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PubMed2004

Aberrant endosperm development in interploidy crosses reveals a timer of differentiation.

The common assumption that the seed failure in interploidy crosses of flowering plants is due to parental genomic imprinting is based on vague interpretations and needs reevaluation since the general question is involved, how differentiation is timed so that cell progenies, while specializing, pass through proper numbers of amplification divisions before proliferation ceases. As recently confirmed, endosperm differentiation is accelerated or de-accelerated, depending upon whether polyploid females are crossed with diploid males, or vice-versa. Unlike the zygote, the first cell of the endosperm is determined to produce a tissue that successively induces growth of maternal tissues, stimulates and nourishes the embryo, and finally ceases cell cycling. Altered timing of endosperm differentiation, thus, perturbs seed development. During fertilization, only the female genomes contribute cytoplasmic equivalents to endosperm development so that in interploidy crosses, the initial amount of cytoplasm per chromosome set is altered, and due to semi-autonomy of cytoplasmic growth, altered numbers of division cycles are needed to provide the amount of cytoplasmic organelles required for differentiation. Cytoplasmic semi-autonomy and dependence of differentiation on an increase in cytoplasm has been shown in other tissues of plants and animals, thus, revealing a common mechanism for intracellular timing of differentiation. As demonstrated, imprinted genes can alter the extent of cell proliferation by interfering with this mechanism.

Cell Differentiation↗
PubMed2000

Enzymic activities and gene expression of enzymes of the acyl-CoA elongase during rapeseed development.

Enzymic activities and gene expression of oleoyl-CoA elongase were studied during seed development using two different rapeseed cultivars, high-erucic-acid rapeseed (HEAR) and low-erucic-acid rapeseed (LEAR). The overall elongase activities were maximal in HEAR between the fourth and eighth weeks after pollination (WAP) and absent in LEAR. The 3-ketoacyl-CoA synthase (condensing enzyme, CE) mRNA levels and the developmental profiles in the two cultivars were different since maximal expression levels were detected in HEAR and LEAR at WAP 4 and WAP 6, respectively. Anti-CE antibodies revealed two proteins of 60 and 67 kDa in both cultivars and an additional reacting protein of 57 kDa in HEAR.

Acyl-Carrier Protein S-Malonyltransferase↗
PubMed1998

cis-isomers of cytokinins predominate in chickpea seeds throughout their development

Trans-isomers of cytokinins (CK) are thought to predominate and have greater biological activity than corresponding cis-isomers in higher plants. However, this study demonstrates a system within which the predominant CK are cis-isomers. CK were measured at four developmental stages in developing chickpea (Cicer arietinum L. cultivar Kaniva) seeds by gas chromatography-mass spectrometry. Concentrations were highest at an early endospermic fluid stage and fell considerably when the cotyledons expanded. The cis-isomers of zeatin nucleotide ([9R-MP]Z), zeatin riboside ([9R]Z), and zeatin (Z) were present in greater concentrations than those of corresponding trans-isomers: (trans)[9R-MP]Z, (trans)[9R]Z, (trans)Z, or dihydrozeatin riboside. Dihydrozeatin, dihydrozeatin nucleotide, and the isopentenyl-type CK concentrations were either low or not detectable. Root xylem exudates also contained predominantly cis-isomers of [9R-MP]Z and [9R]Z. Identities of (cis)[9R]Z and (cis)Z were confirmed by comparison of ion ratios and retention indices, and a full spectrum was obtained for (cis)[9R]Z. Tissues were extracted under conditions that minimized the possibility of RNase hydrolysis of tRNA following tissue disruption, being a significant source of the cis-CK. Since no isomerization of (trans)[2H]CK internal standards occurred, it is unlikely that the cis-CK resulted from enzymic or nonenzymic isomerization during extraction. Although quantities of total CK varied, similar CK profiles were found among three different chickpea cultivars and between adequately watered and water-stressed plants. Developing chickpea seeds will be a useful system for investigating the activity of cis-CK or determining the origin and metabolism of free CK.

