Individual architecture and photosynthetic performance of the submerged form of Drosera intermedia Hayne.

Drosera intermedia grows in acidic bogs in parts of valleys that are flooded in winter, and that often dry out in summer. It is also described as the sundew of the most heavily hydrated habitats in peatlands, and it is often found in water and even underwater. This sundew is the only one that can tolerate long periods of submersion, and more importantly produces a typical submerged form that can live in such conditions for many years. Submerged habitats are occupied by D. intermedia relatively frequently. The aim of the study was to determine the environmental conditions and architecture of individuals in the submerged form of D. intermedia. The features of the morphological and anatomical structure and chlorophyll a fluorescence of this form that were measured were compared with analogous ones in individuals that occurred in emerged and peatland habitats. The submerged form occurred to a depth of 20 cm. Compared to the other forms, its habitat had the highest pH (4.71-4.92; Me = 4.71), the highest temperature and substrate hydration, and above all, the lowest photosynthetically active radiation (PAR; 20.4-59.4%). This form differed from the other forms in almost all of the features of the plant's architecture. It is particularly noteworthy that it had the largest main axis height among all of the forms, which exceeded 18 cm. The number of living leaves in a rosette was notable (18.1 ± 8.1), while the number of dead leaves was very low (6.9 ± 3.8). The most significant differences were in the shape of its submerged leaves, in which the length of the leaf blade was the lowest of all of the forms (0.493 ± 0.15 mm; p < 0.001) and usually the widest. The stem cross-sectional area was noticeably smaller in the submerged form than in the other forms, the xylem was less developed and collaterally closed vascular bundles occurred. Our analysis of the parameters of chlorophyll fluorescence in vivo revealed that the maximum quantum yield of the primary photochemistry of photosystem II is the highest for the submerged form (Me = 0.681), the same as the maximum quantum yield of the electron transport (Me φE0 = 0.183). The efficiency of energy use per one active reaction center of photosystem II (RC) was the lowest in the submerged form (Me = 2.978), same as the fraction of energy trapped by one active RC (Me = 1.976) and the non-photochemical energy dissipation (DI0/RC; Me = 0.916). The ET0/RC parameter, associated with the efficiency of the energy utilization for electron transport by one RC, in the submerged plant reached the highest value (Me = 0.489). The submerged form of D. intermedia clearly differed from the emerged and peatland forms in its plant architecture. The submerged plants had a thinner leaf blade and less developed xylem than the other forms, however, their stems were much longer. The relatively high photosynthetic efficiency of the submerged forms suggests that most of the trapped energy is utilized to drive photosynthesis with a minimum energy loss, which may be a mechanism to compensate for the relatively small size of the leaf blade.

Do Cuticular Gaps Make It Possible to Study the Composition of the Cell Walls in the Glands of Drosophyllum lusitanicum?

Carnivorous plants can survive in poor habitats because they have the ability to attract, capture, and digest prey and absorb animal nutrients using modified organs that are equipped with glands. These glands have terminal cells with permeable cuticles. Cuticular discontinuities allow both secretion and endocytosis. In Drosophyllum lusitanicum, these emergences have glandular cells with cuticular discontinuities in the form of cuticular gaps. In this study, we determined whether these specific cuticular discontinuities were permeable enough to antibodies to show the occurrence of the cell wall polymers in the glands. Scanning transmission electron microscopy was used to show the structure of the cuticle. Fluorescence microscopy revealed the localization of the carbohydrate epitopes that are associated with the major cell wall polysaccharides and glycoproteins. We showed that Drosophyllum leaf epidermal cells have a continuous and well-developed cuticle, which helps the plant inhibit water loss and live in a dry environment. The cuticular gaps only partially allow us to study the composition of cell walls in the glands of Drosophyllum. We recoded arabinogalactan proteins, some homogalacturonans, and hemicelluloses. However, antibody penetration was only limited to the cell wall surface. The localization of the wall components in the cell wall ingrowths was missing. The use of enzymatic digestion improves the labeling of hemicelluloses in Drosophyllum glands.

Differences in the Occurrence of Cell Wall Components between Distinct Cell Types in Glands of Drosophyllum lusitanicum.

