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PubMed2004

Immunolocalisation of arabinogalactan proteins and pectins in Actinidia deliciosa pollen. Short communication.

The cell wall composition of germinating pollen grains of Actinidia deliciosa was studied by immunolocalization with monoclonal antibodies against arabinogalactan proteins (AGPs) and pectins. In ungerminated pollen, the JIM8 epitope (against a subset of AGPs) was located in the intine and in the cytoplasm, while the MAC207 epitope (against AGPs) was located only in the exine. After germination, the JIM8 and MAC 207 epitopes were located in the cytoplasm and in the pollen tube wall. The Yariv reagent that binds to AGPs was added to the germination medium inducing a reduction or inhibition in pollen germination. This indicates that AGPs are present in the growing pollen tube and play an important role in pollen germination. To identify the nature of the pectins found in pollen grains and tubes, four monoclonal antibodies were used. The JIM5 epitope (against unesterifled pectins) was located in the intine, more intensely in the pore region, and along the pollen tube wall, and the JIM7 epitope (against methylesterified pectins) was also observed in the cytoplasm. After germination, the JIM5 epitope was located in the pollen tube wall; although, the tube tip was not labelled. The JIM7 epitope was located in the entire pollen tube wall. LM5 (against galactans) showed a labelling pattern similar to that of JIM5 and the pattern of LM6 (against arabinans) was similar to that of JIM7. Pectins show different distribution patterns when the degree of esterification is considered. Pollen tube wall pectins are less esterified than those of the pollen tube tip. The association of AGPs with pectins in the cell wall of the pollen grain and the pollen tube may play an important role in the maintenance of cell shape during pollen growth and development.

Actinidia↗
PubMed1958

Intracellular amounts of nucleic acids and protein during pollen grain growth in Tradescantia.

The rapid growth, large organelles, and synchronous development of T. paludosa pollen grains make them ideal subjects for cytochemical analysis. A microphotometric study of the nucleoli, chromosomes, and cytoplasm fixed at daily intervals during pollen grain maturation indicated that: 1. DNA (Feulgen) synthesis in the generative nucleus occurred during the first third of interphase, while the DNA content of the vegetative nucleus remained unchanged. 2. Throughout development, changes in RNA (azure B) content, in general, paralleled changes in protein (NYS(1), Millon) content in each organelle of the vegetative cell. Initially, the RNA and protein of all organelles increased up to mid interphase, when chromosomal and nucleolar fractions began to decline despite a continued increase in cytoplasmic RNA and protein. At least 24 hours before anthesis, the vegetative nucleolus had disappeared and chromosomal protein and RNA of the vegetative nucleus were apparently in rapid decline. Such a system offered an opportunity to study the role of the nucleus, especially the nucleolus, in RNA and protein metabolism in the cytoplasm, by noting what cytoplasmic processes could and could not continue at a time when nuclear mechanisms were absent or minimal. It was found that at least 2 fundamental processes continued during this period: both RNA and protein accumulated in the cytoplasm at a rapid rate. It was concluded that the nucleus is not the sole source of cytoplasmic RNA, for the data suggest that there are at least 2 separate and independent, or remotely dependent synthesizing systems, one nuclear and the other cytoplasmic. It is evident that nuclear influence on cytoplasmic synthesis need be neither direct nor immediate.

Cell Nucleolus↗
PubMed2006

Xenia effect on seed and embryo size in cotton (Gossypium hirsutum L.).