Journal Article↗
PubMed1997

Silencing of a beta-1,3-glucanase transgene is overcome during seed formation.

Expression of a beta-1,3-glucanase transgene (gn1) driven by the CaMV 35S promoter is silenced in the T17 homozygous tobacco transgenic line. This silencing process is post-transcriptionally regulated and subject to developmental control. We have examined this phenomenon to investigate the developmental pathways involved in suppression and reactivation of gn1 expression as well as to identify the plant tissues where these processes occur. Analysis of beta-1,3-glucanase activity and gene expression have allowed us to determine that suppression of gn1 is a very efficient process reducing the steady-state gn1 mRNA level, simultaneously, in all leaves of the plant. Gene silencing occurs a few weeks after seed germination, and is maintained throughout vegetative growth and floral development. Expression of gn1 is restored in the maturing fruit some time after fertilization. In situ hybridization analyses show that expression of gn1 is restored within the developing seeds in tissues derived from meiotically divided cells. In contrast to the high level of expression found in seedlings obtained from germinated T17 homozygous seeds, the expression of gn1 is not reactivated in plantlets regenerated in vitro from leaf explants of suppressed T17 homozygous plants that is, in plant tissues obtained by mitotic division. Thus, reactivation of gn1 expression specifically occurs along the developmental programme controlling sexual reproduction and likely throughout epigenetic modifications affecting the state of gene expression during meiosis.

Fruit↗
PubMed2005

The delayed initiation and slow elongation of fuzz-like short fibre cells in relation to altered patterns of sucrose synthase expression and plasmodesmata gating in a lintless mutant of cotton.

Cotton (Gossypium hirsutum L.) seed develops single-celled long fibres (lint) from the seed-coat epidermis at anthesis. Previous studies have shown that the initiation and rapid elongation of these fibres requires the expression of sucrose synthase (Sus) and, potentially, a transient closure of plasmodesmata. This study extends the previous work to examine the patterns of Sus expression and plasmodesmata gating in fuzz-like short fibres of a mutant that shows delayed initiation and much slower and reduced elongation of the fibre cells. Immunolocalization studies revealed delayed expression of Sus in the mutant seed-coat epidermis that correlates temporally and spatially with the initiation of the fibre cells. Anatomically, these short fibres differed from the normal lint in that their basal ends enlarged immediately after initiation, while the majority of the normal lint on wild-type seed did not show this enlargement until the end of elongation. Suppression of Sus expression in the seed-coat epidermis of the transgenic plants reduced the length of both lint and short fuzz fibres at maturity, suggesting that the growth of short fibres also requires high levels of Sus expression. Confocal imaging of the membrane-impermeant fluorescent solute carboxyfluorescein (CF) revealed no closure of plasmodesmata during the entire elongation period of short fibres from the mutant seed. These results show (i) the delayed initiation of fuzz-like short fibres from the mutant seed correlates with delayed or insufficient expression of Sus in a subset of seed-coat epidermal cells destined to become fibres and (ii) the much shortened elongation of the fibres from the mutant may be related to their inability to close plasmodesmata.

Cotton Fiber↗
PubMed1995

Differential regulation of the expression in transgenic tobacco of the gene for beta-glucuronidase under the control of the 5'-upstream regions of two catalase genes from castor bean.