Carnivorous plants are mixotrophs that have developed the ability to lure, trap, and digest small organisms and utilize components of the digested bodies. Leaves of Drosophyllum lusitanicum have two kinds of glands (emergences): stalked mucilage glands and sessile digestive glands. The stalked mucilage glands perform the primary role in prey lure and trapping. Apart from their role in carnivory, they absorb water condensed from oceanic fog; thus, plants can survive in arid conditions. To better understand the function of carnivorous plant emergences, the molecular composition of their cell walls was investigated using immunocytochemical methods. In this research, Drosophyllum lusitanicum was used as a study system to determine whether cell wall immunocytochemistry differs between the mucilage and digestive glands of other carnivorous plant species. Light and electron microscopy were used to observe gland structure. Fluorescence microscopy revealed the localization of carbohydrate epitopes associated with the major cell wall polysaccharides and glycoproteins. The mucilage gland (emergence) consists of a glandular head, a connecting neck zone, and stalk. The gland head is formed by an outer and inner layer of glandular (secretory) cells and supported by a layer of endodermoid (barrier) cells. The endodermoid cells have contact with a core of spongy tracheids with spiral-shaped thickenings. Lateral tracheids are surrounded by epidermal and parenchymal neck cells. Different patterns of cell wall components were found in the various cell types of the glands. Cell walls of glandular cells generally are poor in both low and highly esterified homogalacturonans (HGs) but enriched with hemicelluloses. Cell walls of inner glandular cells are especially rich in arabinogalactan proteins (AGPs). The cell wall ingrowths in glandular cells are significantly enriched with hemicelluloses and AGPs. In the case of cell wall components, the glandular cells of Drosophyllum lusitanicum mucilage glands are similar to the glandular cells of the digestive glands of Aldrovanda vesiculosa and Dionaea muscipula.

Blocking cholesterol formation and turnover improves cellular and mitochondria function in murine heart microvascular endothelial cells and cardiomyocytes.

Background: Statins and proprotein convertase subtilisin/kexin type 9 inhibitors (PCSK9i) are cornerstones of therapy to prevent cardiovascular disease, acting by lowering lipid concentrations and only partially identified pleiotropic effects. This study aimed to analyze impacts of atorvastatin and synthetic peptide PCSK9i on bioenergetics and function of microvascular endothelial cells and cardiomyocytes. Methods: Mitochondrial function and abundance as well as intracellular nucleotides, membrane potential, cytoskeleton structure, and cell proliferation rate were evaluated in mouse heart microvascular endothelial cells (H5V) and cardiomyocytes (HL-1) under normal and hypoxia-mimicking conditions (CoCl2 exposure). Results: In normal conditions PCSK9i, unlike atorvastatin, enhanced mitochondrial respiratory parameters, increased nucleotide levels, prevented actin cytoskeleton disturbances and stimulated endothelial cell proliferation. Under hypoxia-mimicking conditions both atorvastatin and PCSK9i improved the mitochondrial respiration and membrane potential in both cell types. Conclusion: This study demonstrated that both treatments benefited the endothelial cell and cardiomyocyte bioenergetics, but the effects of PCSK9i were superior.

Immunocytochemical Analysis of Bifid Trichomes in Aldrovanda vesiculosa L. Traps.

The two-armed bifids (bifid trichomes) occur on the external (abaxial) trap surface, petiole, and stem of the aquatic carnivorous plant Aldrovanda vesiculosa (Droseracee). These trichomes play the role of mucilage trichomes. This study aimed to fill the gap in the literature concerning the immunocytochemistry of the bifid trichomes and compare them with digestive trichomes. Light and electron microscopy was used to show the trichome structure. Fluorescence microscopy revealed the localization of carbohydrate epitopes associated with the major cell wall polysaccharides and glycoproteins. The stalk cells and the basal cells of the trichomes were differentiated as endodermal cells. Cell wall ingrowths occurred in all cell types of the bifid trichomes. Trichome cells differed in the composition of their cell walls. The cell walls of the head cells and stalk cells were enriched with arabinogalactan proteins (AGPs); however, they were generally poor in both low- and highly-esterified homogalacturonans (HGs). The cell walls in the trichome cells were rich in hemicelluloses: xyloglucan and galactoxyloglucan. The cell wall ingrowths in the basal cells were significantly enriched with hemicelluloses. The presence of endodermal cells and transfer cells supports the idea that bifid trichomes actively transport solutes, which are polysaccharide in nature. The presence of AGPs (which are considered plant signaling molecules) in the cell walls in these trichome cells indicates the active and important role of these trichomes in plant function. Future research should focus on the question of how the molecular architecture of trap cell walls changes in cells during trap development and prey capture and digestion in A. vesiculosa and other carnivorous plants.

