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PubMed1994

Simultaneous induction of postabscission and germination mRNAs in cultured dicotyledonous embryos.

Cloned mRNAs identify three programs of gene expression in cotton (Gossypium hirsutum L.) embryos that are associated with the maturation (reserve accumulation) stage, the postabscission stage, which is marked by expression of Late-embryogenesis-abundant (Lea) mRNAs, and germination (broadly defined as including all events through early postgerminative growth). In order to test if the regulation of these programs is the same in other dicotyledonous species, their expression was studied in normal and cultured maturation-stage, postabscission-stage, and mature embryo-stage embryos or seed of oilseed rape (Brassica napus L.), soybean (Glycine max [L.] Merr.), and tobacco (Nicotiana tabacum L.) using cotton and other cDNA probes. During postabscission, Lea mRNAs accumulated in all test species and were induced in earlier maturation-stage embryos by excision and culture on basal medium. Abscisic acid often enhanced this induction in the test species. Germination-specific mRNAs were induced in cultured maturation-stage and postabscission-stage embryos of all test species. These results indicate that the regulation of embryonic and germination programs is similar in all dicotyledons tested. Because excised embryos simultaneously induced postabscission and germination programs, the effects of exogenous growth regulators and other factors on such embryos probably reflect stress responses of germinating mature embryos rather than the identity of endogenous regulators of embryogenesis.

Abscisic Acid↗
PubMed1992

Expression of the gene encoding the PR-like protein PRms in germinating maize embryos.

The PRms protein is a pathogenesis-related (PR)-like protein whose mRNA accumulates during germination of maize seeds. Expression of the PRms gene is induced after infection of maize seeds with the fungus Fusarium moniliforme. To further our investigations on the expression of the PRms gene we examined the accumulation of PRms mRNA in different tissues of maize seedlings infected with F. moniliforme and studied the effect of fungal elicitors, the mycotoxin moniliformin, the hormone gibberellic acid, and specific chemical agents. Our results indicate that fungal infection, and treatment either with fungal elicitors or with moniliformin, a mycotoxin produced by F. moniliforme, increase the steady-state level of PRms mRNA. PRms mRNA accumulation is also stimulated by the application of the hormone gibberellic acid or by treatment with silver nitrate, whereas acetylsalicylic acid has no effect. In situ RNA hybridization in isolated germinating embryo sections demonstrates that the PRms gene is expressed in the scutellum, particularly in a group of inner cells, and in the epithelium lying at the interface of the scutellum and the endosperm. The pattern of expression of the PRms gene closely resembles that found for hydrolytic enzymes, being confined to the scutellum and the aleurone layer of the germinating maize seed. Our results suggest that the PRms protein has a function during the normal process of seed germination that has become adapted to serve among the defence mechanisms induced in response to pathogens during maize seed germination.

Blotting, Northern↗
PubMed1992

Dark and light zones of germinal centres of the human tonsil: an ultrastructural study with emphasis on heterogeneity of follicular dendritic cells.

The cellular composition of the dark and light zones of germinal centres in human tonsils was quantitatively determined by electron microscopy. In addition to the well known germinal-centre B-cells, we defined the cleaved blast as a new distinct cell type in the germinal centre. The dark and the light zones clearly differed in their content of lymphoid and non-lymphoid germinal-centre cells. The dark zone was characterized by higher frequencies of cleaved blasts and small centroblasts, a higher blast-centrocyte ratio and a higher incidence of mitotic figures. In contrast, the light zone had higher frequencies of centrocytes, centroplasmacytoid cells, lymphocytes and follicular dendritic cells (FDC) and an inverted blast-centrocyte ratio. Seven distinct appearances of FDC (FDC.1-FDC.7) could be recognized on the basis of their ultrastructure. The distribution pattern of these subtypes differed between the dark and light zone. The undifferentiated subtypes FDC.2 and FDC.3 predominated in the dark zone. In contrast, the highly differentiated subtypes FDC.4 and FDC.5 were present at a much higher density in the light zone. These findings suggest that the dark and light zones are different compartments with regard to proliferation and differentiation of germinal-centre B-cells and that both compartments have a specific microenvironment influenced by FDC.