The regulatory functions of the 5'-flanking regions of two genes for catalase (cat1 and cat2) from castor bean were analyzed in transgenic tobacco plants that carried fusion constructs that included the gene for beta-glucuronidase (GUS) for Escherichia coli. Dry mature seeds from transgenic plants carrying the CAT1-GUS or CAT2-GUS constructs, in which the GUS gene was fused to the 5'-flanking region of cat1 or cat2, respectively, contained significant GUS activity, indicating that the promoters of cat1 and cat2 were active during seed development. GUS activity increased in response to germination in the seeds of transgenic tobacco that carried CAT1-GUS, as well as in those that carried CAT2-GUS. During the post-germinative stage the GUS activity directed by CAT2-GUS increased still further, whereas that directed by CAT1-GUS decreased. The changes in GUS activity in the transgenic tobacco plants that carried CAT1-GUS and CAT2-GUS were similar to the changes in the levels of transcripts of cat1 and cat2, respectively, in castor bean. The results suggest that the expression of cat1 and cat2 in the germinating seeds and post-germinative seedlings is regulated mainly at the level of transcription. However, the distribution of GUS activity among the organs of the transgenic tobacco seedlings and plantlets, which was examined by histochemical staining and by enzymatic assays of tissue extracts, was not identical to that of transcripts of cat1 and cat2 in castor bean. Histochemical analysis also revealed the interesting spatial regulation of the expression of the promoter of cat2 in the transgenic tobacco seedlings.

Base Sequence↗
PubMed2002

Ecological and evolutionary conditions for fruit abortion to regulate pollinating seed-eaters and increase plant reproduction.

Coevolved mutualisms, such as those between senita cacti, yuccas, and their respective obligate pollinators, benefit both species involved in the interaction. However, in these pollination mutualisms the pollinator's larvae impose a cost on plants through consumption of developing seeds and fruit. The effects of pollinators on benefits and costs are expected to vary with the abundance of pollinators, because large population sizes result in more eggs and larval seed-eaters. Here, we develop the hypothesis that fruit abortion, which is common in yucca, senita, and plants in general, could in some cases have the function of limiting pollinator abundance and, thereby, increasing fruit production. Using a general steady-state model of fruit production and pollinator dynamics, we demonstrate that plants involved in pollinating seed-eater mutualisms can increase their fecundity by randomly aborting fruit. We show that the ecological conditions under which fruit abortion can improve plants fecundity are not unusual. They are best met when the plant is long-lived, the population dynamics of the pollinator are much faster than those of the plant, the loss of one fruit via abortion kills a larva that would have the expectation of destroying more than one fruit through its future egg laying as an adult moth, and the effects of fruit abortion on pollinator abundance are spatially localized. We then use the approach of adaptive dynamics to find conditions under which a fruit abortion strategy based on regulating the pollinator population could feasibly evolve in this type of plant-pollinator interaction.

Animals↗
PubMed2006

Proteomic analysis of seed filling in Brassica napus. Developmental characterization of metabolic isozymes using high-resolution two-dimensional gel electrophoresis.

Brassica napus (cultivar Reston) seed proteins were analyzed at 2, 3, 4, 5, and 6 weeks after flowering in biological quadruplicate using two-dimensional gel electrophoresis. Developmental expression profiles for 794 protein spot groups were established and hierarchical cluster analysis revealed 12 different expression trends. Tryptic peptides from each spot group were analyzed in duplicate using matrix-assisted laser desorption ionization time-of-flight mass spectrometry and liquid chromatography-tandem mass spectrometry. The identity of 517 spot groups was determined, representing 289 nonredundant proteins. These proteins were classified into 14 functional categories based upon the Arabidopsis (Arabidopsis thaliana) genome classification scheme. Energy and metabolism related proteins were highly represented in developing seed, accounting for 24.3% and 16.8% of the total proteins, respectively. Analysis of subclasses within the metabolism group revealed coordinated expression during seed filling. The influence of prominently expressed seed storage proteins on relative quantification data is discussed and an in silico subtraction method is presented. The preponderance of energy and metabolic proteins detected in this study provides an in-depth proteomic view on carbon assimilation in B. napus seed. These data suggest that sugar mobilization from glucose to acetyl-coenzyme A [corrected] is a collaboration between the cytosol and plastids and that temporal control of enzymes and pathways extends beyond transcription. This study provides a systematic analysis of metabolic processes operating in developing B. napus seed from the perspective of protein expression. Data generated from this study have been deposited into a web database (http://oilseedproteomics.missouri.edu) that is accessible to the public domain.

Arabidopsis↗
PubMed2001

Hsp101 is necessary for heat tolerance but dispensable for development and germination in the absence of stress.

