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Delivery of germinal granules and localized RNAs via the messenger transport organizer pathway to the vegetal cortex of Xenopus oocytes occurs through directional expansion of the mitochondrial cloud.

During Xenopus oogenesis, the message transport organizer (METRO) pathway delivers germinal granules and localized RNAs to the vegetal cortex of the oocyte via the mitochondrial cloud (Balbiani body). According to the traditional model, the mitochondrial cloud is thought to break up at the onset of vitellogenesis and the germinal granules and METRO-localized RNAs are transported within the mitochondrial cloud fragments to the vegetal cortex of the oocyte. We used light and electron microscopy in situ hybridization and three-dimensional reconstruction to show that germinal granules and METRO-localized RNAs are delivered to the oocyte cortex before the onset of mitochondrial cloud fragmentation and that the delivery involves accumulation of localized RNAs and aggregation of germinal granules at the vegetal tip of the mitochondrial cloud and subsequent internal expansion of the mitochondrial cloud between its animal (nuclear) and vegetal tips, which drives the germinal granules and METRO-localized RNAs toward the vegetal cortex. Thus the fragmentation of the cloud that occurs later in oogenesis is irrelevant to the movement of METRO-localized RNAs and germinal granules. On the basis of these findings, we propose here a revised model of germinal granule and localized RNAs delivery to the oocyte vegetal cortex via the METRO pathway.

Animals↗

Inducible nitric oxide synthase-derived nitric oxide regulates germinal vesicle breakdown and first polar body emission in the mouse oocyte.

The present study investigated the subcellular localization of inducible nitric oxide synthase (iNOS) during mouse oocyte meiotic maturation and fertilization using confocal microscopy, and further studied the roles of iNOS-derived NO in oocyte maturation by using an iNOS-specific inhibitor aminoguanidine (AG) and iNOS antibody microinjection. In germinal vesicle-stage oocytes, iNOS immunoreactivity was mainly localized in the germinal vesicle. Shortly after germinal vesicle breakdown, the iNOS immunoreactivity accumulated around the condensed chromosomes. At metaphase I and metaphase II, with the organization of chromosomes to the equatorial plate, iNOS immunoreactivity was concentrated around the aligned chromosomes, putatively the position of the metaphase spindle. The accumulation of iNOS immunoreactivity could not be detected at anaphase I and anaphase II. However, at telophase I and telophase II, the staining of iNOS was concentrated in the region between the separating chromosomes/chromatids. Furthermore, the staining of iNOS also accumulated in the male and female pronuclei in fertilized eggs. Germinal vesicle breakdown and the first polar body emission of the oocytes were significantly blocked by the iNOS-specific inhibitor AG in a dose-dependent manner. The germinal vesicle breakdown in oocytes injected with iNOS antibody was also inhibited. We found that the phosphorylation of mitogen-activated protein kinase in oocytes after germinal vesicle breakdown was inhibited by AG treatment. The control oocytes extruded a normal first polar body, while the AG-treated oocytes exhibited an elongated protrusion or no elongated protrusion. The results of confocal microscopy showed that the AG-treated oocytes were arrested at anaphase I-telophase I. Our results suggest that the iNOS-derived NO pathway plays important roles in mouse oocyte meiotic maturation, especially in germinal vesicle breakdown and the anaphase-telophase transition.

Animals↗

The heterotrimeric G-protein GanB(alpha)-SfaD(beta)-GpgA(gamma) is a carbon source sensor involved in early cAMP-dependent germination in Aspergillus nidulans.

The role of heterotrimeric G-proteins in cAMP-dependent germination of conidia was investigated in the filamentous ascomycete Aspergillus nidulans. We demonstrate that the G alpha-subunit GanB mediates a rapid and transient activation of cAMP synthesis in response to glucose during the early period of germination. Moreover, deletion of individual G-protein subunits resulted in defective trehalose mobilization and altered germination kinetics, indicating that GanB(alpha)-SfaD(beta)-GpgA(gamma) constitutes a functional heterotrimer and controls cAMP/PKA signaling in response to glucose as well as conidial germination. Further genetic analyses suggest that GanB plays a primary role in cAMP/PKA signaling, whereas the SfaD-GpgA (G betagamma) heterodimer is crucial for proper activation of GanB signaling sensitized by glucose. In addition, the RGS protein RgsA is also involved in regulation of the cAMP/PKA pathway and germination via attenuation of GanB signaling. Genetic epistatic analyses led us to conclude that all controls exerted by GanB(alpha)-SfaD(beta)-GpgA(gamma) on conidial germination are mediated through the cAMP/PKA pathway. Furthermore, GanB may function in sensing various carbon sources and subsequent activation of downstream signaling for germination.