The term xenia was coined to describe the effect of foreign pollen on the development and characters of the seed. To study its importance and consequences for various seed traits in cotton (Gossypium hirsutum L.), the effect of pollen genotype on seed and embryo weight was studied with seeds from 15 F1 hybrids. Cross-fertilization changed seed weight by up to 7.0% in relation to self-fertilization. Xenia effect significantly increased embryo weight of cross-fertilized seeds, by up to 14.4% in comparison to self-fertilized seeds. Seeds of some crosses had a lower hull content than corresponding selfed seeds. On average, the xenia effect was greater for embryo weight than for seed weight. However, in some crosses there was no difference between cross- and self-fertilized seeds for seed weight, embryo weight, moisture content and hull content. Positive xenia effects for seed weight and embryo weight may help us to establish uniform stands of vigorous hybrid seedlings, especially under unfavourable conditions. Also, larger seed and embryo weight, along with lower hull content, could result in higher oil yield. Therefore, careful choosing of genotypes as parents and of cross direction in the production of hybrid seed is very important in cotton.

Chimera↗
PubMed2001

Interaction between maternal effect and zygotic effect mutations during maize seed development.

Double fertilization of the embryo sac by the two sperm cells of a pollen grain initiates seed development. Proper development of the seed depends not only on the action of genes from the resulting embryo and endosperm, but also on maternal genes acting at two stages. Mutations with both sporophytic maternal effects and gametophytic maternal effects have been identified. A new maternal effect mutation in maize, maternal effect lethal1 (mel1), causes the production of defective seed from mutant female gametophytes. It shows reduced pollen transmission, suggesting a requirement in the male gametophyte, but has no paternal effect on seed development. Interestingly, the defective kernel phenotype of mel1 is conditioned only in seeds that inherit mel1 maternally and are homozygous for the recessive allele (endogenous to the W22 inbred line) of either of two genes, sporophyte enhancer of mel1 (snm1) or snm2, suggesting redundancy between maternally and zygotically required genes. Both mel1 and snm1 map to the short arm of chromosome 2, and snm2 maps to the long arm of chromosome 10. The mode of action of mel1 and the relationship between mel1 and snm1 and snm2 are discussed.

Alleles↗
PubMed2002

Successive glycosyltransfer activity and enzymatic characterization of pectic polygalacturonate 4-alpha-galacturonosyltransferase solubilized from pollen tubes of Petunia axillaris using pyridylaminated oligogalacturonates as substrates.

Polygalacturonate 4-alpha-galacturonosyltransferase (pectin synthase) was solubilized from pollen tubes of Petunia axillaris and characterized. To accomplish this, an assay method using fluorogenic pyridylaminated-oligogalacturonic acids (PA-OGAs) as acceptor substrates was developed. When the pollen tube enzyme was solubilized with 0.5% (v/v) Triton X-100 and was incubated with PA-OGA and UDP-galacturonic acid (UDP-GalUA), successive transfer activity of more than 10 GalUAs from UDP-GalUA to the nonreducing end of PA-OGA was observed by diethylaminoethyl high-performance liquid chromatography. This activity was time- and enzyme concentration-dependent. The optimum enzyme activity was observed at pH 7.0 and 30 degrees C. Among the PA-OGAs investigated, those with a degree of polymerization of more than 10 were preferred as substrates. The crude pollen tube enzyme had an apparent K(m) value of 13 microM for the PA-OGA with a degree of polymerization 11 and 170 microM for UDP-GalUA. The characteristics of the P. axillaris pollen tube enzyme and the usefulness of fluorogenic PA-OGAs for the assay of this enzyme are discussed.

Cations↗
PubMed2007

The impact of pollen-related food allergens on pollen allergy.

Patients with birch pollen allergy frequently develop hypersensitivity reactions to certain foods, e.g. apples, celery, carrots and hazelnuts. These reactions are mainly caused by IgE-antibodies specific for the major birch pollen allergen, Bet v 1, which cross-react with homologous proteins in these foods. Analyzing the T-cell response to Bet v 1-related food allergens revealed that these dietary proteins contain several distinct T-cell epitopes and activate Bet v 1-specific T cells to proliferate and produce cytokines. Several of these cross-reactive T-cell epitopes were not destroyed by simulated gastrointestinal digestion of food allergens and stimulated Bet v 1-specific T cells despite nonreactivity with IgE antibodies. Similarly, cooked food allergens did not elicit IgE-mediated symptoms (oral allergy syndromes) but caused T-cell-mediated late-phase reactions (deterioration of atopic eczema) in birch pollen-allergic patients with atopic dermatitis because thermal processing affected their conformational structure and not the primary amino acid sequence. Thus, T-cell cross-reactivity between Bet v 1 and related food allergens occurs independently of IgE-cross-reactivity in vitro and in vivo. We speculate that symptom-free consumption of pollen-related food allergens may have implications for the pollen-specific immune response of allergic individuals.