B-Lymphocytes↗
PubMed1978

Cyclic AMP and cyclic GMP in germinating conidia of Neurospora crassa.

A new method for obtaining synchronous germination allowed accurate time-course studies of endogenous levels of cyclic adenosine 3',5'-monophosphate and cyclic guanosine 3',5'-phosphate in germinating conidia of Neurospora crassa. The levels of both cyclic nucleotides remained constant throughout germination, showing that they neither signal nor respond to any of the biochemical changes which are known to occur in the germination process. Conidal germination was approximately normal in three cr-1 mutants of Neurospora which have been shown to be deficient in adenylate cyclase activity. Cyclic AMP levels in the mycelia of cr-1 mutants were low, but surprisingly, the levels in conidia were normal.

Adenylyl Cyclases↗
PubMed1980

Cyclic adenosine 3',5'-monophosphate and germination of sporangiospores from the fungus Mucor.

Cyclic adenosine 3',5'-monophosphate (cAMP) metabolism was examined in germinating sporangiospores of Mucor genevensis and Mucor mucedo. Exogenous cAMP prevented normal hyphal development from sporangiospores. Internal pools of cAMP fluctuated profoundly during development. Spherical growth of the spores was characterized by large pools of cAMP whereas germ tube emergence and hyphal elongation were characterized by small pools of cAMP. These observations suggest a possible role for cAMP in sporangiospore germination. Adenylate cyclase activities fluctuated significantly during germination with maximum values attained during spherical growth. In contrast, cAMP phosphodiesterase activities remained constant throughout germination. Internal cAMP levels may therefore be regulated by adjustment of adenylate cyclase activities. The binding of cAMP by soluble cell proteins was measured. cAMP-binding activity changed greatly during germination. Dormant and spherically growing spores possessed the highest activities. Developing hyphae contained the lowest activities. Use of the photoaffinity label, 8-azido-[32P]cAMP, in conjunction with sodium dodecyl sulfate-polyacrylamide-gel electrophoresis allowed the identification of a small population of morphogenetic-stage-specific proteins which bind cAMP and may be of regulatory significance to development.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1987

[The influence of germination on the nutritional value of wheat, mung beans and chickpeas].

The changes in nutrients during the germination of wheat, mung beans and chickpeas were investigated. Germination was performed under conditions commonly used in the household. The amount of water taken up during 4 days of germination varied from 159 g/100 g (chickpeas) to 450 g/100 g (mung beans). For all three seeds losses of dry matter and carbohydrates were observed. In wheat and mung beans, phytic acid was partially hydrolyzed. In mung beans, the total fat content decreased. Increases in the content of polyunsaturated fatty acids in wheat and of dietary fibre in wheat and mung beans were noted. At a constant level of crude protein, a measurable rise in limiting amino acids was observed in wheat and mung beans. Frequent watering during germination caused losses of Fe, between 9% and 21%, K (27% in chickpeas) and Cu (17% in chickpeas). Except for vitamin B6 in both legumes and vitamin B1 in chickpeas, accumulation of the vitamins under investigation (B1, B2, B6, C, E) was noted. Owing to these changes during germination, the nutritional value of the three seeds has been improved to various extents, most distinctly in wheat and least noticeably in chickpeas. Compared with other vegetables, sprouted seeds can be considered a valuable addition to the diet.

Fabaceae↗
PubMed1993

Slow and ultrarapid freezing of fully grown germinal vesicle-stage mouse oocytes: optimization of survival rate outweighed by defective blastocyst formation.