Aspergillus nidulans↗

Lymph follicles and germinal centers in popliteal lymph nodes and other lymphoid tissues of germ-free and conventional rats.

A study was conducted to evaluate the influence of natural exogenous antigen stimulation on the development of lymph follicles in the peripheral lymphoid organs of the rat. The number of lymph follicles and germinal centers per popliteal node as well as the morphological features of popliteal nodes, mesenteric nodes and Peyer's patches were compared between 8-week-old male Sprague-Dawley rats reared in germ-free, specific pathogen-free (SPF) and conventional environments. In mesenteric nodes and Peyer's patches from conventional and SPF rats, almost every follicle contained a germinal center. In the mesenteric nodes from germ-free rats, only 9 of 195 lymph follicles examined contained a germinal center, but interestingly, in Peyer's patches all the lymph follicles examined showed a fairly well developed germinal center. The popliteal nodes from germ-free rats had no germinal centers and each node contained about 80 lymph follicles. In six conventional rats and two of five SPF rats used, the number of lymph follicles per popliteal node usually ranged from 100 to 130, and some lymph follicles contained a germinal center. However, in the other three SPF rats the popliteal node on both sides showed no germinal center, and each contained almost the same number of lymph follicles as the popliteal nodes of other SPF and conventional rats. The present results are consistent with the view that even in the absence of exogenous antigen stimulation, a regional lymph node of the germ-free rat develops a substantial number of primary lymph follicles by way of a non-immunological cellular activity, and that natural exogenous antigens may influence the development of lymph follicles by stimulating the cellular activity rather than evoking the humoral immune responses.

Animals↗

Predictive model for Clostridium perfringens growth in roast beef during cooling and inhibition of spore germination and outgrowth by organic acid salts.

Spores of foodborne pathogens can survive traditional thermal processing schedules used in the manufacturing of processed meat products. Heat-activated spores can germinate and grow to hazardous levels when these products are improperly chilled. Germination and outgrowth of Clostridium perfringens spores in roast beef during chilling was studied following simulated cooling schedules normally used in the processed-meat industry. Inhibitory effects of organic acid salts on germination and outgrowth of C. perfringens spores during chilling and the survival of vegetative cells and spores under abusive refrigerated storage was also evaluated. Beef top rounds were formulated to contain a marinade (finished product concentrations: 1% salt, 0.2% potassium tetrapyrophosphate, and 0.2% starch) and then ground and mixed with antimicrobials (sodium lactate and sodium lactate plus 2.5% sodium diacetate and buffered sodium citrate and buffered sodium citrate plus 1.3% sodium diacetate). The ground product was inoculated with a three-strain cocktail of C. perfringens spores (NCTC 8238, NCTC 8239, and ATCC 10388), mixed, vacuum packaged, heat shocked for 20 min at 75 degrees C, and chilled exponentially from 54.5 to 7.2 degrees C in 9, 12, 15, 18, or 21 h. C. perfringens populations (total and spore) were enumerated after heat shock, during chilling, and during storage for up to 60 days at 10 degrees C using tryptose-sulfite-cycloserine agar. C. perfringens spores were able to germinate and grow in roast beef (control, without any antimicrobials) from an initial population of ca. 3.1 log CFU/g by 2.00, 3.44, 4.04, 4.86, and 5.72 log CFU/g after 9, 12, 15, 18, and 21 h of exponential chilling. A predictive model was developed to describe sigmoidal C. perfringens growth curves during cooling of roast beef from 54.5 to 7.2 degrees C within 9, 12, 15, 18, and 21 h. Addition of antimicrobials prevented germination and outgrowth of C. perfringens regardless of the chill times. C. perfringens spores could be recovered from samples containing organic acid salts that were stored up to 60 days at 10 degrees C. Extension of chilling time to > or =9 h resulted in >1 log CFU/g growth of C. perfringens under anaerobic conditions in roast beef. Organic acid salts inhibited outgrowth of C. perfringens spores during chilling of roast beef when extended chill rates were followed. Although C. perfringens spore germination is inhibited by the antimicrobials, this inhibition may represent a hazard when such products are incorporated into new products, such as soups and chili, that do not contain these antimicrobials, thus allowing spore germination and outgrowth under conditions of temperature abuse.

