Cell wall thickenings and tylosoid: developmental morphology reveals novelties for secretory canals in Protium ovatum (Burseraceae).

We investigated the structure, histochemistry, and ultrastructure of the secretory canals in the vegetative axis of Protium ovatum from a developmental perspective. Samples of roots, stems, and leaves were analyzed using light and transmission electron microscopy. Secretory canals composed of a uniseriate epithelium and a wide lumen occurred in the phloem of all analyzed organs. Schizogenesis and lysigenesis were merging processes involved in the origin, growth, ramification, and fusion of the secretory canals, forming an anastomosed secretory net. Essential oils, polysaccharides, and proteins were detected in the epithelial cells, as well as plastids with poorly developed thylakoids, dictyosomes, rough endoplasmic reticulum, polysomes, and oil drops, showing the mixed nature of the secretion. Epithelial cells exhibited pectin-cellulosic thickenings in the anticlinal and radial walls. These thickenings may act in directing the secretion flux toward the lumen, protecting the neighboring tissues from the toxicity of secreted metabolites. Structural irregularities observed in the mitochondria cristae in epithelial cells may be associated with processes induced by toxic substances. Epithelial cells protruded into the lumen and became lignified in the outer portion of the secondary phloem, obliterating the non-functional secretory canals. We propose that this phenomenon presents a physiological significance similar to that of tylose, preserving the secretion flow inside the active portions of the secretory system. To our knowledge, epithelial cells with wall thickenings, mitochondria with structural abnormalities, and obliteration of non-functional canals are features reported for the first time for Burseraceae. These features have important functional significance for Burseraceae secretory system and contribute to a deeper knowledge of P. ovatum, a medically and economically important plant.

Roles of Calcium Signaling in Gene Expression and Photosynthetic Acclimatization of Solanum lycopersicum Micro-Tom (MT) after Mechanical Damage.

A momentary increase in cytoplasmic Ca2+ generates an oscillation responsible for the activation of proteins, such as calmodulin and kinases, which interact with reactive oxygen species (ROS) for the transmission of a stress signal. This study investigated the influence of variations in calcium concentrations on plant defense signaling and photosynthetic acclimatization after mechanical damage. Solanum lycopersicum Micro-Tom was grown with 0, 2 and 4 mM Ca2+, with and without mechanical damage. The expression of stress genes was evaluated, along with levels of antioxidant enzymes, hydrogen peroxide, lipid peroxidation, histochemistry, photosynthesis and dry mass of organs. The ROS production generated by mechanical damage was further enhanced by calcium-free conditions due to the inactivation of the oxygen evolution complex, contributing to an increase in reactive species. The results indicated that ROS affected mechanical damage signaling because calcium-free plants exhibited high levels of H2O2 and enhanced expression of kinase and RBOH1 genes, necessary conditions for an efficient response to stress. We conclude that the plants without calcium supply recognized mechanical damage but did not survive. The highest expression of the RBOH1 gene and the accumulation of H2O2 in these plants signaled cell death. Plants grown in the presence of calcium showed higher expression of SlCaM2 and control of H2O2 concentration, thus overcoming the stress caused by mechanical damage, with photosynthetic acclimatization and without damage to dry mass production.

Apoplasmic barrier in the extrafloral nectary of Citharexylum myrianthum (Verbenaceae).

