Virus-induced gene silencing (VIGS) analysis shows involvement of the LsSTPK gene in lettuce (Lactuca sativaL.) in high temperature-induced bolting.

To determine the effect of the serine/threonine protein kinase (STPK) gene on leaf lettuce bolting, we utilized virus-induced gene silencing (VIGS) using the TRV vector to silence the target gene. The 'GB30' leaf lettuce cultivar was the test material, and the methods included gene cloning, bioinformatics analysis, quantitative real-time PCR (qRT-PCR) and VIGS. LsSTPK, was cloned from the 'GB30' leaf lettuce cultivar via reverse transcription-polymerase chain reaction (RT-PCR). qRT-PCR analysis showed that the expression of LsSTPK in the stem of leaf lettuce was significantly greater than that in the roots and leaves, and after high-temperature treatment, the gene expression in the stems in the experimental group was markedly lower than that in the control groups. Following LsSTPK silencing via the VIGS method, the stem length in the treatment group was significantly greater than that in the blank and negative control groups, and the contents of auxin (IAA), GA3 and abscisic acid (ABA) in the treatment group were greater than those in the other two groups. Flower bud differentiation occurred in the treatment group but not in the control group. The above findings suggested that LsSTPK inhibits the bolting of leaf lettuce under high-temperature conditions.

Radio frequency energy inactivates peroxidase in stem lettuce at different heating rates and associate changes in physiochemical properties and cell morphology.

Radio frequency (RF) is an emerging technology applied in blanching treatment as alternative to conventional treatment. This study aimed to investigate the effects of RF heating rate on peroxidase (POD) inactivating efficiency, physiochemical properties (texture, color and vitamin C) and cell structure of stem lettuce. POD enzyme inactivation efficiency increased with increased RF heating rate. When POD activity was reduced to <5%, better physiochemical properties and less cell damage occurred in RF heating compared with conventional hot water (HW) blanching (HWB); Relative electrolyte leakage (REL) analysis and microscopic observation suggested that the loss of integrity of cell walls and membranes, the decrease in turgor pressure and the weakened connections between adjacent cells leaded to the deterioration of physiochemical properties. The degrees of cell destruction varied with RF heating rates, which provide a new idea for RF blanching for producing different types of fruits and vegetables products (Solid and juice products).

Identification of triterpenes and functional characterization of oxidosqualene cyclases involved in triterpene biosynthesis in lettuce (Lactuca sativa).

Lettuce (Lactuca sativa) is a member of the family Asteraceae and is most often used for green salads. Triterpenes are the largest class of natural compounds in plants and have beneficial health effects. Here, we identified various triterpene esters (taraxasterol acetates, ψ-taraxasterol acetates, taraxerol acetates, lupeol acetates, α-amyrin acetates, ß-amyrin acetates, and germanicol acetate) and free triterpenes (α-amyrin, ß-amyrin, taraxerol, and taraxasterol) in both the leaves and roots of lettuce. Triterpene biosynthesis occurs through the action of oxidosqualene cyclase (OSC), which generates various types of triterpenes from 2,3-oxidosqualene. None of the OSC genes involved in triterpene biosynthesis in lettuce have been characterized. Five putative lettuce OSC genes (LsOSC1, LsOSC2, LsOSC3, LsOSC4, and LsOSC5) were selected from a transcriptome database. These five genes were functionally characterized via heterologous expression in yeast. The first two enzymes were multifunctional triterpene synthase and the last three genes were monofunctional. Transgenic yeast expressing LsOSC1 produced five triterpenes, namely, taraxasterol, Ψ-taraxasterol, α-amyrin, ß-amyrin, and dammarenediol-II. Yeast expressing LsOSC2 produced baurenol and Ψ-taraxasterol. LsOSC3, LsOSC4, and LsOSC5 expression led to ß-amyrin, taraxerol, and lupeol production, respectively. Transcriptional activity assessment of the five genes revealed that all the OSC genes were more actively transcribed in roots than in leaves, and LsOSC5 among the five OSC genes showed the highest expression in both the leaves and the roots. In conclusion, we identified structurally diverse free triterpenes and triterpene esters in lettuce plants and characterized five OSC genes, which are key enzymes involved in the biosynthesis of diverse triterpenes in lettuce.

Physiological and Oxidative Stress Responses of Lettuce to Cleomside A: A Thiohydroximate, as a New Allelochemical from Cleome arabica L.