Animals↗

Germicidin, an autoregulative germination inhibitor of Streptomyces viridochromogenes NRRL B-1551.

During germination spores of Streptomyces viridochromogenes NRRL B-1551 excrete a compound, germicidin, which has an inhibitory effect on the germination of its own arthrospores at a concentration as low as 200 pM (40 pg/ml). At higher concentrations germicidin inhibits porcine Na+/K(+)-activated ATPase and retards the germination of the cress Lepidium sativum. Germicidin is the first known autoregulative inhibitor of spore germination in the genus Streptomyces and was isolated from the supernatant of germinated spores, but also from the supernatant of the submerged culture. Spectroscopic analysis and derivatization reactions revealed germicidin to be 6-(2-butyl)-3-ethyl-4-hydroxy-2-pyrone (C11H16O3). Crude isolates of germicidin from the supernatant of submerged culture, but not from the spores, contained a second, structurally very similar compound (C10H14O3), in which in contrast to germicidin a 2-propyl instead of the 2-butyl chain was bound to C-6 and which did not show any activity in the germination and ATPase assay. The germination assay was evaluated as a new screening model for specifically active compounds.

Fermentation↗

Volumetric analysis of the germinal matrix and lateral ventricles performed using MR images of postmortem fetuses.

BACKGROUND AND PURPOSE: The volumetric changes of the ventricular system and germinal matrix are important to understand brain maturation and the mechanism of subependymal hemorrhage. Our purpose was to show the 3D configuration of the brain, germinal matrix, and lateral ventricles and to discuss the volumetric changes of each structure with maturation by using high-resolution MR imaging. METHODS: Three-dimensional MR images of 13 formalin-fixed fetal brains ranging from 7 to 28 weeks' gestational age (GA) were obtained on a 4.7-T unit. Each 3D configuration of the brain surface, germinal matrix, and ventricles was rendered from the cross-sectional imaging data sets and its volume measured. RESULTS: The germinal matrix was detected on MR images at 9 weeks' GA. Its volume exponentially increased by 23 weeks' GA (maximum, 2346 mm3) and then sharply decreased at 28 weeks' GA. The volume of the lateral ventricles increased gradually and reached 2646 mm3 peak volume at 23 weeks' GA. Between 11 and 23 weeks' GA, total brain and germinal matrix volumes were exponentially increasing, but the volume ratio of germinal matrix to brain was stable at about 5%. On the other hand, the volume ratio of lateral ventricles to brain was large between 10 and 13 weeks' GA. This period corresponded to the lateral ventricle showing a "vesicular" aspect with a thin mantle, and the developing mantle thickness of the hemisphere resulted in the decreasing ratio. CONCLUSION: Volumetric information concerning the germinal matrix and lateral ventricles may be useful in the accurate interpretation of clinical echograms and MR images of the fetal brain in utero.

Brain↗

[Germination of basidiospores of Agaricus bisporus].

The type of dormancy and conditions necessary for germination of Agaricus bisporus basidiospores (BS) were studied. BS failed to germinate on starvation agar and required the presence of carbon and nitrogen (asparagine and/or glucose) sources in the medium. Upon 3-week storage, BS germinated after 4-5 days. Heat shock (20 min at 45 degrees C) and the decreased temperature facilitated activation of germination. Heterocyclic compounds stimulating germination of endogenously dormant spores, such as furfural, failed to activate germination. The data obtained suggested an endogenous dormancy of A. bisporus BS differing from zygospores of Mucorales. BS contained 17-19% lipids with a composition of fatty acids differing from those of pileus and stipe of the fruiting body. The soluble carbohydrates of the cytosol amounted to 12% dry spore weight and consisted of mannitol (74%) and trehalose (26%). Unlike BS stored at 2 degrees C, the BS stored for 5 months at 20 degrees C lost their ability to germinate, which correlated with a decrease in the content of trehalose.

Agaricus↗

The ontogeny of germinal centre forming capacity of neonatal rat spleen.