Screening for Cyclotides in Sri Lankan Medicinal Plants: Discovery, Characterization, and Bioactivity Screening of Cyclotides from Geophila repens.

Cyclotides are an intriguing class of structurally stable circular miniproteins of plant origin with numerous potential pharmaceutical and agricultural applications. To investigate the occurrence of cyclotides in Sri Lankan flora, 50 medicinal plants were screened, leading to the identification of a suite of new cyclotides from Geophila repens of the family Rubiaceae. Cycloviolacin O2-like (cyO2-like) gere 1 and the known cyclotide kalata B7 (kB7) were among the cyclotides characterized at the peptide and/or transcript level together with several putative enzymes, likely involved in cyclotide biosynthesis. Five of the most abundant cyclotides were isolated, sequenced, structurally characterized, and screened in antimicrobial and cytotoxicity assays. All gere cyclotides showed cytotoxicity (IC50 of 2.0-10.2 µM), but only gere 1 inhibited standard microbial strains at a minimum inhibitory concentration of 4-16 µM. As shown by immunohistochemistry, large quantities of the cyclotides were localized in the epidermis of the leaves and petioles of G. repens. Taken together with the cytotoxicity and membrane permeabilizing activities, this implicates gere cyclotides as potential plant defense molecules. The presence of cyO2-like gere 1 in a plant in the Rubiaceae supports the notion that phylogenetically distant plants may have coevolved to express similar cytotoxic cyclotides for a specific functional role, most likely involving host defense.

The Localization of Cell Wall Components in the Quadrifids of Whole-Mount Immunolabeled Utricularia dichotoma Traps.

Utricularia (bladderworts) are carnivorous plants. They produce small hollow vesicles, which function as suction traps that work underwater and capture fine organisms. Inside the traps, there are numerous glandular trichomes (quadrifids), which take part in the secretion of digestive enzymes, the resorption of released nutrients, and likely the pumping out of water. Due to the extreme specialization of quadrifids, they are an interesting model for studying the cell walls. This aim of the study was to fill in the gap in the literature concerning the immunocytochemistry of quadrifids in the major cell wall polysaccharides and glycoproteins. To do this, the localization of the cell wall components in the quadrifids was performed using whole-mount immunolabeled Utricularia traps. It was observed that only parts (arms) of the terminal cells had enough discontinuous cuticle to be permeable to antibodies. There were different patterns of the cell wall components in the arms of the terminal cells of the quadrifids. The cell walls of the arms were especially rich in low-methyl-esterified homogalacturonan. Moreover, various arabinogalactan proteins also occurred. Cell walls in glandular cells of quadrifids were rich in low-methyl-esterified homogalacturonan; in contrast, in the aquatic carnivorous plant Aldrovanda vesiculosa, cell walls in the glandular cells of digestive glands were poor in low-methyl-esterified homogalacturonan. Arabinogalactan proteins were found in the cell walls of trap gland cells in all studied carnivorous plants: Utricularia, and members of Droseraceae and Drosophyllaceae.

A Novel Cryptic Clostridial Peptide That Kills Bacteria by a Cell Membrane Permeabilization Mechanism.