PURPOSE: The cryopreservation of mature metaphase II-stage mouse oocytes is associated with decreased fertilizability, spindle damage, and increased polyploidy. Therefore, we investigated the outcome of cryopreservation of immature germinal vesicle-stage mouse oocytes. METHODS: Oocytes were punctured from Graafian follicles in primed F1 hybrid mice and were then released into maturation medium containing the meiotic inhibitor dibutyryl cyclic AMP. Both slow and ultrarapid freezing protocols with dimethyl sulfoxide, 1,2-proponediol, or a mixture of both agents were tested. We recorded morphological survival rates, in vitro maturation rates, and two-cell and blastocyst formation rates. Each group of frozen oocytes was compared with both unfrozen germinal vesicle-stage oocytes and metaphase II-stage oocytes. RESULTS: An optimal cryosurvival rate of 78% was reached after ultrarapid freezing with 3 M dimethyl sulfoxide followed by one-step dilution, but a decreased rate of two-cell formation was observed. Freezing with a combination of dimethyl sulfoxide and 1,2-propanediol did not improve this fertilization-decreasing effect. Very low cryosurvival rates after freezing with 1,2-propanediol indicated its inappropriateness for ultrarapid freezing of immature oocytes. The rates of in vitro maturation were equivalent for frozen-thawed and freshly collected germinal vesicle-stage oocytes, independent of the freezing protocol used. We report, nevertheless, as a general characteristic for both slow and ultrarapid freezing of fully grown germinal vesicle-stage oocytes, that the in vitro development up to the blastocyst stage is inhibited despite full nuclear maturation. CONCLUSION: We report that cryopreservation of immature germinal vesicle-stage oocytes is invariably associated with a low developmental capacity after fertilization. The rate of in vitro nuclear maturation did not equate with developmental competence. This in turn suggests the importance of cytoplasmic maturation for embryonic development.

Analysis of Variance↗
PubMed1992

Radiation effect on ascorbic acid and riboflavin biosynthesis in germinating soybean.

The influence of irradiation on biosynthesis of ascorbic acid and riboflavin in germinating soybean seeds in tap and distilled water at ambient (25-35 degrees C) conditions was investigated. Ascorbic acid was not detectable in the original seeds and the initial level of riboflavin was 3.3 micrograms/g. The rate of synthesis of these vitamins increased with increasing germination time up to 72-96 hr followed by a decreasing pattern depending upon the treatment. The effect of irradiation and germination on the synthesis of these vitamins was statistically significant (P less than 0.01). Maximum amounts of ascorbic acid 16.2 and 15.0 mg/100g (fresh weight basis) were found in the 0.10 kGy sample after 72 hr of germination in tap and distilled water, respectively. However, a radiation dose of 0.20 kGy resulted in the development of maximum values of riboflavin, 30.0 and 27.0 micrograms/g (dry weight basis) on germination in tap and distilled water respectively.

Ascorbic Acid↗
PubMed1979

A comparative study of the lipid composition of isolated rat Sertoli and germinal cells.

The lipid composition of enriched preparations of Sertoli cells and of germinal cells, isolated from the testes of mature rats, has been investigated. Sertoli cells contained a much lower content of phospholipids (in particular, much less phosphatidylcholine and phosphatidylethanolamine) and a higher content of triacylglycerols than did germinal cells. In addition, the Sertoli cells had a higher ratio of esterified to unesterified cholesterol than did germinal cells. Total lipids of Sertoli cells contained considerably lower levels of palmitic and docosa-4,7,10,13,16-pentaenoic acids and higher levels of stearic and oleic acid than did the total lipids of germinal cells. The major phospholipid classes and the triacylglycerols of Sertoli cells similarly contained less palmitic and docosa-4,7,10,13,16-pentaenoic acids, more stearic and oleic acids and also more arachidonic acid than did the corresponding lipid classes of the germinal cells. Minor differences between cell types were also noted for the content of palmitoleic, linoleic, docosa-7,10,13,16-tetraenoic, docosa-4,7,10,13,16,19-hexaenoic and tetracosa-9,12,15,18-tetraenoic acids.

Animals↗
PubMed1980

The metabolism of polyunsaturated fatty acids in rat Sertoli and germinal cells.