Hsp101 is a molecular chaperone that is required for the development of thermotolerance in plants and other organisms. We report that Arabidopsis thaliana Hsp101 is also regulated during seed development in the absence of stress, in a pattern similar to that seen for LEA proteins and small Hsps; protein accumulates during mid-maturation and is stored in the dry seed. Two new alleles of the locus encoding Hsp101 (HOT1) were isolated from Arabidopsis T-DNA mutant populations. One allele, hot1-3, contains an insertion within the second exon and is null for Hsp101 protein expression. Despite the complete absence of Hsp101 protein, plant growth and development, as well as seed germination, are normal, demonstrating that Hsp101 chaperone activity is not essential in the absence of stress. In thermotolerance assays hot1-3 shows a similar, though somewhat more severe, phenotype to the previously described missense allele hot1-1, revealing that the hot1-1 mutation is also close to null for protein activity. The second new mutant allele, hot1-2, has an insertion in the promoter 101 bp 5' to the putative TATA element. During heat stress the hot1-2 mutant produces normal levels of protein in hypocotyls and 10-day-old seedlings, and it is wild type for thermotolerance at these stages. Thus this mutation has not disrupted the minimal promoter sequence required for heat regulation of Hsp101. The hot1-2 mutant also expresses Hsp101 in seeds, but at a tenfold reduced level, resulting in reduced thermotolerance of germinating seeds and underscoring the importance of Hsp101 to seed stress tolerance.

Adaptation, Physiological↗
PubMed1998

Rice bifunctional alpha-amylase/subtilisin inhibitor: characterization, localization, and changes in developing and germinating seeds.

A bifunctional alpha-amylase/subtilisin inhibitor (RASI) was purified to electrophoretic homogeneity from rice (Oryza sativa L.) bran. Its molecular mass was 21 kDa by SDS-PAGE and its isoelectric point was 9.05. Purified RASI inhibited subtilisin Carlsberg strongly and inhibited alpha-amylase from germinating rice seeds weakly. It inhibited rice alpha-amylase more than barley alpha-amylase, and the inhibition of rice alpha-amylase was greater at higher pHs. RASI did not inhibit trypsin, chymotrypsin, cucumisin, or mammalian alpha-amylase. The RASI was in the outermost part of the rice grain and its subcellular site seemed to be aleurone particles in aleurone cells. SDS-PAGE and western blotting showed that RASI was synthesized in the late milky stage in developing seeds, and it remained fairly constant during the first 7 days of germination.

Blotting, Western↗
PubMed1978

Isolation of polyribosomes and messenger RNA active in in vitro synthesis of soybean seed proteins.

Polyribosome preparations containing low proportions of monosomes to polyribosomes have been isolated from developing seeds of Glycine max L. Merrill using a high pH-high KCl buffer. The polyribosomes were functional in in vitro protein synthesis reactions using wheat germ 23,000g supernatant preparations. Results of experiments using aurintricarboxylic acid indicated that most or all of the amino acid incorporation in vitro resulted from the completion of nascent polypeptides associated with the isolated polyribosmes. RNA purified from polyribosome preparations by affinity chromatography on oligo(dT)-cellulose was also active in vitro, and had different Mg and K requirements for translation than did the polyribosomes. Translation of oligo(dT)-cellulose-purified mRNA was inhibited by the addition of 7-methylguanosine 5'-phosphate, suggesting that soybean mRNAs are "capped" at their 5' ends. Some, but not all, of the products of these reactions were identical in electrophoretic mobility to radioactive polypeptides of storage proteins produced in soybean cotyledons grown in culture.

Journal Article↗
PubMed2006

Megasporogenesis, megagametogenesis and ontogeny of the aril in Cytisus striatus and C. multiflorus (Leguminosae: Papilionoideae).