This work reports detailed characteristics of the antimicrobial peptide Intestinalin (P30), which is derived from the LysC enzyme of Clostridium intestinale strain URNW. The peptide shows a broader antibacterial spectrum than the parental enzyme, showing potent antimicrobial activity against clinical strains of Gram-positive staphylococci and Gram-negative pathogens and causing between 3.04 ± 0.12 log kill for Pseudomonas aeruginosa PAO1 and 7.10 ± 0.05 log kill for multidrug-resistant Acinetobacter baumannii KPD 581 at a 5 µM concentration. Moreover, Intestinalin (P30) prevents biofilm formation and destroys 24-h and 72-h biofilms formed by Acinetobacter baumannii CRAB KPD 205 (reduction levels of 4.28 and 2.62 log CFU/mL, respectively). The activity of Intestinalin is combined with both no cytotoxicity and little hemolytic effect against mammalian cells. The nuclear magnetic resonance and molecular dynamics (MD) data show a high tendency of Intestinalin to interact with the bacterial phospholipid cell membrane. Although positively charged, Intestinalin resides in the membrane and aggregates into small oligomers. Negatively charged phospholipids stabilize peptide oligomers to form water- and ion-permeable pores, disrupting the integrity of bacterial cell membranes. Experimental data showed that Intestinalin interacts with negatively charged lipoteichoic acid (logK based on isothermal titration calorimetry, 7.45 ± 0.44), causes membrane depolarization, and affects membrane integrity by forming large pores, all of which result in loss of bacterial viability. IMPORTANCE Antibiotic resistance is rising rapidly among pathogenic bacteria, becoming a global public health problem that threatens the effectiveness of therapies for many infectious diseases. In this respect, antimicrobial peptides appear to be an interesting alternative to combat bacterial pathogens. Here, we report the characteristics of an antimicrobial peptide (of 30 amino acids) derived from the clostridial LysC enzyme. The peptide showed killing activity against clinical strains of Gram-positive and Gram-negative pathogens. Experimental data and computational modeling showed that this peptide forms transmembrane pores, directly engaging the negatively charged phospholipids of the bacterial cell membrane. Consequently, dissipation of the electrochemical gradient across cell membranes affects many vital processes, such as ATP synthesis, motility, and transport of nutrients. This kind of dysfunction leads to the loss of bacterial viability. Our firm conviction is that the presented study will be a helpful resource in searching for novel antimicrobial peptides that could have the potential to replace conventional antibiotics.

Immunocytochemical Analysis of the Wall Ingrowths in the Digestive Gland Transfer Cells in Aldrovanda vesiculosa L. (Droseraceae).

Carnivorous plants are unique due to their ability to attract small animals or protozoa, retain them in specialized traps, digest them, and absorb nutrients from the dissolved prey material; however, to this end, these plants need a special secretion-digestive system (glands). A common trait of the digestive glands of carnivorous plants is the presence of transfer cells. Using the aquatic carnivorous species Aldrovanda vesiculosa, we showed carnivorous plants as a model for studies of wall ingrowths/transfer cells. We addressed the following questions: Is the cell wall ingrowth composition the same between carnivorous plant glands and other plant system models? Is there a difference in the cell wall ingrowth composition between various types of gland cells (glandular versus endodermoid cells)? Fluorescence microscopy and immunogold electron microscopy were employed to localize carbohydrate epitopes associated with major cell wall polysaccharides and glycoproteins. The cell wall ingrowths were enriched with arabinogalactan proteins (AGPs) localized with the JIM8, JIM13, and JIM14 epitopes. Both methylesterified and de-esterified homogalacturonans (HGs) were absent or weakly present in the wall ingrowths in transfer cells (stalk cells and head cells of the gland). Both the cell walls and the cell wall ingrowths in the transfer cells were rich in hemicelluloses: xyloglucan (LM15) and galactoxyloglucan (LM25). There were differences in the composition between the cell wall ingrowths and the primary cell walls in A. vesiculosa secretory gland cells in the case of the absence or inaccessibility of pectins (JIM5, LM19, JIM7, LM5, LM6 epitopes); thus, the wall ingrowths are specific cell wall microdomains. Even in the same organ (gland), transfer cells may differ in the composition of the cell wall ingrowths (glandular versus endodermoid cells). We found both similarities and differences in the composition of the cell wall ingrowths between the A. vesiculosa transfer cells and transfer cells of other plant species.

Chromosome Number and Genome Size Evolution in Brasolia and Sobralia (Sobralieae, Orchidaceae).

