Costas Fasseas

Polyphenol deposition in leaf hairs of Olea europaea (Oleaceae) and Quercus ilex (Fagaceae).

The subcellular localization (cytoplasm, vacuoles, cell walls) of polyphenol compounds during the development of the multicellular nonglandular leaf hairs of Olea europaea (scales) and Quercus ilex (stellates), was investigated. Hairs of all developmental stages were treated with specific inducers of polyphenol fluorescence, and the bright yellow-green fluorescence of individual hairs was monitored with epifluorescence microscopy. During the early ontogenetic stages, bright fluorescence was emitted from the cytoplasm of the cells composing the multicellular shield of the scales of O. europaea. Transmission electron micrographs of the same stages showed that these cells possessed poor vacuolation and thin cell walls. The nucleus of these cells may be protected against ultraviolet-B radiation damage. The progressive vacuolation that occurred during maturation was followed by a shifting of the bright green-yellow fluorescence from the perinuclear region and the cytoplasm to the cell walls. The same trends were observed during the development of the nonglandular stellate hairs of Quercus ilex, in which maturation was also accompanied by a considerable secondary thickening of the cell walls. Despite the differences in morphology, high concentrations of polyphenol compounds are initially located mainly in the cytoplasm of the developing nonglandular hairs, and their deposition on the cell walls takes place during the secondary cell wall thickening. These structural changes during the development of the leaf hairs make them a very effective barrier against abiotic (uv-B radiation) and probably biotic (pathogenic) stresses.

Wettability, Polarity, and Water Absorption of Holm Oak Leaves: Effect of Leaf Side and Age

The highly pubescent abaxial side of holm oak leaves is unwettable and water repellent, while the adaxial side is wettable and can take up water, which may be an adaptation to growing under Mediterranean conditions. Plant trichomes play important protective functions and may have a major influence on leaf surface wettability. With the aim of gaining insight into trichome structure, composition, and function in relation to water-plant surface interactions, we analyzed the adaxial and abaxial leaf surface of holm oak (Quercus ilex) as a model. By measuring the leaf water potential 24 h after the deposition of water drops onto abaxial and adaxial surfaces, evidence for water penetration through the upper leaf side was gained in young and mature leaves. The structure and chemical composition of the abaxial (always present) and adaxial (occurring only in young leaves) trichomes were analyzed by various microscopic and analytical procedures. The adaxial surfaces were wettable and had a high degree of water drop adhesion in contrast to the highly unwettable and water-repellent abaxial holm oak leaf sides. The surface free energy and solubility parameter decreased with leaf age, with higher values determined for the adaxial sides. All holm oak leaf trichomes were covered with a cuticle. The abaxial trichomes were composed of 8% soluble waxes, 49% cutin, and 43% polysaccharides. For the adaxial side, it is concluded that trichomes and the scars after trichome shedding contribute to water uptake, while the abaxial leaf side is highly hydrophobic due to its high degree of pubescence and different trichome structure, composition, and density. Results are interpreted in terms of water-plant surface interactions, plant surface physical chemistry, and plant ecophysiology.

Structural and cytochemical investigation of the leaf of silverleaf nightshade (Solanum elaeagnifolium), a drought-resistant alien weed of the Greek flora

Light and scanning electron microscopy as well as histochemical reactions were employed to investigate the leaf structure and secretory activity of an adventive perennial pest that is rapidly dispersing in the Mediterranean region. Interesting structural characteristics, not common among the well-adapted native Mediterranean phanerophytes, were observed. Although ‘unconventional’, these structural characteristics produce an ecologically equivalent, xeromorphic leaf on a plant that proves to be competitive and very efficient under the Mediterranean climatic conditions.

A Splicing Mutation in the Novel Mitochondrial Protein DNAJC11 Causes Motor Neuron Pathology Associated with Cristae Disorganization, and Lymphoid Abnormalities in Mice