MAIN CONCLUSION: The cytological changes underlying the formation of an apoplasmic barrier in the multi-layered extrafloral nectaries of Citharexylum myrianthum are compatible with the synthesis, transport and deposition of suberin. In terms of ontogenesis and function, the intermediate layers of these nectaries are homologous with the stalks of nectar-secreting trichomes. Anticlinal cell wall impregnations are common in trichomatic nectaries and their functions as endodermis-like barriers have been discussed because of possible direct effects on the nectary physiology, mainly in the nectar secretion and resorption. However, the cytological events linked to nectary wall impregnations remain little explored. This study documents the ontogenesis and the fine structure of the EFN cells, and cytological events linked to the wall impregnations of multi-layered extrafloral nectaries (EFNs) in Citharexylum myrianthum Cham. (Verbenaceae). EFNs are patelliform, and differentiated into (a) a multicellular foot, which is compound in structure and vascularised with phloem strands, (b) a bi-layered intermediate region with thickened cell walls and (c) a single-layered secretory region with palisade-like cells. EFNs are protodermal in origin, starting with a single protodermal cell and ending with the complex, multi-layered structure. The cell wall impregnations first appear in the very young EFN and increase towards maturity. Lipid patches (assumed to be suberin) are deposited on the inner faces of the primary walls, first along the anticlinal walls and then extend to the periclinal walls. On both walls, plasmodesmata remain apparently intact during the maturation of the EFNs. In the peripheral cytoplasm there are abundant polymorphic plastids, well-developed Golgi bodies often close to rough endoplasmic reticulum profiles, mitochondria and polyribosomes. Cytological events linked to the wall impregnations are consistent with suberin synthesis, transport and deposition. Our findings offer new insights into the structure-properties of specialised nectary cell walls and so should contribute to our knowledge of the physiological and protective roles of this structure in nectar glands.

Fusoid cells in the grass family Poaceae (Poales): a developmental study reveals homologies and suggests new insights into their functional role in young leaves.

Background and Aims: In mature grass leaf blades as seen in cross-section, oblong cell-like structures have been interpreted most recently as intercellular gas spaces delimited by successive collapsed fusoid cells. These cells have been reported in at least seven of 12 subfamilies of Poaceae and are considered a synapomorphy for the family; however, no developmental work has been performed to verify their meristematic origin or to assess possible homologies within the graminid clade (= Flagellariaceae + [(Joinvilleaceae + Ecdeiocoleaceae) + Poaceae]) or among subfamilies of Poaceae. A developmental study was therefore carried out, including 20 species in three families (Flagellariaceae, Joinvilleaceae and Poaceae), representing the earlier-diverging and derived branches within the graminid clade and Poaceae. Methods: Light microscopy was combined with scanning electron microscopy, cryoscanning electron microscopy and transmission electron microscopy to study the development of leaves taken from the shoot apex of young plants. Mature leaf blades also were taken from living or dried plants and the mid-portion was studied. Key Results: Developmental results show that, in mature leaf blades as seen in cross-section, one apparent fusoid cell is typically a cavity resulting from the collapse of the initial fusoid cell and its internal divisions, which are herein interpreted as derivative cells with formation of cell plates only. Each cavity is delimited by successive collapsed fusoid cells arranged perpendicularly to the veins. Fusoid cells in all studied Poaceae members originate from the ground meristem, as do the colourless cells in Joinvillea ascendens (Joinvilleaceae). These two types of mesophyll cell have a strongly similar ontogeny, distinguished mainly by the collapse of the fusoid cells in Poaceae, which is not observed in the colourless cells in J. ascendens. Conclusions: Within the Poaceae, the meristematic origin of fusoid cells is the same in the early-diverging lineages, BOP clade and Panicoideae, and thus they are homologous within the family. The same topography and meristematic origin suggest that fusoid cells in Poaceae and colourless cells in Joinvilleaceae are homologous. The results also suggest that the role played by the fusoid cells in young grass leaves is related to synthesis and storage of starch granules at early stages of development.

Potential Plant-Plant Communication Induced by Infochemical Methyl Jasmonate in Sorghum (Sorghum bicolor).

Despite the fact that they are sessile organisms, plants actively move their organs and also use these movements to manipulate the surrounding biotic and abiotic environments. Plants maintain communication with neighboring plants, herbivores, and predators through the emission of diverse chemical compounds by their shoots and roots. These infochemicals modify the environment occupied by plants. Moreover, some infochemicals may induce morphophysiological changes of neighboring plants. We have used methyl-jasmonate (MeJa), a plant natural infochemical, to trigger communication between emitters and receivers Sorghum bicolor plants. The split roots of two plants were allocated to three different pots, with the middle pot containing the roots of both plants. We scored low stomatal conductance (gS) and low CO2 net assimilation (A) using the plants that had contact with the infochemical for the first time. During the second contact, these parameters showed no significant differences, indicating a memory effect. We also observed that the plants that had direct leaf contact with MeJa transmitted sensory information through their roots to neighboring plants. This resulted in higher maximum fluorescence (FM) and structural changes in root anatomy. In conclusion, MeJa emerges as possible trigger for communication between neighboring sorghum plants, in response to the environmental challenges.

