertilization.

Computational identification of Brassica napus pollen specific protein Bnm1 as an allergen

Bnm1 is a pollen specific protein from Brassica napus(oilseed rape) and it is specifically expressed in the bi-cellular and tri-cellular stages of pollen development. Since the incidence of pollinosis due to oilseed rape (Brassica napus) is increasing day by day, keeping pace with its high cultivation rate, the search for its allergens is a demand of time to develop effective immune therapy. In the present study, different computational tools were adopted to predict the po tential of Bnm1 as a candidate allergen. Physico chemical properties of Bnm1 showed its molecular weight (~20kD) and theoretical pI (5.27) along with other properties to be fallen between the ranges essential for a protein to be an allergen. Keeping in min d the capability of allergen to induce both humoral and cell mediated immune response, we checked both the potential B cell and T cell epitope candidates of Bnm1 using different immune -informatics tools housed at IEDB analysis resource. For B cell epitopeprediction, potential antigenic sites on the protein surface were predicted by both propensity scale and machine learning method followed by their mapping on Bnm1 3D structure predicted from homology modeling. In case of T cell epitope prediction, interact ion of the core sequence with seven abundant MHC -II alleles (DRB1*0101, DRB1*0301, DRB1*0401, DRB1*0701, DRB1*1101, DRB1*1301 and DRB1*1501) within an IC50 range (IC50<25) was the basis. We observed multiple epitope interactions with DRB1*0101 allele, impl ying that this interactions might evoke a strong TH2 response ensuring an increased production of IgE response. Based on our prediction we hereby claim that the Bnm1 is a potential allergen having capability to induce both humoral and cell mediated allergi c responses. However, in vitro analysis was out of our scope and therefore it should have to be performed to validate the potency of Bnm1 as an allergen.

Screening and Analysis of Proteins Interacting with TaPDK from Physiological Male Sterility Induced by CHA in Wheat

To further research the regulatory network of pyruvate dehydrogenase kinase (designated as TaPDK) in physiological male-sterility (PHYMS) of wheat induced by chemical hybridizing agent (CHA) SQ-1, an anther cDNA library was constructed, and the proteins interacting with TaPDK were screened via yeast two-hybrid technique. Subsequently, a few candidate proteins in nucleotide expression levels were detected by real-time quantitative PCR. Yeast-two hybrid screening was performed by mating yeast strain Y2HGold containing BD-TaPDK bait plasmid with yeast strain Y187 including anther cDNA library plasmid. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Ade/-His/-Leu/-Trp) (QDO), and further were incubated on QDO medium containing AbA and X-α-Gal. The interactions between TaPDK and the proteins obtained from positive colonies were further confirmed by co-transformation validation. After plasmids DNA were extracted from blue colonies and sequenced, the sequences results were analyzed by bioinformatic methods. Finally, 24 colonies were obtained, including eight genes, namely non-specific lipid-transfer protein precursor (TanLTP), polyubiquitin (TaPUbi), glyceraldehyde-3-phosphate dehydrogenase, proliferating cell nuclear antigen (TaPCNA), CBS domain containing protein (TaCBS), actin, guanine nucleotide-binding protein beta subunit, chalcone synthase, and three new genes with unknown function. The results of quantitative RT-PCR showed that the expression levels of TanLTP, TaPUbi, and TaPCNA were obviously up-regulated in PHYMS anther, and TaCBS expression was only increased at the tricellular stage in PHYMS anther compared with in fertile lines. Whereas, the expression of TaPDK was obviously down-regulated in PHYMS lines. Collectively, these datas indicated that the majority of candidate proteins might be related to pollen abortion in PHYMS lines, which further suggested that TaPDK plays multiple roles in pollen development, besides participating in regulating pyruvate dehydrogenase complex activity.

Arabidopsis FAB1/PIKfyve Proteins Are Essential for Development of Viable Pollen

Phosphatidylinositol 3,5-bisphosphate [PtdIns(3,5)P(2)] is a phospholipid that has a role in controlling membrane trafficking events in yeast and animal cells. The function of this lipid in plants is unknown, although its synthesis has been shown to be up-regulated upon osmotic stress in plant cells. PtdIns(3,5)P(2) is synthesized by the PIKfyve/Fab1 family of proteins, with two orthologs, FAB1A and FAB1B, being present in Arabidopsis (Arabidopsis thaliana). In this study, we attempt to address the role of this lipid by analyzing the phenotypes of plants mutated in FAB1A and FAB1B. It was not possible to generate plants homozygous for mutations in both genes, although single mutants were isolated. Both homozygous single mutant plant lines exhibited a leaf curl phenotype that was more marked in FAB1B mutants. Genetic transmission analysis revealed that failure to generate double mutant lines was entirely due to inviability of pollen carrying mutant alleles of both FAB1A and FAB1B. This pollen displayed severe defects in vacuolar reorganization following the first mitotic division of development. The presence of abnormally large vacuoles in pollen at the tricellular stage resulted in the collapse of the majority of grains carrying both mutant alleles. This demonstrates a crucial role for PtdIns(3,5)P(2) in modulating the dynamics of vacuolar rearrangement essential for successful pollen development. Taken together, our results are consistent with PtdIns(3,5)P(2) production being central to cellular responses to changes in osmotic conditions.