Mitochondrial structure and function is emerging as a major contributor to neuromuscular disease, highlighting the need for the complete elucidation of the underlying molecular and pathophysiological mechanisms. Following a forward genetics approach with N-ethyl-N-nitrosourea (ENU)-mediated random mutagenesis, we identified a novel mouse model of autosomal recessive neuromuscular disease caused by a splice-site hypomorphic mutation in a novel gene of unknown function, DnaJC11. Recent findings have demonstrated that DNAJC11 protein co-immunoprecipitates with proteins of the mitochondrial contact site (MICOS) complex involved in the formation of mitochondrial cristae and cristae junctions. Homozygous mutant mice developed locomotion defects, muscle weakness, spasticity, limb tremor, leucopenia, thymic and splenic hypoplasia, general wasting and early lethality. Neuropathological analysis showed severe vacuolation of the motor neurons in the spinal cord, originating from dilatations of the endoplasmic reticulum and notably from mitochondria that had lost their proper inner membrane organization. The causal role of the identified mutation in DnaJC11 was verified in rescue experiments by overexpressing the human ortholog. The full length 63 kDa isoform of human DNAJC11 was shown to localize in the periphery of the mitochondrial outer membrane whereas putative additional isoforms displayed differential submitochondrial localization. Moreover, we showed that DNAJC11 is assembled in a high molecular weight complex, similarly to mitofilin and that downregulation of mitofilin or SAM50 affected the levels of DNAJC11 in HeLa cells. Our findings provide the first mouse mutant for a putative MICOS protein and establish a link between DNAJC11 and neuromuscular diseases.

Effects of Lactobacillus salivarius, Lactobacillus reuteri, and Pediococcus acidilactici on the nematode Caenorhabditis elegans include possible antitumor activity

This study examined the effects of three lactic acid bacteria (LAB) strains on the nematode Caenorhabditis elegans. Lactobacillus salivarius, Lactobacillus reuteri, and Pediococcus acidilactici were found to inhibit the development and growth of the worm. Compared to Escherichia coli used as the control, L. reuteri and P. acidilactici reduced the lifespan of wild-type and short-lived daf-16 worms. On the contrary, L. salivarius extended the lifespan of daf-16 worms when used live, but reduced it as UV-killed bacteria. The three LAB induced the expression of genes involved in pathogen response and inhibited the growth of tumor-like germ cells, without affecting DAF16 localization or increasing corpse cells. Our results suggest the possible use of C. elegans as a model for studying the antitumor attributes of LAB. The negative effects of these LAB strains on the nematode also indicate their potential use against parasitic nematodes.

Alarm Photosynthesis: Calcium Oxalate Crystals as an Internal CO2 Source in Plants

A new photosynthetic path named “alarm photosynthesis” uses mesophyll calcium oxalate crystals as the CO2 source when stomata are closed, providing adaptive advantages under drought conditions. Calcium oxalate crystals are widespread among animals and plants. In land plants, crystals often reach high amounts, up to 80% of dry biomass. They are formed within specific cells, and their accumulation constitutes a normal activity rather than a pathological symptom, as occurs in animals. Despite their ubiquity, our knowledge on the formation and the possible role(s) of these crystals remains limited. We show that the mesophyll crystals of pigweed (Amaranthus hybridus) exhibit diurnal volume changes with a gradual decrease during daytime and a total recovery during the night. Moreover, stable carbon isotope composition indicated that crystals are of nonatmospheric origin. Stomatal closure (under drought conditions or exogenous application of abscisic acid) was accompanied by crystal decomposition and by increased activity of oxalate oxidase that converts oxalate into CO2. Similar results were also observed under drought stress in Dianthus chinensis, Pelargonium peltatum, and Portulacaria afra. Moreover, in A. hybridus, despite closed stomata, the leaf metabolic profiles combined with chlorophyll fluorescence measurements indicated active photosynthetic metabolism. In combination, calcium oxalate crystals in leaves can act as a biochemical reservoir that collects nonatmospheric carbon, mainly during the night. During the day, crystal degradation provides subsidiary carbon for photosynthetic assimilation, especially under drought conditions. This new photosynthetic path, with the suggested name “alarm photosynthesis,” seems to provide a number of adaptive advantages, such as water economy, limitation of carbon losses to the atmosphere, and a lower risk of photoinhibition, roles that justify its vast presence in plants.

Leaf anatomy and histochemistry of Lippia citriodora (Verbenaceae).

Lippia citriodora H.B.K. is an aromatic plant indigenous to South America. It is cultivated and commercialised as an ornamental for its lemon-like scent emitted from its leaves and flowers. The present morphoanatomical and histochemical study revealed that leaves of L. citriodora possess one type of setae (non-glandular) and at least five types of glandular trichomes, with the latter differing anatomically and in the composition of their secondary metabolites. Scanning electron microscopy revealed that the same types of trichomes exist also on the calyces. Histochemical tests indicate that the glandular trichomes contain a combination of terpenoids, flavonoids, carbohydrates, phenolics and alkaloids. However, within the vacuoles of the mesophyll cells, terpenoids, phenolics, flavonoids and tannins are stored. The presence of lamellar cytoplasmic inclusions in the mesophyll cells, possibly precursors of tannins stored in the vacuoles, is reported for the first time.