Allergens↗
PubMed1987

Phytic Acid Metabolism in Lily (Lilium longiflorum Thunb.) Pollen.

The accumulation of phytic acid during development of lily (Lilium longiflorum Thunb.) pollen and its degradation during germination have been studied. A substantial amount of phytic acid accumulates in lily pollen by 5 days before anthesis, and little change occurs during subsequent maturation. Mature lily pollen contains 7 to 8 micrograms phytic acid per milligram pollen. Considerable degradation of phytic acid occurs by 15 minutes of incubation in glucose culture medium, and very little is left by 3 hours. No partially phosphorylated myo-inositol accumulates during germination. The breakdown of phytic acid proceeds at a constant rate during this time period. The rate is calculated to be 0.037 microgram phytic acid/milligram pollen/minute. Two phytases are detected in germinated lily pollen extract using high performance liquid chromatography with an anion exchange column (diethylaminoethyl-5PW). The results suggest that one of the phytases is already present in mature ungerminated lily pollen and the other one is newly synthesized during germination from a long-lived, pre-existing mRNA.

Journal Article↗
PubMed2000

Unusual core histones specifically expressed in male gametic cells of Lilium longiflorum.

We have cloned three novel histone genes using antibodies that recognize only nuclei of the male gametic (generative and sperm) cells of Lilium longiflorum. The deduced amino acid sequence of each clone shows only between 40% and 50% identity with the H2A, H2B and H3 somatic core histones of other plant species. Transcripts of these genes were first detected in bicellular pollen soon after microspore mitosis, and their mRNAs, as revealed by in situ hybridization, were observed only in the cytoplasm of the generative cells. As expression of these three genes was specific to generative cells within the bicellular pollen, we designated the clones gH2A, gH2B and gH3. Immunocytochemistry further revealed that the proteins encoded by these genes accumulated in the elongating and condensing generative nucleus during development of bicellular pollen, and were most abundant in the two sperm nuclei within an elongated pollen tube. We therefore propose that these male gamete-specific core histones contribute to chromatin condensation of male gametes or to chromatin remodeling, and result in the repression of gene expression in male gametes.

Amino Acid Sequence↗
PubMed1994

Gibberellin-induced changes in the translatable mRNA populations of stamens and shoots of gibberellin-deficient tomato.

The gib1 mutant of tomato (Lycopersicon esculentum Mill.) is deficient in endogenous gibberellins and exhibits phenotypes including extreme dwarfism, reduced germination, and abnormal flower development, which are reversed by the application of gibberellic acid (GA3). Previous work has demonstrated that, in stamens of the gib1 mutant, pollen mother-cell development arrests at the premeiotic G1 stage (Jacobsen and Olszewski 1991, Plant Physiol. 97, 409-414). Following GA3 treatment of developmentally arrested flowers, pollen mother-cell development resumes and is synchronous. The present study examines gibberellin-induced changes in the translatable mRNA populations of developmentally arrested stamens and of vegetative shoots of the gib1 mutant. Following rescue of developmentally arrested stamens by treatment with GA3, we consistently detected increases and decreases in the abundance of 14 and 20 in-vitro translation products, respectively. Some of these changes were first detected 8 h post treatment and therefore represent the first changes observed in stamens whose development has been rescued by GA3 treatment. In vegetative gib1 shoots, the abundance of 13 in-vitro translation products decreased within 6-24 h after GA3 treatment. However, no in-vitro translation products that increased in abundance after GA3 treatment were detected.