Despite the clear circumscription of tribe Sobralieae (Orchidaceae), its internal relationships are still dubious. The recently delimited genus Brasolia, based on previous Sobralia species, is now assumed to be paraphyletic, with a third genus, Elleanthus, nested in it. The morphology of these three genera is significantly different, indicating the necessity of new data for a better genera delimitation. Though morphology and molecular data are available, cytogenetics data for Sobralieae is restricted to two Sobralia and one Elleanthus species. Aiming to evaluate the potential of cytogenetic data for Brasolia-Elleanthus-Sobralia genera delimitation, we present chromosome number and genome size data for 21 and 20 species, respectively, and used such data to infer the pattern of karyotype evolution in these genera. The analysis allowed us to infer x = 24 as the base chromosome number and genome size of average 1C-value of 5.0 pg for the common ancestor of Brasolia-Elleanthus-Sobralia. The recurrent descending dysploidy in Sobralieae and the punctual genome upsize suggest a recent diversification in Sobralieae but did not allow differing between Brasolia and Sobralia. However, the basal position of tribe Sobralieae in the subfamily Epidendroideae makes this tribe of interest to further studies clarifying the internal delimitation and pattern of karyotype evolution.

Jasmonate-Dependent Response of the Flower Abscission Zone Cells to Drought in Yellow Lupine.

Lipid membranes, as primary places of the perception of environmental stimuli, are a source of various oxygenated polyunsaturated fatty acids-oxylipins-functioning as modulators of many signal transduction pathways, e.g., phytohormonal. Among exogenous factors acting on plant cells, special attention is given to drought, especially in highly sensitive crop species, such as yellow lupine. Here, we used this species to analyze the contribution of lipid-related enzymes and lipid-derived plant hormones in drought-evoked events taking place in a specialized group of cells-the flower abscission zone (AZ)-which is responsible for organ detachment from the plant body. We revealed that water deficits in the soil causes lipid peroxidation in these cells and the upregulation of phospholipase D, lipoxygenase, and, concomitantly, jasmonic acid (JA) strongly accumulates in AZ tissue. Furthermore, we followed key steps in JA conjugation and signaling under stressful conditions by monitoring the level and tissue localization of enzyme providing JA derivatives (JASMONATE RESISTANT1) and the JA receptor (CORONATINE INSENSITIVE1). Collectively, drought-triggered AZ activation during the process of flower abscission is closely associated with the lipid modifications, leading to the formation of JA, its conjugation, and induction of signaling pathways.

The involvement of cyclotides in mutual interactions of violets and the two-spotted spider mite.

Plants employ different chemicals to protect themselves from herbivory. These defenses may be constitutive or triggered by stress. The chemicals can be toxic, act as repellents, phagosuppressants and/or phago-deterrents. The two-spotted spider mite (Tetranychus urticae) is a generalist arthropod herbivorous pest and its feeding causes extensive damage both to crops and wild plants. Cyclotides are cyclic peptides involved in host-plant defenses. A single Viola sp. can produce more than a hundred cyclotides with different biological activities and roles. The organ and tissue specific cyclotide patterns change over the seasons and/or with environment, but the role of biotic/abiotic stress in shaping them remains unclear. Here, we demonstrate the involvement of cyclotides in mutual interactions between violets and mites. We used immunohistochemistry and mass spectrometry imaging to show the ingested cyclotides in T. urticae and assess the Viola odorata response to mite feeding. Moreover, to assess how mites are affected by feeding on violets, acceptance and reproductive performance was compared between Viola uliginosa, V. odorata and Phaseolus vulgaris. We demonstrate that cyclotides had been taken in by mites feeding on the violets. The ingested peptides were found in contact with epithelial cells of the mite digestive system, in the fecal matter, feces, ovary and eggs. Mites preferred common bean plants (P. vulgaris) to any of the violet species; the latter affected their reproductive performance. The production of particular cyclotides in V. odorata (denoted by molecular weights: 2979, 3001, 3017, 3068, 3084, 3123) was activated by mite feeding and their levels were significantly elevated compared to the control after 5 and 21 days of infestation. Specific cyclotides may affect mites by being indigestible or through direct interaction with cells in the mite digestive tract and reproductive organs. A group of particular peptides in V. odorata appears to be involved in defense response against herbivores.