Embryology ofHelianthus annuus(Asteraceae)

Abortion of sunflower fruits during different phases of their development could be due to abnormalities in the reproductive process. We report a comparative study of the sporogenesis, gametogenesis and the development of the related sporophytic structures in three commercial hybrids, DK 4050, CF 17 and P30, and a line, HA 89. The anther wall consists of epidermis, endothecium, one middle layer and a plasmodial tapetum with binucleate cells before integrating. A peritapetal membrane with orbicules encloses the pollen grains, which are triporate, angulaperturate and shed at tri cellular stage. HA 89 also presents tetraporate pollen grains. The ovule is tenuinucellate, unitegmic and anatropous. The young female gametophyte consists of six cells and eight nuclei. Antipodals, which vary in number and in the number of nuclei in each cell, usually have thick walls between themselves and the central cell. Differences among the genotypes studied may explain the contradictions found on previous accounts.

Characterization and expression of Arabidopsis UDP-sugar pyrophosphorylase

At5g52560, a homolog of pea ( Pisum sativum) UDP-sugar pyrophosphorylase ( PsUSP) was functionally annotated by expression in Escherichia coli and subsequent characterization of substrate specificity and kinetic properties. Arabidopsis contains a single USP gene ( AtUSP) and evaluation of gene databases suggests that USP is unique to plants. The 69 kDa AtUSP gene product exhibited high activity with Glc-1-P, GlcA-1-P and Gal-1-P, but low activity with GlcNAc-1-P, Fuc-1-P, Man-1-P, inositol-1-P or Glc-6-P. AtUSP was activated by magnesium and preferred UTP as co-substrate. Apparent K m values for GlcA-1-P, Glc-1-P and UTP were 0.13 mM, 0.42 mM and 0.14 mM, respectively. In the reverse direction (pyrophosphorolysis), the apparent K m values for UDP-GlcA, UDP-Glc and pyrophosphate were 0.56 mM, 0.72 mM and 0.15 mM, respectively. USP enzyme activity (UDP-GlcA → GlcA-1-P) was detected in Arabidopsis tissues with highest activity found in the inflorescence. As determined by semi-quantitative RT-PCR, AtUSP transcript is widely expressed with high levels detected in the inflorescence. To evaluate tissue-specific expression of AtUSP, histochemical GUS staining of plants transformed with AtUSPprom:GUS constructs was performed. In 7-day-old seedlings, GUS staining was detected in cotyledons, trichomes and vascular tissues of the primary root. In the inflorescence of older plants, high levels of GUS staining were detected in cauline leaves, the epidermis of the stem and in pollen. In silico analysis of AtUSP expression in developing pollen indicates that transcript levels increase as development proceeds from the uninucleate to the tricellular stage. The results suggest that AtUSP plays an important role in pollen development in Arabidopsis.

Localisation of allergens in ryegrass pollen and in airborne micronic particles

In Melbourne, Australia, grass pollen allergens, especially from ryegrass, are a major cause of allergic hayfever and asthma. This review outlines recent developments in our understanding of how grass pollen allergens find their way into the atmosphere and how they are transported in particulate form. Much of this work has relied on antibody technology in immunological and immunocytochemical investigations. The localisation of allergens in situ has proved difficult due to their water-soluble character. Recently, allergens have been localised in developing ryegrass pollen by dryfixation, rapid-freeze and freeze-substitution techniques. This involved anthers being substituted in a mixture of aldehydes, organic solvents, and 2,2-dimethoxypropane. Incubation in dimethylsulfoxide prior to embedding in LR Gold resin provided good infiltration with freeze-substituted material. Immunogold-labelled sections show that the major allergens, Lol p 1 and Lol p 5, are synthesised in the pollen cytoplasm from the early bicellular stage, soon after the first starch granules are formed. From the early tricellular stage, Lol p 5 moves into the starch granules where it remains until maturity. Lol p 1 is localised in the cytoplasm of mature pollen grains. The incidence of airborne grass pollen, as measured in pollen traps, correlates with hayfever symptoms. Forecasting models which rely on rainfall and temperature data have been produced for the grass pollen (daily and seasonal) counts in Melbourne. Research over the past six years has shed light on the causes of grass-pollen-induced asthma. Micronic particles in the atmosphere may be starch granules originating from pollen grains osmotically ruptured by rainwater. Ultrastructural and immunological characterisation of micronic particles collected from outdoor air filters confirm the presence of airborne starch granules. These are loaded with grass pollen allergens, occur in the atmosphere especially after rainfall, and correlate significantly with instances of allergic asthma. Diesel particles might also play a role in the transmission of grass pollen allergens and thus become an extra asthma trigger. A variation in the mode of release of micronic particles occurs in other species, such as birch, where such particles are derived from burst birch pollen tubes. These particles are positive for Bet v 1 and are starch granules which are released into the atmosphere after light rain as a result of pollen germination on, e.g., leaves. After subsequent rupture of pollen tubes their contents are released when conditions become drier.