Gene Expression↗
PubMed2003

Expression pattern of diacylglycerol acyltransferase-1, an enzyme involved in triacylglycerol biosynthesis, in Arabidopsis thaliana.

Triacylglycerol (TAG) is the major carbon storage reserve in oilseeds such as Arabidopsis. Acyl-CoA:diacylglycerol acyltransferase (DGAT) catalyses the final step of the TAG synthesis pathway. Although TAG is mainly accumulated during seed development, and DGAT has presumably the highest activity in developing seeds, we show here that TAG synthesis is also actively taking place during germination and seedling development in Arabidopsis. The expression pattern of the DGAT1 gene was studied in transgenic plants containing the reporter gene beta-glucuronidase (GUS) fused with DNA sequences flanking the DGAT1 coding region. GUS activity was not only detected in developing seeds and pollen, which normally accumulate storage TAG, but also in germinating seeds and seedlings. Western blots showed that DGAT1 protein is present in several tissues, though is most abundant in developing seeds. In seedlings, DGAT1 is expressed in shoot and root apical regions, correlating with rapid cell division and growth. The expression of GUS in seedlings was consistent with the results of RNA gel blot analyses, precursor feeding and DGAT assay. In addition, DGAT1 gene expression is up-regulated by glucose and associated with glucose-induced changes in seedling development.

Acyltransferases↗
PubMed2002

Ectopic expression of BABY BOOM triggers a conversion from vegetative to embryonic growth.

The molecular mechanisms underlying the initiation and maintenance of the embryonic pathway in plants are largely unknown. To obtain more insight into these processes, we used subtractive hybridization to identify genes that are upregulated during the in vitro induction of embryo development from immature pollen grains of Brassica napus (microspore embryogenesis). One of the genes identified, BABY BOOM (BBM), shows similarity to the AP2/ERF family of transcription factors and is expressed preferentially in developing embryos and seeds. Ectopic expression of BBM in Arabidopsis and Brassica led to the spontaneous formation of somatic embryos and cotyledon-like structures on seedlings. Ectopic BBM expression induced additional pleiotropic phenotypes, including neoplastic growth, hormone-free regeneration of explants, and alterations in leaf and flower morphology. The expression pattern of BBM in developing seeds combined with the BBM overexpression phenotype suggests a role for this gene in promoting cell proliferation and morphogenesis during embryogenesis.

Amino Acid Sequence↗
PubMed1999

A male gametophyte-specific monosaccharide transporter in Arabidopsis.

The AtSTP2 gene (sugar transport protein 2) of Arabidopsis thaliana encodes a high affinity, low specificity monosaccharide carrier that can transport a number of hexoses and pentoses at similar rates. AtSTP2 has 12 putative transmembrane helices and a molecular mass of 55.0 kDa. AtSTP2 expression was localized in AtSTP2 promoter-beta-glucuronidase (GUS) Arabidopsis plants showing AtSTP2-driven GUS activity during pollen maturation and also in germinating pollen. Immunohistochemical studies with anti-AtSTP2 antiserum as well as RNA in situ hybridization analyses modified these results and showed that AtSTP2 expression is confined to the early stages of gametophyte development. Both AtSTP2 mRNA and AtSTP2 protein are first seen at the time of beginning callose degradation and microspore release from the tetrades. AtSTP2 mRNA and AtSTP2 protein are no longer detected after the mitotic divisions and the formation of the trinucleate gametophyte. No AtSTP2 mRNA or AtSTP2 protein is seen in fully developed or germinating pollen. The putative role of AtSTP2 in the uptake of glucose units resulting from callose degradation during pollen maturation is discussed.

Amino Acid Sequence↗
PubMed2006

[Natural course from rhinitis to asthma in the patients with autumnal pollinosis: a clinical study of 1096 patients].