Remodeling of Cell Wall Components in Root Nodules and Flower Abscission Zone under Drought in Yellow Lupine.

We recently showed that yellow lupine is highly sensitive to soil water deficits since this stressor disrupts nodule structure and functioning, and at the same time triggers flower separation through abscission zone (AZ) activation in the upper part of the plant. Both processes require specific transformations including cell wall remodeling. However, knowledge about the involvement of particular cell wall elements in nodulation and abscission in agronomically important, nitrogen-fixing crops, especially under stressful conditions, is still scarce. Here, we used immuno-fluorescence techniques to visualize dynamic changes in cell wall compounds taking place in the root nodules and flower AZ of Lupinus luteus following drought. The reaction of nodules and the flower AZ to drought includes the upregulation of extensins, galactans, arabinans, xylogalacturonan, and xyloglucans. Additionally, modifications in the localization of high- and low-methylated homogalacturonans and arabinogalactan proteins were detected in nodules. Collectively, we determined for the first time the drought-associated modification of cell wall components responsible for their remodeling in root nodules and the flower AZ of L. luteus. The involvement of these particular molecules and their possible interaction in response to stress is also deeply discussed herein.

Spatiotemporal Distribution of Homogalacturonans and Hemicelluloses in the Placentas, Ovules and Female Gametophytes of Utricularia nelumbifolia during Pollination.

Utricularia nelumbifolia is a large carnivorous plant that is endemic to Brazil. It forms an extra-ovular female gametophyte, which surpasses the entire micropylar canal and extends beyond the limit of the integument and invades the placenta tissues. Due to the atypical behavior of the female gametophyte, it is interesting to determine the interaction between the gametophyte and sporophytic tissue. Therefore, the aim of this study was to evaluate the role of the placenta, the ovular tissues, the hypertrophied central cell and the integument in guiding the pollen tube in Utricularia nelumbifolia Gardner by studying the distribution of homogalacturonans and hemicelluloses. It was also determined whether the distribution of the homogalacturonans (HG) and hemicelluloses in Utricularia are dependent on pollination. The antibodies directed against the wall components (anti-pectin: JIM5, JIM7, LM19, LM20 and the anti-hemicelluloses: LM25, LM11, LM15, LM20, LM21) were used. Because both low- and high-esterified HG and xyloglucan were observed in the placenta, ovule (integument, chalaza) and female gametophyte of both pollinated and unpollinated flowers, the occurrence of these cell-wall components was not dependent on pollination. After fertilization, low methyl-esterified HGs were still observed in the cell walls of somatic cells and female gametophyte. However, in the case of high-esterified HG, the signal was weak and occurred only in the cell walls of the somatic cells. Because xyloglucans were observed in the cell walls of the synergids and egg cells, this suggests that they play a role in sexual reproduction. Utricularia nelumbifolia with an extra ovule-female gametophyte is presented as an attractive model for studying the male-female dialogue in plants.

Proteomic Studies of Roots in Hypoxia-Sensitive and -Tolerant Tomato Accessions Reveal Candidate Proteins Associated with Stress Priming.

Tomato (Solanum lycopersicum L.) is a vegetable frequently exposed to hypoxia stress induced either by being submerged, flooded or provided with limited oxygen in hydroponic cultivation systems. The purpose of the study was to establish the metabolic mechanisms responsible for overcoming hypoxia in two tomato accessions with different tolerance to this stress, selected based on morphological and physiological parameters. For this purpose, 3-week-old plants (plants at the juvenile stage) of waterlogging-tolerant (WL-T), i.e., POL 7/15, and waterlogging-sensitive (WL-S), i.e., PZ 215, accessions were exposed to hypoxia stress (waterlogging) for 7 days, then the plants were allowed to recover for 14 days, after which another 7 days of hypoxia treatment was applied. Root samples were collected at the end of each time-point and 2D-DIGE with MALDI TOF/TOF, and expression analyses of gene and protein-encoded alcohol dehydrogenase (ADH2) and immunolabelling of ADH were conducted. After collating the obtained results, the different responses to hypoxia stress in the selected tomato accessions were observed. Both the WL-S and WL-T tomato accessions revealed a high amount of ADH2, which indicates an intensive alcohol fermentation pathway during the first exposure to hypoxia. In comparison to the tolerant one, the expression of the adh2 gene was about two times higher for the sensitive tomato. Immunohistochemical analysis confirmed the presence of ADH in the parenchyma cells of the cortex and vascular tissue. During the second hypoxia stress, the sensitive accession showed a decreased accumulation of ADH protein and similar expression of the adh2 gene in comparison to the tolerant accession. Additionally, the proteome showed a greater protein abundance of glyceraldehyde-3-phosphate dehydrogenase in primed WL-S tomato. This could suggest that the sensitive tomato overcomes the oxygen limitation and adapts by reducing alcohol fermentation, which is toxic to plants because of the production of ethanol, and by enhancing glycolysis. Proteins detected in abundance in the sensitive accession are proposed as crucial factors for hypoxia stress priming and their function in hypoxia tolerance is discussed.