The inclination toward natural products have led the onset for the discovery of new bioactive metabolites that could be targeted for specific therapeutic or agronomic applications. This study aimed to isolate bioactive compounds from Cleome arabica L., and subsequently determine the unexplored mechanism of action of the newly identified compounds on Lactuca sativa L. Chemical investigation of the ethyl acetate fraction of methanolic silique extract of C. arabica afforded seven secondary metabolites belonging to different classes such as flavonoids, triterpene, and a new thiohydroximate derivative, named cleomside A. Among phytotoxic assays, the growth of lettuce was totally inhibited by cleomside A compared to the other identified compounds. This effect was associated with the increased levels of electrolyte leakage, malondialdehyde, and hydrogen peroxide indicating disruption of membrane integrity and induction of oxidative stress. Activities of the antioxidant enzymes SOD, CAT, and APX were also elevated, thereby demonstrating the enhanced generation of reactive oxygen species upon identified allelochemical exposure. Thus, the changes caused by cleomside A described herein can contribute to better understanding the allelochemical actions of thiohydroximate and the potential use of these substances in the production of natural herbicides compared to the other identified flavonoids and triterpene.

Effects of exogenous spermidine on antioxidants and glyoxalase system of lettuce seedlings under high temperature.

In this research, the lettuce high-temperature-sensitive variety Beisan San 3 was used as a test material. The effects of exogenous spermidine (Spd) on membrane lipid peroxidation, the antioxidant system, the ascorbic acid-glutathione (AsA-GSH) system and the glyoxalase (Glo) system in lettuce seedlings under high-temperature stress were studied by spraying either 1 mM spermidine or ionized water as a control. The results showed that, under high-temperature stress, the growth of lettuce seedlings was weak, and the dry weight (DW) and fresh weight (FW) were reduced by 68.9% and 82%, respectively, compared with those of the normal-temperature controls. In addition, the degree of membrane lipid peroxidation increased, and the reactive oxygen species (ROS) level increased, both of which led to a significant increase in malondialdehyde (MDA) content and lipoxygenase (LOX) activity. Under high-temperature stress, the activity of superoxide dismutase (SOD) decreased, the activities of peroxidase (POD) and catalase (CAT) increased first but then decreased, and the activity of ascorbic acid peroxidase (APX) decreased first but then increased. Glutathione reductase (GR) activity, ascorbic acid (AsA) and glutathione (GSH) content showed an upward trend under high-temperature stress. The activities of glyoxalase (GloI and GloII) in the lettuce seedling leaves increased significantly under high-temperature stress. In contrast, the application of exogenous Spd alleviated the oxidative damage to the lettuce seedlings, which showed a decrease in MDA content and LOX activity and an increase in SOD, POD, CAT, APX, GR, GloI, and GloII activities. In addition, the antioxidant AsA and GSH contents also increased to varying degrees. It can be seen from the results that high temperature stress leads to an increase in the level of ROS and cause peroxidation in lettuce seedlings, and exogenous Spd can enhance the ability of lettuce seedlings to withstand high temperature by enhancing the antioxidant system, glyoxalase system and AsA-GSH cycle system.

The NADPH-oxidase LsRbohC1 plays a role in lettuce (Lactuca sativa) seed germination.

Reactive oxygen species (ROS) are increasingly recognized as non-enzymatic players in the processes of radicle elongation growth and endosperm weakening during seed germination. NADPH oxidases (EC 1.6.3.1), also known as respiratory burst oxidase homologues (Rbohs), are key enzymes for the production of ROS. We previously reported that ROS are involved in the radicle elongation growth and endosperm weakening during lettuce seed germination. However, the function of the gene(s) encoding Rbohs during lettuce seed germination remains to be elucidated. In this study, one lettuce Rboh gene LsRbohC1 was cloned, and over-expression and RNAi-lines of this gene were generated. It was found that LsRbohC1 was abundantly expressed in germinating seeds, especially in the endosperm cap and hypocotyl. Over-expression/knock-down of this gene observably increased/decreased the production of superoxide radicals in the radicle and endosperm cap, and significantly promoted/delayed the germination process. The results suggest that LsRbohC1 plays a role in lettuce seed germination.

Bacillus subtilis strain L1 promotes nitrate reductase activity in Arabidopsis and elicits enhanced growth performance in Arabidopsis, lettuce, and wheat.