OBJECTIVE: To investigate the natural course from rhinitis to asthma in the patients with autumnal pollinosis. METHODS: 1096 patients with autumnal pollinosis, 488 males and 632 females, aged 38 +/- 14 (5 - 77), 511 with pure allergic rhinitis and 585 with allergic rhinitis complicated with asthma, underwent skin test of inhalant allergens, serum IgE specific to autumnal pollens, and questionnaire survey. RESULTS: The age range of rhinitis induced by autumnal pollens was 2 - 75. The peak onset age of rhinitis was 15 - 44, and the peak onset age of asthma was 25 - 54. 33% of the rhinitis patients complicated with asthma (194/585) had their first attack of rhinitis and the first attack of asthma in the same year, 66% of them (386/585) had the first attack of asthma later than the first attack of rhinitis, and only 0.8% of them (5/585) had their first attack of asthma earlier than that of rhinitis. 37% of the patients with autumnal pollen allergic rhinitis (410/1096) developed asthma within 5 years, 47% (511/1096), within 9 years, 5% (58/1096) within 10 approximately 19 years, and 1.5% (16/1096) within 20 - 40 years. CONCLUSION: Almost half of the patients with autumnal pollen allergic rhinitis develop to seasonal allergic asthma within 9 years.

Adolescent↗
PubMed2001

Monoclonal antibodies against the major allergen of Plantago lanceolata pollen, Pla l 1: affinity chromatography purification of the allergen and development of an ELISA method for Pla l 1 measurement.

BACKGROUND: Plantago lanceolata (English plantain) pollen is a relevant cause of pollinosis in temperate regions. The major allergen of this pollen, Pla l 1, is recognized by the specific IgE from more than 80% of plantain-sensitive patients. It displays significant sequence homology with the major olive-pollen allergen Ole e 1. The objective was to develop a monoclonal antibody-based ELISA to quantify Pla l 1, and to assess the correlation of Pla l 1 content with the biologic activity of plantain pollen extracts. We also aimed to establish the specificity of the monoclonal antibodies against the potentially cross-reactive allergen Ole e 1, and to investigate the presence of Pla l 1-like proteins in psyllium and melon that have been reported to cross-react with P. lanceolata pollen. METHODS: After fusion of myeloma cells with spleen cells from a BALB/c mouse, two Pla l 1-specific monoclonal antibodies secreting hybridomas were selected, and the antibodies characterized. One of them (2A10) was used as the capture antibody in an ELISA for Pla l 1 quantitation. An anti-P. lanceolata rabbit serum was used as the second antibody. Pla l 1 was purified by immunoaffinity chromatography and used as the standard in the assay. RESULTS: The ELISA developed was highly reproducible and sensitive, with a detection limit of 0.1 ng/ml, and a practical working range of 0.4-12 ng/ml. The specificity was demonstrated against a large battery of allergens, including Ole e 1. The concentration of Pla l 1 was measured in 19 extracts of P. lanceolata pollen, and a good correlation was observed between the Pla l 1 content and the allergenic activity of the extracts. Pla l 1 was not detected in psyllium or melon extracts. CONCLUSION: The results prove the usefulness of the Pla l 1-ELISA for the standardization of extracts of P. lanceolata pollen intended for clinical use.

Allergens↗
PubMed1982

Birch pollen allergy in children. Role of milk feeding during the first birch season of life.

The effect of exclusive breast-feeding throughout the first birch pollen season of life was examined in 59 children compared to 67 children on cow milk formula and to 27 children weaned to cow milk-based formula during their first birch season. The infants were about 3 months of age in their first birch season, and allergy to birch pollen was evaluated at 5 years of age by history and prick and provocation tests. Breast-feeding throughout the first birch season did not prevent birch pollen allergy; a similar birch allergy prevalence of about 10% was found in children initially fed on human or cow milk. Instead, weaning to cow milk-based formula during the first birch season seemed to protect from subsequent development of birch pollen allergy. Analogous results were obtained regarding allergy to grass pollen. This unexpected finding may be related to the immunologic stress on the young infant provided by introduction of cow milk proteins at an early age.