Arabinogalactan Proteins in the Digestive Glands of Dionaea muscipula J.Ellis Traps.

The arabinogalactan proteins (AGP) play important roles in plant growth and developmental processes. However, to the best of our knowledge, there is no information on the spatial distribution of AGP in the plant organs and tissues of carnivorous plants during their carnivorous cycle. The Dionaea muscipula trap forms an "external stomach" and is equipped with an effective digestive-absorbing system. Because its digestive glands are composed of specialized cells, the hypothesis that their cell walls are also very specialized in terms of their composition (AGP) compared to the cell wall of the trap epidermal and parenchyma cells was tested. Another aim of this study was to determine whether there is a spatio-temporal distribution of the AGP in the digestive glands during the secretory cycle of D. muscipula. Antibodies that act against AGPs, including JIM8, JIM13 and JIM14, were used. The localization of the examined compounds was determined using immunohistochemistry techniques and immunogold labeling. In both the un-fed and fed traps, there was an accumulation of AGP in the cell walls of the gland secretory cells. The epitope, which is recognized by JIM14, was a useful marker of the digestive glands. The secretory cells of the D. muscipula digestive glands are transfer cells and an accumulation of specific AGP was at the site where the cell wall labyrinth occurred. Immunogold labeling confirmed an occurrence of AGP in the cell wall ingrowths. There were differences in the AGP occurrence (labeled with JIM8 and JIM13) in the cell walls of the gland secretory cells between the unfed and fed traps.

Stellate Trichomes in Dionaea muscipula Ellis (Venus Flytrap) Traps, Structure and Functions.

The digestive organs of carnivorous plants have external (abaxial) glands and trichomes, which perform various functions. Dionaea muscipula Ellis (the Venus flytrap) is a model carnivorous plant species whose traps are covered by external trichomes. The aim of the study was to fill in the gap regarding the structure of the stellate outer trichomes and their immunocytochemistry and to determine whether these data support the suggestions of other authors about the roles of these trichomes. Light and electron microscopy was used to show the trichomes' structure. Fluorescence microscopy was used to locate the carbohydrate epitopes that are associated with the major cell wall polysaccharides and glycoproteins. The endodermal cells and internal head cells of the trichomes were differentiated as transfer cells, and this supports the idea that stellate trichomes transport solutes and are not only tomentose-like trichomes. Trichome cells differ in the composition of their cell walls, e.g., the cell walls of the internal head cells are enriched with arabinogalactan proteins (AGPs). The cell walls of the outer head cells are poor in both low and highly homogalacturonans (HGs), but the immature trichomes are rich in the pectic polysaccharide (1-4)-ß-D-galactan. In the immature traps, young stellate trichomes produce mucilage which may protect the trap surface, and in particular, the trap entrance. However, the role of these trichomes is different when the outer head cells collapse. In the internal head cells, a thick secondary wall cell was deposited, which together with the thick cell walls of the outer head cells played the role of a large apoplastic space. This may suggest that mature stellate trichomes might function as hydathodes, but this should be experimentally proven.

Spatio-Temporal Distribution of Cell Wall Components in the Placentas, Ovules and Female Gametophytes of Utricularia during Pollination.