BACKGROUND AND AIMS: There are few embryological reports on wild legumes and even fewer on their seminal appendages. There are no existing studies on the complete ontogeny of these appendages in Cytiseae, a very important Papilionoideae tribe in Mediterranean ecosystems. In this work megasporogenesis, megagametogenesis and aril ontogeny were studied in Cytisus multiflorus and C. striatus, endemics from the western Mediterranean region. METHODS: Ovaries and ovules from flower buds, flowers at anthesis and hand cross-pollinated flowers were sectioned with a rotary microtome and studied under light and fluorescence microscopy. KEY RESULTS: A monosporic Polygonum-type of megagametogenesis is observed in both species but with megasporogenesis characterized by formation of a triad of cells after incomplete meiosis. The original cell wall of the megaspore mother cell and triad, including the transverse walls between the latter, are surrounded by a callose layer that isolates them from the surrounding diploid tissue; this callose layer gradually disappears during embryo sac formation. There are no antipodals in the mature embryo sac. Aril ontogeny starts in pre-anthesis with the formation of the aril primordium, and its normal development will occur only after fertilization, more specifically after endosperm initiation. After fertilization, a reactivation of meristem capacity takes place in the aril cells resulting in slow and sparse growth. Later, this type of development gradually decreases but the aril cells continue to grow by cell expansion, which in the last period of seed development is the only type of growth of the aril. In the mature seed, the seminal appendage acquires an irregular U-shape in transverse section, showing vacuolated cells with a large central vacuole that stores lipids and some proteins. CONCLUSIONS: Meiotic triad formation is due to a failure in meiosis II of the chalazal cell of the dyad. In Cytisus seeds the aril has a funicular origin with predominantly post-fertilization development, but a normal growth of the endosperm is needed for proper aril development.

Cytisus↗
PubMed1997

An extracellular insoluble inhibitor of cysteine proteinases in cell cultures and seeds of carrot.

An 18 kDa extracellular insoluble protein (EIP18) was found previously in amorphous particles suspended in the culture medium and in the interspaces of cell clusters of carrot (Daucus carota L.) callus, as well as in the extracellular spaces of carrot seeds, being located both in the embryo and at the inner edge of the endosperm. We purified EIP18 by washing the amorphous particles with the mixture of Triton X-100, NaCl and ethylenediaminetetraacetic acid (EDTA). We determined several partial amino acid sequences, and then we cloned and sequenced a cDNA for EIP18. EIP18 was found to consist of 133 amino acid residues that included a signal sequence, but it did not contain cysteine, sites for N-linked glycosylation or hydrophobic regions. Since its sequence was found to be homologous to that of inhibitors of cysteine proteinases, namely cystatins, EIP18 was renamed EICC (extracellular insoluble cystatin of carrot). EICC expressed in yeast was also found in an insoluble form in yeast cell walls. EICC prepared from the culture medium of carrot cells inhibited commercial cysteine proteinases and a proteinase extracted from germinating carrot seeds. The expression of the gene for EICC was detected in developing seeds, and the level of its transcript was markedly enhanced upon treatment of somatic embryos with abscisic acid.

Amino Acid Sequence↗
PubMed2000

AtZFP1, encoding Arabidopsis thaliana C2H2 zinc-finger protein 1, is expressed downstream of photomorphogenic activation.

C2H2 zinc-finger proteins play important roles in plant development including floral organogenesis, leaf initiation, lateral shoot initiation, gametogenesis and seed development. The gene for one such protein from Arabidopsis, AtZFP1 (Arabidopsis thaliana zinc-finger protein 1), is expressed at high levels in the shoot apex, including the apical meristem, developing leaves and the developing vascular system. In light-grown seedlings, AtZFP1 expression is induced about three days after germination, before the expansion of the true leaves. Dark-grown plants, in which photomorphogenesis is repressed, have no detectable AtZFP1 expression in the shoot apex. Under conditions which induce or mimic photomorphogenic development including growth in the light, shifting dark-grown plants to continuous light or growth on cytokinin in the dark, high levels of AtZFP1 expression are detected. Furthermore, AtZFP1 expression does not depend on active photosynthesis as shown by analysis of plants grown on the carotenoid biosynthetic inhibitor norflurazon. These results are discussed in relation to a possible role for AtZFP1 in shoot development, downstream of photomorphogenic activation.