Animals↗
PubMed2006

Genetic transformation of barley (Hordeum vulgare L.) via infection of androgenetic pollen cultures with Agrobacterium tumefaciens.

A novel genetic transformation method for barley (Hordeum vulgare L.), based on infection of androgenetic pollen cultures with Agrobacterium tumefaciens, is presented. Winter-type barley cv. 'Igri' was amenable to stable integration of transgenes mediated by A. tumefaciens strain LBA4404 harbouring a vector system that confers hypervirulence, or by the non-hypervirulent strain GV3101 with a standard binary vector. The efficacy of gene transfer was substantially influenced by pollen pre-culture time, choice of Agrobacterium strain and vector system, Agrobacterium population density, medium pH and the concentrations of acetosyringone, CaCl(2) and glutamine. After co-culture, rapid removal of viable agrobacteria was crucial for subsequent development of the pollen culture. To this end, the growth of agrobacteria was suppressed by the concerted effects of appropriate antibiotics, low pH, reduced level of glutamine and high concentrations of CaCl(2) and acetosyringone. Following infection with LBA4404 and GV3101, about 31% and 69%, respectively, of the primary transgenic (T(0)) plants carried a single copy of the sequence integrated. The use of hypervirulent A. tumefaciens and hygromycin resistance as a selectable marker resulted in 3.7 T(0) plants per donor spike. About 60% of the primary transgenic plants set seed, indicating spontaneous genome doubling. An analysis of 20 T(1) populations revealed that four progenies did not segregate for reporter gene expression. This indicates that the approach pursued enables the generation of instantly homozygous primary transgenic plants. The method established will be a valuable tool in functional genomics as well as for the biotechnological improvement of barley.

Agrobacterium tumefaciens↗
PubMed2006

[Functional analysis of rice P0491E01 gene regulating anther development].

The rice P0491E01 gene shares high similarity in amino acid sequence with Arabidopsis gene AtDAD1 (DEFECTIVE IN ANTHER DEHISCENCE1) which plays a key role in the biosynthesis of jasmonic acid. In this paper, we investigated the function of this gene in the anther development of rice using RNA interference strategy. An exon fragment of 263bp was cloned from genomic DNA of P0491E01 to construct RNAi vector pP0491RNAi. Then, pP0491RNAi was transformed into O. sativa japonica by Agrobacterium-mediated transformation and ten transgenic plants were obtained. GUS-staining and PCR analysis confirmed that T-DNA was integrated into rice genome. Three of the transgenic plants were male sterile, and the other transgenic plants showed reduced fertility. Cytological observation indicated that anther development in the early stage of male sterile transgenic plants was normal, however, the microspores could not develop into mature pollen grains. Further investigations of the expression of P0491E01 gene in these transgenic lines by RT-PCR revealed that its transcription was significantly reduced. The results suggest that P0491E01 may play an important role during the late stage of anther development.

Cyclopentanes↗
PubMed2004

Asynchronous meiosis in an interspecific hybrid of Brachiaria ruziziensis and B. brizantha.

Male meiosis is generally synchronous in higher plants. The regulation of the cell cycle is still not well understood, and a powerful tool for gaining an understanding of this regulation is the development of mutations that affect cell-cycle synchrony. We report here asynchronous microsporogenesis in an interspecific hybrid between two important tropical grasses. In young spikelets of the interspecific hybrid 49.10% of anther meiocytes entered meiosis, exhibiting typical phases of the first and second divisions, while the other 50.90% showed distinctive features of early prophase. In older spikelets, anthers containing mature pollen grains also displayed meiocytes still undergoing meiosis. At this time, the latter cells were enclosed by the exine wall. Despite asynchrony, all cells completed meiosis. Old anthers contained only pollen grains that appeared to be in the same stage of development. Pollen fertility was estimated to be 52.76% in dehiscent anthers. An independent genetic control for meiosis synchrony and meiotic stages is suggested.

Brachiaria↗