The metabolism of [1-14C] linoleic, [1-14C] arachidonic and [3-14C] docosa-4,7,10,13,16-pentaenoic acids was investigated after intratesticular injection of the labeled compounds and isolation of rat Sertoli and germinal cell. Following injection of either 14C-linoleate or 14C-arachidonate, the specific activity (sp act) of docosa-4,7,10,13,16-pentaenoic acid of Sertoli cells was greater than that of the germinal cells. The data suggest that the Sertoli cells are more active in the biosynthesis of the 22-carbon pentaene than the germinal cells. Differences between these 2 cell types were also noted in the distribution of the incorporated 14C among the various lipid classes. Following intra-testicular injection of 14C-docosapentaenoic acid, a greater proportion of the recovered 14C in Sertoli cells than in germinal cells was present in 20-carton fatty acids, suggesting a greater activity in Sertoli cells in the metabolism of the pentaene. The major portion of the recovered 14C in both cell types was present in triacylglycerols during early time periods and in phospholipids after 24 hr. The possibility of transfer of biosynthesized docosapentaenoic acid from Sertoli to germinal cells is discussed.

Animals↗
PubMed1989

Uptake and metabolism of retinol in isolated cells of germinal epithelium in vitro.

Retinyl esters were quantitatively the most significant product formed in short term incubations of isolated rat germinal cells administered 3H-retinol. 3H-retinyl palmitate was found to be the single most abundant metabolite accounting for approximately 80% of the total label 2 h after administration of 3H-retinol bound to bovine serum albumin. Differences were found in the relative activity of retinol uptake and metabolism between the various subpopulations of germinal cells separated by the staput sedimentation technique. The strongest correlation was between the magnitude of 3H-retinyl palmitate synthesis and the distribution of round spermatids. Large pachytene spermatocytes also appeared to correlate with 3H-retinyl palmitate synthesis although this correlation was ambiguous due to the presence of a small population of Sertoli cells and spermatogonia in these fractions. The concentration of cellular 3H-retinol was more than 10 fold greater in germinal cells administered 3H-retinol bound to bovine serum albumin than when 3H-retinol bound to serum retinol-binding protein was administered, and likewise the concentration of cellular 3H-retinyl palmitate was approximately 30 fold higher. In comparison to germinal cells, Sertoli cells administered 3H-retinol bound to serum retinol-binding protein incorporated 20 fold more 3H-retinol and synthesized more 3H-retinyl palmitate. However, both germinal cells and Sertoli cells were found to have incorporated similar concentrations of total label when administered 3H-retinol bound to bovine serum albumin. (ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2002

Photoperiodic control of germination in the unicell Chlamydomonas.

Photoperiodic time measurement is a well-documented adaptation of multicellular plants and animals to seasonal changes in the environment, but it is unclear whether unicellular organisms can exhibit bona fide photoperiodic responses. We demonstrate that the occurrence of zygospore germination of the unicellular alga Chlamydomonas is a genuine photoperiodic response. Germination efficiency is enhanced in long days as compared with short days. While the total amount of light exposure influences the efficiency of germination, the photoperiod is a significant cue for germination. The developmental stage that senses the photoperiod is just prior to mating and during the first days of zygospore development, so there may be a critical window of zygospore maturation that is regulated by photoperiod. Because zygospores are resistant to freezing injury, whereas vegetative cells are not, it is likely that the suppression of germination by short days is an adaptive response for overwintering of Chlamydomonas. Therefore, Chlamydomonas is a single-celled organism that is capable of photoperiodic responses.

Animals↗
PubMed2004

Flow cytometry analysis of germinating Bacillus spores, using membrane potential dye.

