Pollen germination as a crucial process in plant development strongly depends on the accessibility of carbon as energy source. Carbohydrates, however, function not only as a primary energy source, but also as important signaling components. In a comprehensive study, we analyzed various aspects of the impact of 32 different sugars on in vitro germination of Arabidopsis pollen comprising about 150 variations of individual sugars and combinations. Twenty-six structurally different mono-, di- and oligosaccharides, and sugar analogs were initially tested for their ability to support pollen germination. Whereas several di- and oligosaccharides supported pollen germination, hexoses such as glucose, fructose and mannose did not support and even considerably inhibited pollen germination when added to germination-supporting medium. Complementary experiments using glucose analogs with varying functional features, the hexokinase inhibitor mannoheptulose and the glucose-insensitive hexokinase-deficient Arabidopsis mutant gin2-1 suggested that mannose- and glucose-mediated inhibition of sucrose-supported pollen germination depends partially on hexokinase signaling. The results suggest that, in addition to their role as energy source, sugars act as signaling molecules differentially regulating the complex process of pollen germination depending on their structural properties. Thus, a sugar-dependent multilayer regulation of Arabidopsis pollen germination is supported, which makes this approach a valuable experimental system for future studies addressing sugar sensing and signaling.

• MeSH
• Arabidopsis účinky léků   fyziologie   MeSH
• hexosy metabolismus   farmakologie   MeSH
• klíčení účinky léků   fyziologie   MeSH
• mannosa metabolismus   farmakologie   MeSH
• metabolismus sacharidů * MeSH
• oligosacharidy chemie   metabolismus   farmakologie   MeSH
• pyl metabolismus   fyziologie   MeSH
• sacharidy MeSH
• sacharosa metabolismus   farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH

Plant-derived smoke stimulates seed germination in numerous plant species. Smoke also has a positive stimulatory effect on pollen germination and pollen tube growth. The range of plant families affected my smoke still needs to be established since the initial study was restricted to only three species from the Amaryllidaceae. The effects of smoke-water (SW) and the smoke-derived compounds, karrikinolide (KAR1 ) and trimethylbutenolide (TMB) on pollen growth characteristics were evaluated in seven different plant families. Smoke-water (1:1000 and 1:2000 v:v) combined with either Brewbaker and Kwack's (BWK) medium or sucrose and boric acid (SB) medium significantly improved pollen germination and pollen tube growth in Aloe maculata All., Kniphofia uvaria Oken, Lachenalia aloides (L.f.) Engl. var. aloides and Tulbaghia simmleri P. Beauv. Karrikinolide (10(-6) and 10(-7) m) treatment significantly improved pollen tube growth in A. maculata, K. uvaria, L. aloides and Nematanthus crassifolius (Schott) Wiehle compared to the controls. BWK or SB medium containing TMB (10(-3) m) produced significantly longer pollen tubes in A. maculata, K. uvaria and N. crassifolius. These results indicate that plant-derived smoke and the smoke-isolated compounds may stimulate pollen growth in a wide range of plant species.

• MeSH
• furany farmakologie   MeSH
• gama-butyrolakton analogy a deriváty   farmakologie   MeSH
• kouř MeSH
• kultivační média MeSH
• liliovité účinky léků   MeSH
• pyl účinky léků   růst a vývoj   MeSH
• pylová láčka účinky léků   růst a vývoj   MeSH
• pyrany farmakologie   MeSH
• voda MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Plant-derived smoke and smoke-isolated compounds stimulate germination in seeds from over 80 genera. It has also been reported that smoke affects overall plant vigour and has a stimulatory effect on pollen growth. The effect of smoke on orchid seeds, however, has not been assessed. In South Africa, orchid seeds from several genera may be exposed to smoke when they are released from their seedpods. It is therefore possible that smoke may affect their germination and growth. Therefore, the effects of smoke [applied as smoke-water (SW)] and two smoke-derived compounds, karrikinolide (KAR1 ) and trimethylbutenolide (TMB), were investigated on the germination and growth of orchid seeds in vitro. The effect of SW, KAR1 and TMB were investigated on the endangered epiphytic orchid, Ansellia africana, which is indigenous to tropical areas of Africa. Smoke-water, KAR1 and TMB were infused in half-strength MS medium. The number of germinated seeds and number of seeds and protocorm bodies to reach predetermined developmental stages were recorded on a weekly basis using a dissecting microscope for a 13-week period. Infusing SW 1:250 (v:v) into half-strength MS medium significantly increased the germination rate index (GRI) and the development rate index (DRI) of the A. africana seeds. All the SW treatments significantly increased the number of large protocorm bodies at the final stage of development. Infusing KAR1 into the growing medium had no significant effect on germination or development of the seeds. The TMB treatment, however, significantly reduced the GRI and DRI of A. africana seeds.