In most angiosperms, the female gametophyte is hidden in the mother tissues and the pollen tube enters the ovule via a micropylar canal. The mother tissues play an essential role in the pollen tube guidance. However, in Utricularia, the female gametophyte surpasses the entire micropylar canal and extends beyond the limit of the integument. The female gametophyte then invades the placenta and a part of the central cell has direct contact with the ovary chamber. To date, information about the role of the placenta and integument in pollen tube guidance in Utricularia, which have extra-ovular female gametophytes, has been lacking. The aim of this study was to evaluate the role of the placenta, central cell and integument in pollen tube pollen tube guidance in Utricularia nelumbifolia Gardner and Utricularia humboldtii R.H. Schomb. by studying the production of arabinogalactan proteins. It was also determined whether the production of the arabinogalactan proteins is dependent on pollination in Utricularia. In both of the examined species, arabinogalactan proteins (AGPs) were observed in the placenta (epidermis and nutritive tissue), ovule (integument, chalaza), and female gametophyte of both pollinated and unpollinated flowers, which means that the production of AGPs is independent of pollination; however, the production of some AGPs was lower after fertilization. There were some differences in the production of AGPs between the examined species. The occurrence of AGPs in the placental epidermis and nutritive tissue suggests that they function as an obturator. The production of some AGPs in the ovular tissues (nucellus, integument) was independent of the presence of a mature embryo sac.

EPIP-Evoked Modifications of Redox, Lipid, and Pectin Homeostasis in the Abscission Zone of Lupine Flowers.

Yellow lupine is a great model for abscission-related research given that excessive flower abortion reduces its yield. It has been previously shown that the EPIP peptide, a fragment of LlIDL (INFLORESCENCE DEFICIENT IN ABSCISSION) amino-acid sequence, is a sufficient molecule to induce flower abortion, however, the question remains: What are the exact changes evoked by this peptide locally in abscission zone (AZ) cells? Therefore, we used EPIP peptide to monitor specific modifications accompanied by early steps of flower abscission directly in the AZ. EPIP stimulates the downstream elements of the pathway-HAESA and MITOGEN-ACTIVATED PROTEIN KINASE6 and induces cellular symptoms indicating AZ activation. The EPIP treatment disrupts redox homeostasis, involving the accumulation of H2O2 and upregulation of the enzymatic antioxidant system including superoxide dismutase, catalase, and ascorbate peroxidase. A weakening of the cell wall structure in response to EPIP is reflected by pectin demethylation, while a changing pattern of fatty acids and acyl lipids composition suggests a modification of lipid metabolism. Notably, the formation of a signaling molecule-phosphatidic acid is induced locally in EPIP-treated AZ. Collectively, all these changes indicate the switching of several metabolic and signaling pathways directly in the AZ in response to EPIP, which inevitably leads to flower abscission.

Immunodetection of Pectic Epitopes, Arabinogalactan Proteins, and Extensins in Mucilage Cells from the Ovules of Pilosella officinarum Vaill. and Taraxacum officinale Agg. (Asteraceae).

The main aim of this study was to compare the cytological difference between ovular mucilage cells in two Asteraceae species-Pilosella officinarum and Taraxacum officinale-in order to determine whether pectic epitopes, arabinogalactan proteins, or extensins are present. The immunocytochemical technique was used. Both the Taracacum and Pilosella genera have been used recently as models for understanding the mechanisms of apomixis. Knowledge of the presence of signal molecules (pectic epitopes, arabinogalactan proteins, and extensins) can help better understand the developmental processes in these plants during seed growth. The results showed that in Pilosella officinarum, there was an accumulation of pectins in the mucilage, including both weakly and highly esterified pectins, which was in contrast to the mucilage of Taraxacum officinale, which had low amounts of these pectins. However, Taraxacum protoplasts of mucilage cells were rich in weakly methyl-esterified pectins. While the mucilage contained arabinogalactan proteins in both of the studied species, the types of arabinogalactan proteins were different. In both of the studied species, extensins were recorded in the transmitting tissues. Arabinogalactan proteins as well as weakly and highly esterified pectins and extensins occurred in close proximity to calcium oxalate crystals in both Taraxacum and Pilosella cells.