In non-specifically immunized rats, bred under conventional conditions, the first 'spontaneous' germinal centres were observed by 21 days after birth. Deliberate antigenic stimulation led to an earlier appearance of germinal centres in neonatal spleen: immunization with sheep red blood cells as early as 7 days after birth resulted in germinal centre formation in the spleen as observed 7 days later. By that time the first primary follicles could also be observed, in both immunized and non-immunized rats. Although 3-day-old rats upon antigenic stimulation failed to generate germinal centres in their spleen, transfer experiments of 3-day-old spleen cells to lethally X-irradiated syngeneic adult recipients indicated that 3-day-old spleens at least contained all the essential lymphoid elements (B and T cells) needed for germinal centre formation. These results strongly suggest that the failure to induce germinal centres in 3-day-old rats is most probably due to an immaturity of their splenic microenvironment. Immunohistochemical staining of frozen sections of neonatal rat spleen using mAb ED 5 and MRC OX-2 showed that follicular dendritic cells (FDC) were found as soon as primary follicles were found (i.e. by 14 days after birth). The appearance of FDC in neonatal spleens was not influenced by deliberate antigenic stimulation nor by the administration of adult spleen cells. We postulate that, during the development of FDC, a splenic microenvironment is created that allows primary follicle formation and the generation of germinal centres.

Animals↗

Changes of enzymes and factors involved in DNA synthesis during wheat embryo germination.

We have previously purified and characterized wheat germ DNA polymerases A and B. To determine the role played by DNA polymerases A and B in DNA replication, we have measured the level of their activities during wheat embryo germination. The level of cellular proteins known to be associated with DNA synthesis such as PCNA and DNA primase were also investigated. The activity of DNA polymerase A gradually increased reaching a maximal level at 12 h after germination. Three days later, only a residual activity was detected. DNA polymerase B showed the same pattern during germination with very similar changes in activity. Our results indicate a striking correlation between maximal activities of DNA polymerase A, DNA polymerase B and optimal levels of DNA synthesis. These results support a replicative role of these enzymes. The activity of wheat DNA primase that copurifies with DNA polymerase A also increases during wheat germination. Taking together all its properties, and in spite of its behaviour with some inhibitors. DNA polymerase A may be considered as the plant counterpart of animal DNA polymerase alpha. Concerning DNA polymerase B we have previously shown that PCNA stimulates its processivity. Besides studying the changes of DNA polymerases A and B and DNA primase we have also studied changes in PCNA during germination. We show that PCNA is present in wheat embryos at a constant relatively high level during the first 24 h of germination. After 48 h, the absence of PCNA is concomitant with an important decrease in DNA polymerase B activity. In this report we confirm the behaviour of DNA polymerase B as a delta-like activity.

Animals↗

Biological and chemical evaluation of chick pea seed proteins as affected by germination, extraction and alpha-amylase treatment.

The effects of germination, extraction (double extraction with 70% ethanol and water at isoelectric point) and alpha-amylase treatments of chick pea seed flours on crude protein, total carbohydrate, protein efficiency ratio (PER), biological value (BV), true digestibility (TD), net protein utilization (NPU), essential amino acid composition, in-vitro protein digestibility (IVPD) and actual amino acid indices (essential amino acid index or amino acid score) were evaluated. Crude protein content was increased (8-149%), while total carbohydrate was decreased (11-62%) by germination, extraction and alpha-amylase treatments. Alpha-amylase treatment was more efficient in reducing total carbohydrate and increasing the protein content than that of extraction treatment. The protein quality of chick pea flours as measured by PER, BV, TD, NPU, IVPD and corrected amino acid indices (actual amino acid indices x IVPD) was significantly improved by these treatments. The protein quality of germinated-alpha-amylase treatment was comparable with casein, while germinated-alpha-amylase treaded seeds appeared nutritionally superior to casein. The results indicate that the germinated-alpha-amylase and germinated-alpha-amylase-extracted treatments could be used successfully as a source of concentrated high quality protein for baby food production. The corrected amino acid indices gave better prediction of PER, BV, TD and NPU (r = 93 to 97) than actual amino acid indices (r = 45 to 71). PER was highly correlated with corrected amino acid score (r = 0.93). The PER could be predicted from the following simple regression equation: PER = -1.827 + 0.0561 x corrected amino acid score.