• MeSH
• klíčení * MeSH
• kouř * MeSH
• květy fyziologie   MeSH
• Orchidaceae růst a vývoj   MeSH
• semena rostlinná růst a vývoj   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Polarized exocytosis is critical for pollen tube growth, but its localization and function are still under debate. The exocyst vesicle-tethering complex functions in polarized exocytosis. Here, we show that a sec3a exocyst subunit null mutant cannot be transmitted through the male gametophyte due to a defect in pollen tube growth. The green fluorescent protein (GFP)-SEC3a fusion protein is functional and accumulates at or proximal to the pollen tube tip plasma membrane. Partial complementation of sec3a resulted in the development of pollen with multiple tips, indicating that SEC3 is required to determine the site of pollen germination pore formation. Time-lapse imaging demonstrated that SEC3a and SEC8 were highly dynamic and that SEC3a localization on the apical plasma membrane predicts the direction of growth. At the tip, polar SEC3a domains coincided with cell wall deposition. Labeling of GFP-SEC3a-expressing pollen with the endocytic marker FM4-64 revealed the presence of subdomains on the apical membrane characterized by extensive exocytosis. In steady-state growing tobacco (Nicotiana tabacum) pollen tubes, SEC3a displayed amino-terminal Pleckstrin homology-like domain (SEC3a-N)-dependent subapical membrane localization. In agreement, SEC3a-N interacted with phosphoinositides in vitro and colocalized with a phosphatidylinositol 4,5-bisphosphate (PIP2) marker in pollen tubes. Correspondingly, molecular dynamics simulations indicated that SEC3a-N associates with the membrane by interacting with PIP2 However, the interaction with PIP2 is not required for polar localization and the function of SEC3a in Arabidopsis (Arabidopsis thaliana). Taken together, our findings indicate that SEC3a is a critical determinant of polar exocytosis during tip growth and suggest differential regulation of the exocytotic machinery depending on pollen tube growth modes.

• MeSH
• Arabidopsis genetika   růst a vývoj   metabolismus   MeSH
• buněčná membrána metabolismus   MeSH
• časosběrné zobrazování metody   MeSH
• exocytóza * MeSH
• fosfatidylinositol-4,5-difosfát metabolismus   MeSH
• fosfatidylinositoly metabolismus   MeSH
• fylogeneze MeSH
• geneticky modifikované rostliny MeSH
• konfokální mikroskopie MeSH
• mutace MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• protein - isoformy genetika   metabolismus   MeSH
• proteiny huseníčku klasifikace   genetika   metabolismus   MeSH
• pyl genetika   růst a vývoj   metabolismus   MeSH
• pylová láčka genetika   růst a vývoj   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční homologie aminokyselin MeSH
• sekvenční homologie nukleových kyselin MeSH
• simulace molekulární dynamiky MeSH
• stanovení celkové genové exprese metody   MeSH
• vazba proteinů MeSH
• vazebná místa genetika   MeSH
• vezikulární transportní proteiny klasifikace   genetika   metabolismus   MeSH
• zelené fluorescenční proteiny genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH

Zm-p60.1 is maize cDNA coding cytokinin-glucoside specific beta-glucosidase. Indigogenic method was used for histochemical localization of Zm-p60.1 beta-glucosidase activity in various developmental stages of transgenic tobacco anthers. Expression of Zm-p60.1 cDNA in T7 tobacco plants is controlled by the CaMV 35S promoter. Another type of tobacco transformant expresses Zm-p60.1 under the control of LAT 52 promoter. Histochemical detection has proved different patterns of beta-glucosidase activity during tobacco pollen development in these two types of transformants. Zm-p60.1 beta-glucosidase activity had not direct influence on pollen germinability.