Plant growth promoting rhizobacteria (PGPR) are a group of bacteria that promote plants growth in the rhizosphere. PGPRs are involved in various mechanisms that reinforce plant development. In this study, we screened for PGPRs that were effective in early growth of Arabidopsis thaliana when added to the media and one Bacillus subtilis strain L1 (Bs L1) was selected for further study. When Bs L1 was placed near the roots, seedlings showed notably stronger growth than that in the control, particularly in biomass and root hair. Quantitative reverse transcription polymerase chain reaction analysis revealed a high level of expression of the high affinity nitrate transporter gene, NRT2.1 in A. thaliana treated with Bs L1. After considering how Bs L1 could promote plant growth, we focused on nitrate, which is essential to plant growth. The nitrate content was lower in A. thaliana treated with Bs L1. However, examination of the activity of nitrate reductase revealed higher activity in plants treated with PGPR than in the control. Bs L1 had pronounced effects in representative crops (wheat and lettuce). These results suggest that Bs L1 promotes the assimilation and use of nitrate and plant growth.

Fine monitoring of major phenolic compounds in lettuce and escarole leaves during storage.

Two varieties of lettuce (Lactuca sativa L. var. crispa and capitata) and one variety of escarole (Cichorium endivia var. latifolium) were chosen due to their different browning sensitivity during storage as minimally processed products. The changes in the compositions and contents of the primary polyphenolic compounds were investigated during the first few hours of storage and then after 1, 2, 3, and 6 days of storage at 6°C and revealed sharp variations. Browning development, activity of phenylalanine ammonia lyase, and concentration of ascorbic acid were also examined during storage. The content of chicoric acid, which was the most abundant phenolic compound, varied substantially during the first 24 hr of storage and between the different varieties. Oak leaf lettuce, which was the most sensitive variety to browning, was characterized by a higher maintained chicoric acid level with a constant decreased chlorogenic acid level during the storage period. PRACTICAL APPLICATIONS: Browning development is responsible for the short shelf life of minimally processed vegetables, such as lettuce (Lactuca sativa). Polyphenolic compounds, as substrates of enzymatic reactions, are involved in the browning susceptibility of leaves. Comparisons of the profiles and contents of these compounds in different leaves showed dramatic variations during storage. This study provides additional information to better control, optimize minimally processed produce and select more suitable leaves for the fresh-cut industry.

Effect of mild heat treatment on browning-related parameters in fresh-cut Iceberg lettuce.

Enzymatic browning of Iceberg lettuce was studied by subjecting midrib tissues to a series of mild heat treatments. The effects of wounding and subsequent application of a mild heat treatment were examined by monitoring the browning potential (BP) and the activity of three browning-related enzymes (i.e., phenylalanine ammonia lyase [PAL], polyphenol oxidase [PPO], and peroxidase [POD]) during refrigerated storage up to 10 days. Efficient inhibition of browning was achieved by treatment at 50°C for 60 s. The wound-induced increase of the BP and the activity of PAL and POD was effectively suppressed, maintaining their values at initial levels up to 7 days of storage. PPO activity, on the contrary, remained unchanged after wounding, whether or not followed by heat treatment. BP, PAL activity and POD were found to be strongly correlated, whereas meaningful associations for PPO with the other parameters could not be established. PRACTICAL APPLICATIONS: In an attempt to answer to the growing demand in the fresh-cut produce industry to control browning, heat treatment was investigated as interesting alternative to chemical preservation methods. Efficient control of enzymatic browning in fresh-cut Iceberg lettuce could be achieved by heat treatment at 50°C for 60 s. Experimental data are provided showing the effects of wounding and subsequent heat treatment on visual browning, the BP and the activity of PAL, PPO, and POD during refrigerated storage up to 10 days. Using this data, correlations were found for BP, PAL activity, and POD activity, but not for PPO. Although undesired side effects of heat treatment (e.g., tissue softening) cannot be excluded, the obtained information might be useful for further research, serving as a baseline for wound-induced effects on browning-related parameters in fresh-cut lettuce and possible mechanisms of action of inhibitory treatments.

Study of the contamination rate and change in growth features of lettuce (Lactuca sativa Linn.) in response to cadmium and a survey of its phytochelatin synthase gene.