Comparative studies on the male and female gametophyte development in three different Triticum species

The in planta and in vitro development of the male and female gametophytes of three Triticum species (T. aestivum cv. Mv 15/6n/, T. araraticum/4n/ and T. monococcum/2n/) was traced from just after meiosis until the development of the mature pollen or embryo sac. The dynamics of differentiation in the male gametophytes was not influenced by the different ploidy levels up to the tricellular stage. The maturing process of differentiated haploid male gametophytes, resulting in the shedding of mature pollen from the anthers, proceeds most rapidly in the hexaploid species. The development of the initial microspore slowed in vitro, remaining in the uninucleate stage for a longer period, with a delay in first pollen mitosis compared to the control. There was no substantial difference between the three species in the dynamics of in vitro pollen development. In all cases the uninucleate microspore developed into a functional tricellular pollen grain within a week. However, there were differences in the lengths of the various developmental stages under in vitro and natural conditions. The seven-cellular female gametophyte developed far more rapidly than the male gametophyte, which means that the mitotic divisions leading to the differentiation of male and female gametophytes were not synchronised within the plants. In species with lower ploidy levels macrosporogenesis took longer than microsporogenesis, and the development of the seven-celled embryo sac was completed 2 days later than in hexaploid species. The differing ploidy levels also influenced the size of the embryo sac. The embryo sacs achieved their maximum dimensions 2–3 days prior to male gametophyte maturation. These observations raise several questions as to when gametophytes can be regarded as functionally mature, and which phase of gametophytic development is most suitable for cell manipulation.

Genetic control of the rate of transmission of rye B chromosomes. III. Male meiosis and gametogenesis

Male meiosis and gametogenesis were studied at metaphase I, metaphase and anaphase of the first pollen grain mitosis, and bicellular and tricellular pollen grain stages in 2B rye plants belonging to the low (L) and high (H) B transmission rate lines previously selected. Our results show that B chromosome behaviour significantly differs in both lines whereas the behaviour of the normal complement does not differ. In the L line the Bs form univalents in 81.07 per cent of the metaphase I cells, and are conserved in 44.14 per cent of the pollen grains at first metaphase whereas the remaining Bs are eliminated as micronuclei. In the H line the Bs form bivalents in 87.71 per cent of the metaphase I cells, and are present in 82.48 per cent of pollen grains at first metaphase. The Bs of the L and H lines do not differ in their ability to undergo nondisjunction at first pollen grain anaphase. This indicates that the different B transmission in the L and H lines results from their differential ability to form uni- or bivalents at metaphase I, which determines their loss or conservation in the pollen grains. The L and H lines also differ in pollen viability at the tricellular stage because 19.75 per cent of pollen grains of the L line and only 1.2 per cent of the H line are inviable.

Genetic control of the rate of transmission of rye B chromosomes. III. Male meiosis and gametogenesis

Male meiosis and gametogenesis were studied at metaphase I, metaphase and anaphase of the first pollen grain mitosis, and bicellular and tricellular pollen grain stages in 2B rye plants belonging to the low (L) and high (H) B transmission rate lines previously selected. Our results show that B chromosome behaviour significantly differs in both lines whereas the behaviour of the normal complement does not differ. In the L line the Bs form univalents in 81.07 per cent of the metaphase I cells, and are conserved in 44.14 per cent of the pollen grains at first metaphase whereas the remaining Bs are eliminated as micronuclei. In the H line the Bs form bivalents in 87.71 per cent of the metaphase I cells, and are present in 82.48 per cent of pollen grains at first metaphase. The Bs of the L and H lines do not differ in their ability to undergo nondisjunction at first pollen grain anaphase. This indicates that the different B transmission in the L and H lines results from their differential ability to form uni- or bivalents at metaphase I, which determines their loss or conservation in the pollen grains. The L and H lines also differ in pollen viability at the tricellular stage because 19.75 per cent of pollen grains of the L line and only 1.2 per cent of the H line are inviable.

The Critical Requirement for Linolenic Acid Is Pollen Development, Not Photosynthesis, in an Arabidopsis Mutant.

The very high proportions of trienoic fatty acids found in chloroplast membranes of all higher plants suggest that these lipid structures might be essential for photosynthesis. We report here on the production of Arabidopsis triple mutants that contain negligible levels of trienoic fatty acids. Photosynthesis at 22[deg]C was barely affected, and vegetative growth of the mutants was identical with that of the wild type, demonstrating that any requirement for trienoic acyl groups in membrane structure and function is relatively subtle. Although vegetative growth and development were unaffected, the triple mutants are male sterlle and produce no seed under normal conditions. Comparisons of pollen development in wild-type and triple mutant flowers established that pollen grains in the mutant developed to the tricellular stage. Exogenous applications of [alpha]-llnolenate or jasmonate restored fertility. Taken together, the results demonstrate that the critical role of trienoic acids in the life cycle of plants is as the precursor of oxylipin, a signaling compound that regulates final maturation processes and the release of pollen.