Anatomical and ultrastructural studies reveal temporal and spatial variation in the oil production in leaves of the diesel tree (Copaifera langsdorffii, Leguminosae).

The oily resin produced by Copaifera langsdorffii, commonly called oil of copaiba, is widely exploited by the drug, cosmetic, and biodiesel industries. The distribution of oily secretory cavities and canals (secretory spaces) over the vegetative body characterizes this species. Oil is stored inside the lumen of the secretory spaces and only reaches the organ surface after injuries. Nonetheless, translucent oily deposits occur on the adaxial surface of intact young leaves. In this study, we searched for further sources of oil production in C. langsdorffii leaves in addition to the well-known secretory cavities and investigated the mechanisms of secretion. Leaves in different developmental stages were collected from adult plants and processed for studies on light and transmission electron microscopies. The primary finding of this study was the involvement of the chlorenchyma cells in lipid biosynthesis, in addition to the secretory cavities. The secretory activity of cavities and chlorenchyma cells overlapped in young leaves. Ultrastructurally, secretory cavity cells exhibited abundant smooth endoplasmic reticulum profiles and oleoplasts, whereas the chlorenchyma cells had large chloroplasts with oil inclusions. Our data suggest that the oily material on the leaf surface arose from the chlorenchyma and was transported via the apoplast. These findings open new avenues for understanding oil biosynthesis regulation in mesophyll cells and planning of future strategies for the biotechnological application of C. langsdorffii leaves.

Resin secretory canals in Protium heptaphyllum (Aubl.) Marchand. (Burseraceae): a tridimensional branched and anastomosed system.

Protium heptaphyllum is a Burseraceae species known by the production of aromatic resin with medicinal, economic, and ecological values. Information on the development, architecture, and lifetime of the secretory system are crucial to understand the resin production and contribute to a more sustainable tapping regime. We investigated the histology and ultrastructure of the secretory canals under a developmental point of view. Stem samples were analyzed under light and transmission electron microscopy by conventional and cytochemical methods. Secretory canals, originated from procambium and cambium, occurred immersed in the primary and secondary phloem. Mature canals have a secretory epithelium and a wide lumen where the exudate is accumulated. A sheath of parenchyma cells with meristematic features surrounds the epithelium. The canals originate by schizogenesis and develop by schyzolysigenesis. Canals active in secretion occurred since the shoot apex and near the cambium. In the dilation zone of the secondary phloem, secretory canals exhibit sclerified epithelial and sheath cells and are inactive in secretion. Secreting epithelial cells have subcellular apparatus consistent with oleoresin, polysaccharides, and enzymes secretion. Pectinase and cellulase were cytochemically detected in developing canals and are involved in cell wall changes associated to canal growth and release of exudate. In P. heptaphyllum, the secretory system has a complex structure resultant from longitudinal growth, lateral ramification, and fusion of the adjacent canals, in addition to intrusive growth of both epithelial and sheath cells. Although some anatomical results are already known, ultrastructural data represent the novelty of this work. Our findings can contribute to the establishment of more efficient and sustainable techniques for resin extraction in this species.

Structure and functioning of oil cavities in the shoot apex of Metrodorea nigra A. St.-Hil. (Rutaceae).

This study investigates the histology and subcellular features of secretory cavities during the development of the shoot apex of Metrodorea nigra A. St.-Hil. in order to better understand the functioning of these glands. This Rutaceae species is a very suitable model for studying secretory cavity life span, since the shoot apex exhibits both dormant and growth stages during its annual cycle. Shoot apices were collected during the dormant and growth stages from populations of M. nigra growing under natural conditions. Materials were processed using standard techniques for light and electron microscopy. The secretory cavities originate under the protodermis, and their initiation is restricted to the early developmental stage of shoot organs, which are protected by a hood-shaped structure. Secretory cavities have a multi-seriate epithelium surrounding a lumen that expands schizolysigenously. Oil production begins before lumen formation. When the shoot apex resumes development after the dormant stage, the glands remain active in oil secretion in the developing shoot apex and fully expanded leaves. The mature epithelial cells are flattened and exhibit very thin walls, large oil bodies, leucoplasts surrounded by endoplasmic reticulum, and mitochondria with unusual morphology. The tangential walls of the epithelial cells facing the lumen undergo continuous peeling. The vacuole extrusion appears to be the primary mode of release oil into the lumen, in an exocytotic way. The continuity of oil secretion is ensured by the replacement of the damaged inner epithelial cells by divisions in the parenchyma layer that surround the oil gland, likely a meristematic sheath.

PEG-induced osmotic stress in Mentha x piperita L.: Structural features and metabolic responses.

The present study investigated whether osmotic stress induced by the exposure of peppermint (Mentha x piperita L.) to moderate and severe stress for short periods of time changes the plant's physiological parameters, leaf anatomy and ultrastructure and essential oil. Plants were exposed to two levels of polyethyleneglycol (50 g L(-1) and 100 g L(-1) of PEG) in a hydroponic experiment. The plants exposed to 50 g L(-1) maintained metabolic functions similar to those of the control group (0 g L(-1)) without changes in gas exchange or structural characteristics. The increase in antioxidant enzyme activity reduced the presence of free radicals and protected membranes, including chloroplasts and mitochondria. In contrast, the osmotic stress caused by 100 g L(-1) of PEG inhibited leaf gas exchange, reduced the essential oil content and changed the oil composition, including a decrease in menthone and an increase in menthofuran. These plants also showed an increase in peroxidase activity, but this increase was not sufficient to decrease the lipid peroxidation level responsible for damaging the membranes of organelles. Morphological changes were correlated with the evaluated physiological features: plants exposed to 100 g L(-1) of PEG showed areas with collapsed cells, increases in mesophyll thickness and the area of the intercellular space, cuticle shrinkage, morphological changes in plastids, and lysis of mitochondria. In summary, our results revealed that PEG-induced osmotic stress in M. x piperita depends on the intensity level of the osmotic stress applied; severe osmotic stress changed the structural characteristics, caused damage at the cellular level, and reduced the essential oil content and quality.

Flavonoids modify root growth and modulate expression of SHORT-ROOT and HD-ZIP III.

Flavonoids are a class of distinct compounds produced by plant secondary metabolism that inhibit or promote plant development and have a relationship with auxin transport. We showed that, in terms of root development, Copaifera langsdorffii leaf extracts has an inhibitory effect on most flavonoid components compared with the application of exogenous flavonoids (glycosides and aglycones). These compounds alter the pattern of expression of the SHORT-ROOT and HD-ZIP III transcription factor gene family and cause morpho-physiological alterations in sorghum roots. In addition, to examine the flavonoid auxin interaction in stress, we correlated the responses with the effects of exogenous application of auxin and an auxin transport inhibitor. The results show that exogenous flavonoids inhibit primary root growth and increase the development of lateral roots. Exogenous flavonoids also change the pattern of expression of specific genes associated with root tissue differentiation. These findings indicate that flavonoid glycosides can influence the polar transport of auxin, leading to stress responses that depend on auxin.

The role of the parenchyma sheath and PCD during the development of oil cavities in Pterodon pubescens (Leguminosae-Papilionoideae).

Pterodon pubescens cavities are constituted by lumen and uniseriated epithelium surrounded by multiseriate parenchyma sheath. We studied the development of secretory cavities, including the role of parenchyma sheath, using light and transmission electron microscopy. A Tunel assay was performed to verify whether programmed cell death (PCD) occurs during the process. The lumen is formed by schizogeny and lysigeny occur in later developmental stages of the secretory cavities. Ultrastructurally, epithelial cells in later developmental stages become dark and with sinuous walls; the protoplast becomes retracted and the cytoplasm shows low organelle definition. Degenerated cells are released toward the lumen. Our results showed that PCD occurs during later developmental stages of cavities and plays a critical role in functioning of these glands. New cells originated from the parenchyma sheath differentiate into secretory cells and replace those degenerated ones. This fact associated to PCD guarantees epithelium renovation during the secretory cycle and the maintenance of secretory activity of cavities.