Dietary Proteins↗

Evolution and kinetics of monosaccharides, disaccharides and alpha-galactosides during germination of lentils.

The effect of light and seed rinsing during the germination of lentil seeds (Lens culinaris var. vulgaris, cultivar Magda-20) on the level of monosaccharides, disaccharides and alpha-galactosides (raffinose, ciceritol and stachyose) was investigated. The total soluble sugar content corresponded to about 9% of the mature seed weight, about 65% of which was alpha-galactosides. Germination brought about a large decrease in alpha-galactosides: 18% to 40% losses after 3 days and 100% after 6 days. However, glucose, which was not detected in ungerminated seeds, as well as fructose and sucrose gradually increased during germination. The content of alpha-galactosides decreased more rapidly when germinating seeds were given 6 h light per day, but under these conditions there was also a major reduction in the levels of fructose, glucose and sucrose. Seeds rinsed daily showed a greater reduction of alpha-galactosides and an increase in the levels of fructose, glucose and sucrose. When seeds were germinated for 10 days in the dark with daily rinsing, the content of alpha-galactosides decreased gradually during the first 4 days and they were not detected after 6 days. Under these conditions, fructose, glucose and sucrose, which represented about 3% of the mature seed weight, started increasing after day 2 and represented more than 13% of the germinated seed dry weight after 10 days.

Disaccharides↗

Distinct cis-acting elements direct the germination and sugar responses of the cucumber malate synthase gene.

The malate synthase gene (ms) promoter in cucumber (Cucumis sativus L.) was investigated with the aim of distinguishing DNA sequences mediating regulation of gene expression by sugar, and expression following seed germination. Promoter deletions were constructed and their ability to direct expression of the beta-glucuronidase (gus) reporter gene was investigated in transgenic Nicotiana plumbaginifolia. Gene expression was assayed in germinating seeds and developing seedlings (the germination response) and in seedlings transferred from light into darkness with and without sucrose (the sugar response). As progressively more of the promoter was deleted from the 5' end, first the sugar response and then the germination response was lost. Thus, distinct regions of the promoter are required for carbohydrate control and for regulation of gene expression in response to germination. Sequence comparisons of the ms promoter with that of the isocitrate lyase gene (icl) of cucumber have previously identified four IMH(ICL-MS-Homology) sequences. One such sequence, IMH2, is shown here to be implicated in the sugar response of the ms gene. The 17 bp sequences which when deleted from the ms gene results in loss of the germination response, contains a 14 bp sequence which is similar to a sequence in the icl promoter, which we refer to as IMH5. Furthermore, this sequence has similarity with amdI9-like sequences in filamentous fungi, which confer facB-mediated acetate inducibility on several genes, including those encoding ICL and MS.

Base Sequence↗

Glucose delays seed germination in Arabidopsis thaliana.

Here we report that glucose delays germination of Arabidopsis thaliana (L.) Heynh. seeds at concentrations below those known to inhibit early seedling development. This inhibition acts on embryo growth and is independent of hexokinase (HXK) function. Hormones and hormone inhibitors were applied to the germination media and several hormone biosynthesis and signalling mutants were tested on glucose media to investigate a possible role of abscisic acid (ABA), gibberellin and ethylene in the glucose-induced germination delay. Results indicate that the germination inhibition by glucose cannot be antagonized by ethylene or gibberellin and is independent of the HXK1/ABA/ ABI4 signalling cascade. These findings suggest that there is a separate regulatory pathway independent of ABI2/ ABI4/ ABI5. Thus, in a relatively short time frame sugars utilize different signalling cascades to inhibit germination and post-germination growth, underlining the complexity of sugar responses.

Abscisic Acid↗

Differentially expressed genes associated with dormancy or germination of Arabidopsis thaliana seeds.

Differential display analysis using dormant and non-dormant Arabidopsis thaliana (L.) Heynh seeds resulted in a set of genes that were associated with either dormancy or germination. Expression of the germination-associated genes AtRPL36B and AtRPL27B, encoding two ribosomal proteins, was undetectable in the dry seed, low in dormant seed, and high under conditions that allowed completion of germination. Expression of these genes was also found to be light-regulated and to correlate with germination speed. Expression of the dormancy-associated genes ATS2 and ATS4, encoding a caleosin-like protein and a protein similar to a low-temperature-induced protein respectively, was high in the dry seed and decreased during germination. Expression of ATS2 and ATS4 was high in primary and secondary dormant seed but low in after-ripened or chilled seed. The expression of both genes was also light-regulated, but no relationship with temperature-dependent germination speed was found.