• MeSH
• beta-glukosidasa genetika   MeSH
• DNA primery genetika   MeSH
• exprese genu MeSH
• geneticky modifikované rostliny MeSH
• jedovaté rostliny MeSH
• kukuřice setá enzymologie   genetika   růst a vývoj   MeSH
• messenger RNA genetika   metabolismus   MeSH
• pyl enzymologie   genetika   růst a vývoj   MeSH
• RNA rostlin genetika   metabolismus   MeSH
• sekvence nukleotidů MeSH
• tabák MeSH
• transformace genetická MeSH

Developmental processes are closely connected to certain states of epigenetic information which, among others, rely on methylation of chromatin. S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) are key cofactors of enzymes catalyzing DNA and histone methylation. To study the consequences of altered SAH/SAM levels on plant development we applied 9-(S)-(2,3-dihydroxypropyl)-adenine (DHPA), an inhibitor of SAH-hydrolase, on tobacco seeds during a short phase of germination period (6 days). The transient drug treatment induced: (1) dosage-dependent global DNA hypomethylation mitotically transmitted to adult plants; (2) pleiotropic developmental defects including decreased apical dominance, altered leaf and flower symmetry, flower whorl malformations and reduced fertility; (3) dramatic upregulation of floral organ identity genes NTDEF, NTGLO and NAG1 in leaves. We conclude that temporal SAH-hydrolase inhibition deregulated floral genes expression probably via chromatin methylation changes. The data further show that plants might be particularly sensitive to accurate setting of SAH/SAM levels during critical developmental periods.

• MeSH
• adenin analogy a deriváty   toxicita   MeSH
• adenosylhomocysteinasa antagonisté a inhibitory   metabolismus   MeSH
• DNA primery genetika   MeSH
• epigeneze genetická účinky léků   fyziologie   MeSH
• klíčení účinky léků   fyziologie   MeSH
• komplementární DNA genetika   MeSH
• květy anatomie a histologie   fyziologie   MeSH
• metylace DNA MeSH
• neparametrická statistika MeSH
• pyl fyziologie   MeSH
• regulace genové exprese u rostlin účinky léků   genetika   fyziologie   MeSH
• rostlinné proteiny metabolismus   MeSH
• Southernův blotting MeSH
• tabák enzymologie   fyziologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

In our previous study we applied the Agilent 44K tobacco gene chip to introduce and analyze the tobacco male gametophyte transcriptome in mature pollen and 4h pollen tubes. Here we extended our analysis post-pollen mitosis II (PMII) by including a new data set obtained from more advanced stage of the ongoing progamic phase - pollen tubes cultivated in vitro for 24 h. Pollen mitosis II marks key events in the control of male gametophyte development, the production of two sperm cells. In bicellular species covering cca 70% of angiosperms including Nicotiana tabacum, PMII takes place after pollen germination in growing pollen tube. We showed the stable and even slightly increasing complexity of tobacco male gametophyte transcriptome over long period of progamic phase-24 h of pollen tube growth. We also demonstrated the ongoing transcription activity and specific transcript accumulation in post-PMII pollen tubes cultivated in vitro. In all, we have identified 320 genes (2.2%) that were newly transcribed at least after 4h of pollen tube cultivation in vitro. Further, 699 genes (4.8%) showed over 5-fold increased accumulation after the 24h of cultivation.

• MeSH
• analýza hlavních komponent MeSH
• mitóza genetika   MeSH
• pylová láčka cytologie   genetika   MeSH
• regulace genové exprese u rostlin MeSH
• tabák cytologie   genetika   MeSH
• transkriptom genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Many flowering plants produce bicellular pollen. The two cells of the pollen grain are destined for separate fates in the male gametophyte, which provides a unique opportunity to study genetic interactions that govern guided single-cell polar expansion of the growing pollen tube and the coordinated control of germ cell division and sperm cell fate specification. We applied the Agilent 44 K tobacco gene chip to conduct the first transcriptomic analysis of the tobacco male gametophyte. In addition, we performed a comparative study of the Arabidopsis root-hair trichoblast transcriptome to evaluate genetic factors and common pathways involved in polarized cell-tip expansion. RESULTS: Progression of pollen grains from freshly dehisced anthers to pollen tubes 4 h after germination is accompanied with > 5,161 (14.9%) gametophyte-specific expressed probes active in at least one of the developmental stages. In contrast, > 18,821 (54.4%) probes were preferentially expressed in the sporophyte. Our comparative approach identified a subset of 104 pollen tube-expressed genes that overlap with root-hair trichoblasts. Reverse genetic analysis of selected candidates demonstrated that Cu/Zn superoxide dismutase 1 (CSD1), a WD-40 containing protein (BP130384), and Replication factor C1 (NtRFC1) are among the central regulators of pollen-tube tip growth. Extension of our analysis beyond the second haploid mitosis enabled identification of an opposing-dynamic accumulation of core regulators of cell proliferation and cell fate determinants in accordance with the progression of the germ cell cycle. CONCLUSIONS: The current study provides a foundation to isolate conserved regulators of cell tip expansion and those that are unique for pollen tube growth to the female gametophyte. A transcriptomic data set is presented as a benchmark for future functional studies using developing pollen as a model. Our results demonstrated previously unknown functions of certain genes in pollen-tube tip growth. In addition, we highlighted the molecular dynamics of core cell-cycle regulators in the male gametophyte and postulated the first genetic model to account for the differential timing of spermatogenesis among angiosperms and its coordination with female gametogenesis.