Crops can become contaminated when grown in soils containing heavy metals. Cadmium is a heavy metal that poses a significant health risk to humans. The purpose of this study was to evaluate the effect of cadmium on lettuce (Lactuca sativa Linn) and the contamination risk of lettuce grown in cadmium environments. The results showed that photosynthesis and growth parameters were significantly affected by cadmium. Lettuce has the ability to absorb large amounts of cadmium from the contaminated environment and so is a cadmium hyperaccumulator plant. The study showed that approximately 35% of the total absorbed cadmium is transmitted to aerial and edible parts of lettuce. This study was undertaken as lettuce has the ability to absorb and accumulate high levels of cadmium. There are however are no reports on the PCS gene and the potential for high cadmium accumulation in lettuce. The bioinformatics study revealed that lettuce has two phytochelatin synthase genes that produce 6 PCSs through splicing leading to the ability of lettuce to store high levels of cadmium. These six sequences although different in length have high similarity. Sequence structure, cellular location, three-dimensional structure, phylogeny and a comparison of their catalytic power were evaluated. The high accumulation of cadmium in lettuce and the presence of several PCSs contribute to the accumulation of cadmium in aerial tissues. The cultivation of lettuce in contaminated environments led us to evaluate suspected farms for the presence of cadmium in produce. Lettuce grown in industrial environments contaminated with cadmium can pose a serious threat to human health.

Validation of leaf and microbial pectinases: commercial launching of a new platform technology.

Almost all current genetically modified plant commercial products are derived from seeds. The first protein product made in leaves for commercial use is reported here. Leaf pectinases are validated here with eight liquid commercial microbial enzyme products for textile or juice industry applications. Leaf pectinases are functional in broad pH/temperature ranges as crude leaf extracts, while most commercial enzyme products showed significant loss at alkaline pH or higher temperature, essential for various textile applications. In contrast to commercial liquid enzymes requiring cold storage/transportation, leaf pectinase powder was stored up to 16 months at ambient temperature without loss of enzyme activity. Commercial pectinase products showed much higher enzyme protein PAGE than crude leaf extracts with comparable enzyme activity without protease inhibitors. Natural cotton fibre does not absorb water due to hydrophobic nature of waxes and pectins. After bioscouring with pectinase, measurement of contact-angle water droplet absorption by the FAMAS videos showed 33 or 63 (leaf pectinase), 61 or 64 (commercial pectinase) milliseconds, well below the 10-second industry requirements. First marker-free lettuce plants expressing pectinases were also created by removal of the antibiotic resistance aadA gene. Leaf pectinase powder efficiently clarified orange juice pulp similar to several microbial enzyme products. Commercial pilot scale biomass production of tobacco leaves expressing different pectinases showed that hydroponic growth at Fraunhofer yielded 10 times lower leaf biomass per plant than soil-grown plants in the greenhouse. Pectinase enzyme yield from the greenhouse plants was double that of Fraunhofer. Thus, this leaf-production platform offers a novel, low-cost approach for enzyme production by elimination of fermentation, purification, concentration, formulation and cold chain.

Cadmium-accumulator straw application alleviates cadmium stress of lettuce (Lactuca sativa) by promoting photosynthetic activity and antioxidative enzyme activities.

The effects of application of straw derived from cadmium (Cd) accumulator plants (Siegesbeckia orientalis, Conyza canadensis, Eclipta prostrata, and Solanum photeinocarpum) on growth and Cd accumulation of lettuce plants grown under Cd exposure were studied. Treatment with straw of the four Cd-accumulator species promoted growth, photosynthesis, and soluble protein contents and enhanced the activities of peroxidase in leaves of lettuce seedlings. The biomass of shoot of lettuce from high to low in turn is the treatment of C. canadensis straw > S. photeinocarpum straw > S. orientalis > E. prostrata > Control. The Cd content in edible parts (shoots) of the lettuce plants was significantly decreased in the presence of straw from the Cd-accumulator species, except the presence of the straw of E. prostrata. And, the greatest reduction in Cd content in shoots was 27.09% in the S. photeinocarpum straw treatment compared with that of the control. Therefore, application of straw of S. orientalis, C. canadensis, and S. photeinocarpum can promote the growth of lettuce seedlings, and decrease their Cd accumulation, when grown in Cd-contaminated soil, which is beneficial for production of lettuce safe for human consumption.

Involvement of S-nitrosothiols modulation by S-nitrosoglutathione reductase in defence responses of lettuce and wild Lactuca spp. to biotrophic mildews.