Arabidopsis↗
PubMed1995

Effect of the Growth Retardant 3,5-Dioxo-4-butyryl-cyclohexane Carboxylic Acid Ethyl Ester, an Acylcyclohexanedione Compound, on Fruit Growth and Gibberellin Content of Pollinated and Unpollinated Ovaries in Pea.

Treatment of pollinated pea (Pisum sativum L. cv Alaska, line V1) ovaries with 3,5-dioxo-4-butyryl-cyclohexane carboxylic acid ethyl ester (LAB), an acylcyclohexanedione derivative that competitively inhibits 2-oxoglutarate-dependent gibberellin (GA) dioxygenases, caused a reduction of pod elongation proportional to the amount of inhibitor applied. The effect of LAB was counteracted by GA1 and GA3, and partially by GA20. The inhibitor decreased the contents of GA1 and GA3 (the purported active GAs) and GA8, increased those of GA19 and GA20, and did not affect that of GA29 in both the pod and the developing seeds. These results provide evidence that GA1 and/or GA3 control pod development in pea and show that GA20 is not active per se. In contrast to its effect on pollinated ovaries, LAB promoted parthenocarpic development of unpollinated ovaries, which is associated with an increase of GA1 and GA8 content. The inhibitor enhanced the response of unpollinated ovaries to GA1 and GA20, but it did not alter the response to GA3. LAB is proposed to promote parthenocarpic development and enhance the response to exogenous GAs by blocking the 2[beta]-hydroxylation of GA1 more efficiently than 3[beta]-hydroxylation of GA20.

Journal Article↗
PubMed2002

Contrapuntal networks of gene expression during Arabidopsis seed filling.

We have used cDNA microarrays to examine changes in gene expression during Arabidopsis seed development and to compare wild-type and mutant wrinkled1 (wri1) seeds that have an 80% reduction in oil. Between 5 and 13 days after flowering, a period preceding and including the major accumulation of storage oils and proteins, approximately 35% of the genes represented on the array changed at least twofold, but a larger fraction (65%) showed little or no change in expression. Genes whose expression changed most tended to be expressed more in seeds than in other tissues. Genes related to the biosynthesis of storage components showed several distinct temporal expression patterns. For example, a number of genes encoding core fatty acid synthesis enzymes displayed a bell-shaped pattern of expression between 5 and 13 days after flowering. By contrast, the expression of storage proteins, oleosins, and other known abscisic acid-regulated genes increased later and remained high. Genes for photosynthetic proteins followed a pattern very similar to that of fatty acid synthesis proteins, implicating a role in CO(2) refixation and the supply of cofactors for oil synthesis. Expression profiles of key carbon transporters and glycolytic enzymes reflected shifts in flux from cytosolic to plastid metabolism. Despite major changes in metabolism between wri1 and wild-type seeds, <1% of genes differed by more than twofold, and most of these were involved in central lipid and carbohydrate metabolism. Thus, these data define in part the downstream responses to disruption of the WRI1 gene.

Arabidopsis↗
PubMed1980

Agmatine iminohydrolase activity during development and germination of groundnut seeds.

During the development of groundnut seeds agmatine iminohydrolase activity increased in the cotyledons but remained constant in the embryo. In seeds stored for 1 year, decreased activity of the enzyme was found in both cotyledons and embryo. During germination, the enzyme activity increased in the cotyledons, but in the embryo it increased up to day 3 and then decreased to the initial level.

Arachis↗
PubMed1981

Morphology and ultrastructure of maternal seed tissues of soybean in relation to the import of photosynthate.

Cytological observations were made on developing seeds of soybean (Glycine max (L.) Merr. "Amsoy 71") using scanning and transmission electron microscopy and light microscopy. Attention was focused on the maternal tissues of the seed coat and embryo sac. An hypothesis of photosynthate import, unloading, and movement to the embryo is presented based on the results of these studies.

Journal Article↗