• MeSH
• Arabidopsis genetika   MeSH
• buněčný cyklus genetika   MeSH
• gametogeneze rostlin MeSH
• genový knockdown MeSH
• klíčení MeSH
• kořeny rostlin genetika   MeSH
• pyl genetika   MeSH
• pylová láčka růst a vývoj   MeSH
• regulace genové exprese u rostlin MeSH
• RNA rostlin genetika   MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• tabák genetika   MeSH
• transkriptom MeSH
• vývojová regulace genové exprese MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

Heat shock transcription factors (Hsfs) are involved in multiple aspects of stress response and plant growth. However, their role during male gametophyte development is largely unknown, although the generative phase is the most sensitive and critical period in the plant life cycle. Based on a wide screen of T-DNA mutant lines, we identified the atren1 mutation (restricted to nucleolus1) in early male gametophytic gene At1g77570, which has the closest homology to HSFA5 gene, the member of a heat shock transcription factor (HSF) gene family. The mutation causes multiple defects in male gametophyte development in both structure and function. Because the mutation disrupts an early acting (AtREN1) gene, these pollen phenotype abnormalities appear from bicellular pollen stage to pollen maturation. Moreover, the consequent progamic phase is compromised as well as documented by pollen germination defects and limited transmission via male gametophyte. In addition, atren1/- plants are defective in heat stress (HS) response and produce notably higher proportion of aberrant pollen grains. AtREN1 protein is targeted specifically to the nucleolus that, together with the increased size of the nucleolus in atren1 pollen, suggests that it is likely to be involved in ribosomal RNA biogenesis or other nucleolar functions.

• MeSH
• alely MeSH
• Arabidopsis cytologie   růst a vývoj   metabolismus   MeSH
• buněčné jadérko metabolismus   MeSH
• DNA bakterií genetika   MeSH
• DNA vazebné proteiny genetika   metabolismus   MeSH
• exony genetika   MeSH
• fenotyp MeSH
• klíčení MeSH
• mutace genetika   MeSH
• penetrance MeSH
• proteiny huseníčku genetika   metabolismus   MeSH
• pyl cytologie   genetika   růst a vývoj   MeSH
• pylová láčka cytologie   genetika   růst a vývoj   MeSH
• reakce na tepelný šok * genetika   MeSH
• regulace genové exprese u rostlin MeSH
• segregace chromozomů genetika   MeSH
• testy genetické komplementace MeSH
• transport proteinů MeSH
• vývojová regulace genové exprese MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The transition between the quiescent mature and the metabolically active germinating pollen grain most probably involves changes in protein phosphorylation status, since phosphorylation has been implicated in the regulation of many cellular processes. Given that, only a minor proportion of cellular proteins are phosphorylated at any one time, and that phosphorylated and nonphosphorylated forms of many proteins can co-exist within a cell, the identification of phosphoproteins requires some prior enrichment from a crude protein extract. Here, we have used metal oxide/hydroxide affinity chromatography (MOAC) based on an aluminum hydroxide matrix for this purpose, and have generated a population of phosphoprotein candidates from both mature and in vitro activated tobacco pollen grains. Both electrophoretic and nonelectrophoretic methods, allied to MS, were applied to these extracts to identify a set of 139 phosphoprotein candidates. In vitro phosphorylation was also used to validate the spectrum of phosphoprotein candidates obtained by the MOAC phosphoprotein enrichment. Since only one phosphorylation site was detected by the above approach, titanium dioxide phosphopeptide enrichment of trypsinized mature pollen crude extract was performed as well. It resulted in a detection of additional 51 phosphorylation sites giving a total of 52 identified phosphosites in this set of 139 phosphoprotein candidates.

• MeSH
• 2D gelová elektroforéza MeSH
• chromatografie afinitní metody   MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• fosfoproteiny analýza   chemie   izolace a purifikace   MeSH
• fosforylace MeSH
• molekulární sekvence - údaje MeSH
• proteom analýza   chemie   MeSH
• pyl chemie   MeSH
• rostlinné proteiny analýza   chemie   MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• tabák chemie   MeSH
• titan MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH