Phospholipases C and D and Their Role in Biotic and Abiotic Stresses.

Plants, as sessile organisms, have adapted a fine sensing system to monitor environmental changes, therefore allowing the regulation of their responses. As the interaction between plants and environmental changes begins at the surface, these changes are detected by components in the plasma membrane, where a molecule receptor generates a lipid signaling cascade via enzymes, such as phospholipases (PLs). Phospholipids are the key structural components of plasma membranes and signaling cascades. They exist in a wide range of species and in different proportions, with conversion processes that involve hydrophilic enzymes, such as phospholipase-C (PLC), phospholipase-D (PLD), and phospholipase-A (PLA). Hence, it is suggested that PLC and PLD are highly conserved, compared to their homologous genes, and have formed clusters during their adaptive history. Additionally, they generate responses to different functions in accordance with their protein structure, which should be reflected in specific signal transduction responses to environmental stress conditions, including innate immune responses. This review summarizes the phospholipid systems associated with signaling pathways and the innate immune response.

Capsaicin Synthesis Requires in Situ Phenylalanine and Valine Formation in in Vitro Maintained Placentas from Capsicum chinense.

Capsaicinoids (CAP) are nitrogenous metabolites formed from valine (Val) and phenylalanine (Phe) in the placentas of hot Capsicum genotypes. Placentas of Habanero peppers can incorporate inorganic nitrogen into amino acids and have the ability to secure the availability of the required amino acids for CAP biosynthesis. In order to determine the participation of the placental tissue as a supplier of these amino acids, the effects of blocking the synthesis of Val and Phe by using specific enzyme inhibitors were analyzed. Isolated placentas maintained in vitro were used to rule out external sources' participation. Blocking Phe synthesis, through the inhibition of arogenate dehydratase, significantly decreased CAP accumulation suggesting that at least part of Phe required in this process has to be produced in situ. Chlorsulfuron inhibition of acetolactate synthase, involved in Val synthesis, decreased not only Val accumulation but also that of CAP, pointing out that the requirement for this amino acid can also be fulfilled by this tissue. The presented data demonstrates that CAP accumulation in in vitro maintained placentas can be accomplished through the in situ availability of Val and Phe and suggests that the synthesis of the fatty acid chain moiety may be a limiting factor in the biosynthesis of these alkaloids.

Fructans and other water soluble carbohydrates in vegetative organs and fruits of different Musa spp. accessions.

The water soluble carbohydrates (WSC) glucose, fructose, and sucrose are well-known to the great public, but fructans represent another type of WSC that deserves more attention given their prebiotic and immunomodulatory properties in the food context. Although the occurrence of inulin-type fructo-oligosaccharides (FOS) was proposed in the fruit of some banana accessions, little or no information is available neither on the exact identity of the fructan species, nor on the fructan content in different parts of banana plants and among a broader array of banana cultivars. Here, we investigated the WSC composition in leaves, pulp of ripe fruits and rhizomes from mature banana plants of 11 accessions (I to XI), including both cultivated varieties and wild Musa species. High performance anion exchange chromatography with integrated pulsed amperometric detection (HPAEC-IPAD) showed the presence of 1-kestotriose [GF2], inulobiose [F2], inulotriose [F3], 6-kestotriose and 6G-kestotriose (neokestose) fructan species in the pulp of mature fruits of different accessions, but the absence of 1,1-nystose and 1,1,1 kestopentaose and higher degree of polymerization (DP) inulin-type fructans. This fructan fingerprint points at the presence of one or more invertases that are able to use fructose and sucrose as alternative acceptor substrates. Quantification of glucose, fructose, sucrose and 1-kestotriose and principal component analysis (PCA) identified related banana groups, based on their specific WSC profiles. These data provide new insights in the biochemical diversity of wild and cultivated bananas, and shed light on potential roles that fructans may fulfill across species, during plant development and adaptation to changing environments. Furthermore, the promiscuous behavior of banana fruit invertases (sucrose and fructose as acceptor substrates besides water) provides a new avenue to boost future work on structure-function relationships on these enzymes, potentially leading to the development of genuine banana fructosyltransferases that are able to increase fructan content in banana fruits.

Nitrate promotes capsaicin accumulation in Capsicum chinense immobilized placentas.