Characterization of a Mesorhizobium loti α-Type Carbonic Anhydrase and Its Role in Symbiotic Nitrogen Fixation

Carbonic anhydrase (CA) (EC 4.2.1.1) is a widespread enzyme catalyzing the reversible hydration of CO(2) to bicarbonate, a reaction that participates in many biochemical and physiological processes. Mesorhizobium loti, the microsymbiont of the model legume Lotus japonicus, possesses on the symbiosis island a gene (msi040) encoding an alpha-type CA homologue, annotated as CAA1. In the present work, the CAA1 open reading frame from M. loti strain R7A was cloned, expressed, and biochemically characterized, and it was proven to be an active alpha-CA. The biochemical and physiological roles of the CAA1 gene in free-living and symbiotic rhizobia were examined by using an M. loti R7A disruption mutant strain. Our analysis revealed that CAA1 is expressed in both nitrogen-fixing bacteroids and free-living bacteria during growth in batch cultures, where gene expression was induced by increased medium pH. L. japonicus plants inoculated with the CAA1 mutant strain showed no differences in top-plant traits and nutritional status but consistently formed a higher number of nodules exhibiting higher fresh weight, N content, nitrogenase activity, and delta(13)C abundance. Based on these results, we propose that although CAA1 is not essential for nodule development and symbiotic nitrogen fixation, it may participate in an auxiliary mechanism that buffers the bacteroid periplasm, creating an environment favorable for NH(3) protonation, thus facilitating its diffusion and transport to the plant. In addition, changes in the nodule delta(13)C abundance suggest the recycling of at least part of the HCO(3)(-) produced by CAA1.

Factors affecting the efficacy of agar-based substrates for the study of tomato pollen germination

Summary The present work examines the effects of substrate additions and incubation conditions on in vitro germination of tomato pollen on semi-solid, agar-based substrates with the ultimate aim of defining a standard semi-solid substrate for tomato pollen germination studies. Partial replacement of sucrose by polyethylene glycol (PEG-6000) for osmotic regulation of the substrate significantly increased pollen germination of both the low temperature-susceptible F1 cultivar ‘Dombito’, and the more temperature-resistant cv.‘Supermarmande’, to a level that varied according to the season of pollen harvest. In contrast, partial substitution of sucrose by mannitol was inhibitory to an extent that depended on the final concentration of mannitol in the medium. The optimum pH for germination was 6.5 and the optimum incubation temperature was 15°C. Among the vitamins (riboflavin, thiamine, pyridoxine, niacin, pantothenic acid, p-aminobenzoic acid), amino acids (glutamic acid, aspartic acid), casein hydrolysate, plant growth regulators (gibberellic acid, indole-3-acetic acid, indole-3-butyric acid, indole-3-propionic acid, 1-naphthaleneacetic acid, kinetin) and flavonoids (naringin, myricetin, naringenin, quercetin, rutin) tested, a significant increase in pollen germination was induced only by the flavonoids quercetin and myricetin and, to a lesser extent, by indole-3-acetic acid. In contrast, both kinetin and naringenin were inhibitory, as were both amino acids at 100 mg l–1 and casein hydrolysate (at 1.0 – 10.0 g l–1). On the basis of the above results, we conclude that, for both tomato cultivars tested, the most suitable semi-solid substrate among those examined was that containing 10% (w/v) sucrose, 15.1% (w/v) PEG-6000 and 1.5% (w/v) agar, with the possible addition of 5 mg l–1 quercetin, or 5 mg l–1 myricetin.

Pollen polymorphism in Thymus capitatus (Lamiaceae)

Polymorphism with respect to size and structural characteristics occurs in pollen grains of several species. Pollen grains of Thymus capitatus (L.) Hoffmans & Link. from three different areas of Greece were collected, and their palynological characteristics were measured. Using light and scanning electron microscopy, a quantification of pollen polymorphism of T. capitatus was achieved by measuring the size and the characteristics of exine sculpture. The measurements were repeated during 3 successive years, and statistical analyses with one-way ANOVA and Duncan’s multiple range test were applied. Results revealed differences in the exine pattern between the locations; also, three palynological characteristics (surface, perimeter, and equatorial diameter) differed statistically. The results clearly indicate that pollen grains of T. capitatus are polymorphic between the sampling areas. The polymorphism has been related to environmental parameters and mainly with water availability. Average rainfall seems to ...