Arabidopsis↗

Ethylene promotes ethylene biosynthesis during pea seed germination by positive feedback regulation of 1-aminocyclo-propane-1-carboxylic acid oxidase.

Increased ethylene evolution accompanies seed germination of many species including Pisum sativum L., but only a little is known about the regulation of the ethylene biosynthetic pathway in different seed tissues. Biosynthesis of the direct ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), the expression of ACC oxidase (ACO), and ethylene production were investigated in the cotyledons and embryonic axis of germinating pea seeds. An early onset and sequential induction of ACC biosynthesis, accumulation of Ps-ACO1 mRNA and of ACO activity, and ethylene production were localized almost exclusively in the embryonic axis. Maximal levels of ACC, Ps-ACO1 mRNA, ACO enzyme activity and ethylene evolution were found when radicle emergence was just complete. Treatment of germinating seeds with ethylene alone or in combination with the inhibitor of ethylene action 2,5-norbornadiene showed that endogenous ethylene regulates its own biosynthesis through a positive feedback loop that enhances ACO expression. Accumulation of Ps-ACO1 mRNA and of ACO enzyme activity in the embryonic axis during the late phase of germination required ethylene, whereas Ps-ACS1 mRNA levels and overall ACC contents were not induced by ethylene treatment. Ethylene did not induce ACO in the embryonic axis during the early phase of germination. Ethylene-independent signalling pathways regulate the spatial and temporal pattern of ethylene biosynthesis, whereas the ethylene signalling pathway regulates high-level ACO expression in the embryonic axis, and thereby enhances ethylene evolution during seed germination.

Amino Acid Oxidoreductases↗

Phytochrome A-mediated inhibition of seed germination in tomato.

Far-red light (FR) inhibition of seed germination of tomato (Solanum lycopersicum L.) was studied with the phytochrome (phy)-hypersensitive mutants, hp-1w, hp-1w,fri1, a phyA-deficient double mutant, and hp-1w,tri1, a phyB1-deficient double mutant. Seeds of all mutants germinated readily in the dark at 25 degrees C, and the germination was retarded by a single 100-s FR pulse given 1-3 h after sowing. The effect of an FR pulse was red-light reversible in all mutants used. After 24 h where a single FR pulse was no longer effective, prolonged FR exposure or hourly FR pulses suppressed germination in hp-1w and hp-1w,tri1, whereas in hp-1w,fri1 the suppressive effect of FR was almost absent. The effect of the prolonged FR was greater than that of the hourly 3-min FR pulses having equal photon fluence, and was fluencerate dependent. Thus we conclude that the germination inhibition by FR in tomato seed consists of a low-fluence response and a high irradiance response (HIR); the latter is controlled by phyA, but not phyB1. This is the first indication of phyA being involved in the HIR of seed germination inhibition.

Apoproteins↗

Behavior of storage lipids during development and germination of olive ( Olea europaea L.) pollen.

The presence of abundant oil bodies in the mature olive pollen grain has led us to focus on the behavior of these lipid bodies during pollen development and in vitro pollen germination. The appearance, increase, and accumulation of lipid bodies have been determined by following the sequential development of the pollen grain. Semithin slices of anthers and pollen grains were stained with Sudan Black B in order to identify neutral lipids. Ultrastructural studies were also carried out. Our results show a notable increase in lipid bodies between the young-pollen-grain stage and the mature-pollen-grain stage. Substantial polarization of lipid bodies was observed after 1 or 2 h of pollen incubation in germination medium. During pollen tube growth, the lipid bodies are located near the germinative aperture after 3 h of incubation, as well as inside the pollen tube, thus suggesting that the lipid bodies move from the pollen grain to the pollen tube. After 7 h of germination the presence of lipid bodies inside the pollen tube is no longer substantial. Our results support the idea that lipid bodies are involved in pollen germination, stigma penetration, and pollen tube growth. These results are discussed in connection with their implications for the pollen germination process.

Cell Polarity↗