Germination of Bacillus anthracis spores is necessary for the transcription of plasmidic genes essential to the infection. Assessing germination potential is crucial to predict the risk associated with pathogenic Bacillus exposure. The aim of this study was to set up a viability assay based on membrane potential in order to predict the earliest germination event of spores. B. cereus and two strains of B. subtilis were used. The spores were isolated with a sodium bromide gradient. Approximately 10(7) spores were incubated at 37 degrees C in tryptic soy broth (TSB). Aliquots were harvested at predetermined times and stained with 3,3'-dihexyloxacarbocyanine iodide [DiOC(6)(3)] or with bis-(1,3-dibutylbarbituric acid) trimethine oxonol [DiBAC(4)(3)]. Fluorescence characteristics were obtained using flow cytometry. The earliest detectable activation of membrane potential occurred after 15 min of incubation in TSB using DiOC(6)(3). Using DiBAC(4)(3), the earliest detectable signal was after 4 h of incubation. Control experiments using carbonyl cyanide m-chlorophenylhydrazone (CCCP)-treated spores did not show any change in the fluorescence intensity over time. Since no membrane potential and no germination were detected in CCCP-treated spores, the activation of membrane potential seems to be associated with germination. DiOC(6)(3) can be used as an early membrane potential indicator for spores. DiBAC(4)(3), by contrast, is not a early membrane potential marker.

Bacillus↗
PubMed2000

Arabidopsis thaliana germin-like proteins: common and specific features point to a variety of functions.

Germin-like proteins (GLPs) are ubiquitous plant proteins encoded by diverse multigene families. It is not known whether they share germin's unusual biochemical properties and oxalate oxidase activity. Using specific antibodies, we have studied three GLPs (AtGER1. AtGER2 and AtGER3) in Arabidopsis thaliana (L.) Heynh. as well as in transgenic tobacco (Nicotiana tabacum L.) plants overexpressing these proteins. Like wheat ( Triticum aestivum L.) germin, these Arabidopsis GLPs are associated with the extracellular matrix (ECM) and they also seem to exist as two glycosylated isoforms. However, none of them is an oxalate oxidase. Although GLPs display several conserved features, each has its specific characteristics. Both AtGER2 and AtGER3 are oligomeric proteins that share germin's resistance to pepsin and to dissociation by heat and SDS. In contrast, AtGER1 seems to exist as a monomer. The GLPs may interact with the ECM in a variety of ways, since each is efficiently extracted by different conditions. In addition, germins and GLPs all bind Cibacron Blue, a dye often but not exclusively used for the purification of enzymes having nucleotide cofactors. In the case of AtGER2, binding to the dye is so tight that it almost allows a one-step purification of this protein. The variety of sequences, expression patterns and biochemical features indicates that GLPs could be a class of receptors localized in the ECM and involved in physiological and developmental processes as well as stress response.

Arabidopsis↗
PubMed2004

Aviglycine and propargylglycine inhibit conidial germination and mycelial growth of Fusarium oxysporumf. sp. luffae.

Two inhibitors, aviglycine and propargylglycine, were tested for their ability to suppress methionine synthesis thus inhibit conidial germination and mycelial growth of Czapek-Dox liquid medium grown Fusarium oxysporum f. sp. luffaemuM. The linear inhibition range for mycelial growth was about 7.6-762.9 microM. Although aviglycine did not completely inhibit both conidial germination and mycelial growth, it showed significant inhibitory effect at 1.5 microM. The inhibition range for propargylglycine against conidial germination and mycelial growth were from 0.08 to 8841 microM and from 0.8 to 884.1 microM, respectively. Propargylglycine inhibited conidial germination and mycelial growth at a concentration of 8841 muM. The EC(50) values of aviglycine were 1 microM for conidial growth and 122 microM for mycelial growth, and the EC(50) values of propargylglycine were 47.7 microM for conidial growth and 55.6 muM for mycelial growth. Supplement of methionine released inhibition of aviglycine or propargylglycine to conidial germination. In addition, a mixture of aviglycine (1.5 microM) and propargylglycine (8841 microM) showed additive inhibitive effect than applied alone on 10 isolates. From these results, both aviglycine and propargylglycine exhibited inhibitory activity, and suggest that they can provide potential tools to design novel fungicide against fungal pathogens.

Alkynes↗
PubMed2006

A direct observation technique for evaluating sclerotium germination by Macrophomina phaseolina and effects of biocontrol materials on survival of sclerotia in soil.