In chili pepper's pods, placental tissue is responsible for the synthesis of capsaicinoids (CAPs), the compounds behind their typical hot flavor or pungency, which are synthesized from phenylalanine and branched amino acids. Placental tissue sections from Habanero peppers (Capsicum chinense Jacq.) were immobilized in a calcium alginate matrix and cultured in vitro, either continuously for 28 days or during two 14-day subculture periods. Immobilized placental tissue remained viable and metabolically active for up to 21 days, indicating its ability to interact with media components. CAPs contents abruptly decreased during the first 7 days in culture, probably due to structural damage to the placenta as revealed by scanning electron microcopy. CAPs levels remained low throughout the entire culture period, even though a slight recovery was noted in subcultured placentas. However, doubling the medium's nitrate content (from 40 to 80 mM) resulted in an important increment, reaching values similar to those of intact pod's placentas. These data suggest that isolated pepper placentas cultured in vitro remain metabolically active and are capable of metabolizing inorganic nitrogen sources, first into amino acids and, then, channeling them to CAP synthesis.

Antioxidant capacity and total phenolic content in fruit tissues from accessions of Capsicum chinense Jacq. (Habanero pepper) at different stages of ripening.

In the past few years, there has been a renewed interest in studying a wide variety of food products that show beneficial effects on human health. Capsicum is an important agricultural crop, not only because its economic importance, but also for the nutritional values of its pods, mainly due to the fact that they are an excellent source of antioxidant compounds, and also of specific constituents such as the pungent capsaicinoids localized in the placental tissue. This current study was designed to evaluate the antioxidant capacity and total phenolic contents from fruits tissues of two Capsicum chinense accessions, namely, Chak k'an-iik (orange) and MR8H (red), at contrasting maturation stages. Results showed that red immature placental tissue, with a Trolox equivalent antioxidant capacity (TEAC) value of 55.59 µmols TE g(-1) FW, exhibited the strongest total antioxidant capacity using both the 2,2-diphenyl-1-picrylhydrazyl (DPPH) and the CUPRAC methods. Placental tissue also had the highest total phenolic content (27 g GAE 100 g(-1) FW). The antioxidant capacity of Capsicum was directly related to the total amount of phenolic compounds detected. In particular, placentas had high levels of capsaicinoids, which might be the principal responsible for their strong antioxidant activities.

Measurement of cell viability.

An overview of the methods for assessing cell viability is presented. Different protocols of the most commonly used assays are described in detail so that the readers may be able to determine which assay is suitable for their own projects in plant biotechnology.

Exogenous nitrate induces root branching and inhibits primary root growth in Capsicum chinense Jacq.

The effects of nitrate (NO3⁻) on the root system are complex and depend on several factors, such as the concentration available to the plant, endogenous nitrogen status and the sensitivity of the species. Though these effects have been widely documented on Arabidopsis and cereals, no reports are available in the Capsicum genus. In this paper, we have determined the effect of an exogenous in vitro application of this nutrient on root growth in habanero pepper (Capsicum chinense Jacq.). Exposure to NO3⁻ inhibited primary root growth in both, dose- and time-dependent manners. The highest inhibition was attained with 0.1 mM NO3⁻ between the fourth and fifth days of treatment. Inhibition of primary root growth was observed by exposing the root to both homogeneous and heterogeneous conditions of the nutrient; in contrast, ammonium was not able to induce similar changes. NO3⁻-induced inhibition of primary root growth was reversed by treating the roots with IAA or NPA, a polar auxin transport inhibitor. Heterogeneous NO3⁻ application stimulated the formation and elongation of lateral roots in the segment where the nutrient was present, and this response was influenced by exogenous phytohormones. These results demonstrate that habanero pepper responds to NO3⁻ in a similar fashion to other species with certain particular differences. Therefore, studies in this model could help to elucidate the mechanisms by which roots respond to NO3⁻ in fluctuating soil environments.

Aluminum induces changes in oxidative burst scavenging enzymes in Coffea arabica L. suspension cells with differential Al tolerance.

The accumulation of reactive oxygen species (ROS) and concomitant oxidative stress have been considered deleterious consequences of aluminum toxicity. However, several lines of evidence suggest that ROS can function as important signaling molecules in the plant defense system for protection from abiotic stress and the acquisition of tolerance. The role of ROS-scavenging enzymes was assayed in two different coffee cell suspension lines. We treated L2 (Al-sensitive) and LAMt (Al-tolerant) Coffea arabica suspension cells with 100 µM AlCl(3) and observed significant differences in catalase activity between the two cell lines. However, we did not observe any differences in superoxide dismutase or glutathione reductase activity in either cell line following Al treatment. ROS production was diminished in the LAMt cell line. Taken together, these results indicate that aluminum treatment may impair the oxidative stress response in L2 cells but not in LAMt cells. We suggest a possible role for Al-induced oxidative bursts in the signaling pathways that lead to Al resistance and protection from Al toxicity.

Measurement of cell viability in in vitro cultures.

An overview of the methods for assessing cell viability in in vitro cultures is presented. The protocols of four of the most commonly used assays are described in detail, so the readers may be able to determine which assay is suitable for their own projects using plant cell cultures.