Germination of sclerotia of Macrophomina phaseolina was quantified by direct microscopic observation following application of experimental treatments in vitro and incubation of sclerotia in soil. To assay germination, pieces of agar containing sclerotia were macerated in dilute, liquid cornmeal agar on glass slides; thinly spread; and incubated in a saturated atmosphere for 18-22 h. Germinated sclerotia then were identified by morphological features of germ hyphae. Frequencies of germination were similar in three dilute agar media. Germination was not affected by air-drying sclerotia for 2 weeks, but it was significantly reduced after 4 weeks and greatly reduced or eliminated after 6 or 8 weeks. Survival of sclerotia for 14 days in soil was greatest at 50, 75, and 100% moisture-holding capacity, less at 0 and 25%, and least at 125% (flooded soil). Incorporation of ground poultry litter into soil at 5% by weight reduced survival of sclerotia after 13 days, and incorporation of litter at 10% nearly eliminated it. These results indicate that the direct-observation technique may be used to evaluate animal wastes and other agricultural byproducts for biocontrol activity against sclerotia of M. phaseolina in soil.

Animals↗
PubMed2004

Germin-like protein gene family of a moss, Physcomitrella patens, phylogenetically falls into two characteristic new clades.

We identified 77 EST clones encoding germin-like proteins (GLPs) from a moss, Physcomitrella patens in a database search. These Physcomitrella GLPs ( PpGLP s) were separated into seven groups based on DNA sequence homology. Phylogenetic analysis showed that these groups were divided into two novel clades clearly distinguishable from higher plant germins and GLPs, named bryophyte subfamilies 1 and 2. PpGLPs belonging to bryophyte subfamilies 1 lacked two cysteines at the conserved positions observed in higher plant germins or GLPs. PpGLPs belonging to bryophyte subfamily 2 contained two cysteines as observed in higher plant germins and GLPs. In bryophyte subfamily 1, 12 amino acids, in which one of two cysteines is included, were deleted between boxes A and B. Further, we determined the genomic structure of all of seven PpGLP genes. The sequences of PpGLP s of bryophyte subfamily 1 contained one or two introns, whereas those of bryophyte subfamily 2 contained no introns. Other GLPs from bryophytes, a liverwort GLP from Marchantia polymorpha , and two moss GLPs from Barbula unguiculata and Ceratodon purpureus also fell into bryophyte subfamily 1 and bryophyte subfamily 2, respectively. No higher plant germins and GLPs were grouped into the bryophyte subfamilies 1 and 2 by our analysis. Moreover, we revealed that PpGLP6 had manganese-containing extracellular superoxide dismutase activity. These results indicated that bryophyte possess characteristic GLPs, which phylogenetically are clearly distinguishable from higher plant GLPs.

Amino Acid Sequence↗
PubMed1975

Characteristics of developing mitochondrial genetic and respiratory functions in germinating fungal spores.

Spores of the fungus Botryodiplodia theobromae began a cyanide-sensitive oxygen consumption immediately upon exposure to a liquid medium, and spore germination and respiration were not affected by ethidium bromide, D-threochloramphenicol, and acriflavin until later during germ tube emergence. These inhibitors of the mitochondrial genetic system all inhibited total cell protein synthesis to the same intermediate degree from the outset of incubation. When spores were incubated in water under non-germinating conditions, protein synthesis and oxygen uptake proceeded at initial rates almost identical to those seen in spores germinating in the presence of the three mitochondrial system inhibitors. Although the spores respired at rapid rates from the onset of incubation, no cytochrome absorption peaks could be observed in mitochondrial fractions prepared from ungerminated spores; they were readily observed in germinated spores, however. When the spores were germinated in the presence of inhibitors of the mitochondrial system, an excess of cytochrome c was observed in the near absence of cytochromes a and b. The results indicate that the ungerminated spores of this organism contain a preserved, potentially functional aerobic respiratory system which requires cycloheximide-sensitive ribosome activity to become functional when the spores are inoculated into a liquid medium.

Acriflavine↗