MAIN CONCLUSION: Resistant Lactuca spp. genotypes can efficiently modulate levels of S-nitrosothiols as reactive nitrogen species derived from nitric oxide in their defence mechanism against invading biotrophic pathogens including lettuce downy mildew. S-Nitrosylation belongs to principal signalling pathways of nitric oxide in plant development and stress responses. Protein S-nitrosylation is regulated by S-nitrosoglutathione reductase (GSNOR) as a key catabolic enzyme of S-nitrosoglutathione (GSNO), the major intracellular S-nitrosothiol. GSNOR expression, level and activity were studied in leaves of selected genotypes of lettuce (Lactuca sativa) and wild Lactuca spp. during interactions with biotrophic mildews, Bremia lactucae (lettuce downy mildew), Golovinomyces cichoracearum (lettuce powdery mildew) and non-pathogen Pseudoidium neolycopersici (tomato powdery mildew) during 168 h post inoculation (hpi). GSNOR expression was increased in all genotypes both in the early phase at 6 hpi and later phase at 72 hpi, with a high increase observed in L. sativa UCDM2 responses to all three pathogens. GSNOR protein also showed two-phase increase, with highest changes in L. virosa-B. lactucae and L. sativa cv. UCDM2-G. cichoracearum pathosystems, whereas P. neolycopersici induced GSNOR protein at 72 hpi in all genotypes. Similarly, a general pattern of modulated GSNOR activities in response to biotrophic mildews involves a two-phase increase at 6 and 72 hpi. Lettuce downy mildew infection caused GSNOR activity slightly increased only in resistant L. saligna and L. virosa genotypes; however, all genotypes showed increased GSNOR activity both at 6 and 72 hpi by lettuce powdery mildew. We observed GSNOR-mediated decrease of S-nitrosothiols as a general feature of Lactuca spp. response to mildew infection, which was also confirmed by immunohistochemical detection of GSNOR and GSNO in infected plant tissues. Our results demonstrate that GSNOR is differentially modulated in interactions of susceptible and resistant Lactuca spp. genotypes with fungal mildews and uncover the role of S-nitrosylation in molecular mechanisms of plant responses to biotrophic pathogens.

Expression analysis of polyphenol oxidase isozymes by active staining method and tissue browning of head lettuce (Lactuca sativa L.).

Browning of plant tissue is generally considered attributable to enzymatic oxidation by polyphenol oxidase (PPO). Electrophoresis followed by activity staining has been used as an effective procedure to visually detect and isolate isozymes; however, it has not been applied for examination of various PPO isozymes in lettuce. Our study demonstrated that different lettuce PPO isozymes could be detected at different pH in active staining, and multiple isozymes were detected only under alkaline conditions. As a result, we concluded that activity staining with approximately pH 8 enabled to detect various PPO isozymes in lettuce. By expression analysis of the PPO isozymes after wounding, PPO isozymes that correlated with time-course of tissue browning were detected. The wound-induced PPO may play a key role in enzymatic browning.

Characterization of S-nitrosoglutathione reductase from Brassica and Lactuca spp. and its modulation during plant development.

Cellular homeostasis of S-nitrosoglutathione (GSNO), a major cache of nitric oxide bioactivity in plants, is controlled by the NADH-dependent S-nitrosoglutathione reductase (GSNOR) belonging to the family of class III alcohol dehydrogenases (EC 1.1.1.1). GSNOR is a key regulator of S-nitrosothiol metabolism and is involved in plant responses to abiotic and biotic stresses. This study was focused on GSNOR from two important crop plants, cauliflower (Brassica oleracea var. botrytis, BoGSNOR) and lettuce (Lactuca sativa, LsGSNOR). Both purified recombinant GSNORs were characterized in vitro and found to exists as dimers, exhibit high thermal stability and substrate preference towards GSNO, although both enzymes have dehydrogenase activity with a broad range of long-chain alcohols and ω-hydroxy fatty acids in presence of NAD+. Data on enzyme affinities to their cofactors NADH and NAD+ obtained by isothermal titration calorimetry suggest the high affinity to NADH might underline the GSNOR capacity to function in the intracellular environment. GSNOR activity and gene expression peak during early developmental stages of lettuce and cauliflower at 20 and 30 days after germination, respectively. GSNOR activity was also measured in four other Lactuca spp. genotypes with different degree of resistance to biotrophic pathogen Bremia lactucae. Higher GSNOR activities were found in non-infected plants of susceptible genotypes L. sativa UCDM2 and L. serriola as compared to resistant genotypes. GSNOR and GSNO were localized by confocal laser scanning microscopy in vascular bundles and in epidermal and parenchymal cells of leaf cross-sections. The presented results bring new insight in the role of GSNOR in the regulation of S-nitrosothiol levels in plant growth and development.

Zn-biofortification enhanced nitrogen metabolism and photorespiration process in green leafy vegetable Lactuca sativa L.

BACKGROUND: Excessive rates of nitrogen (N) fertilizers may result in elevated concentrations of nitrate (NO3- ) in plants. Considering that many programs of biofortification with trace elements are being performed, it has become important to study how the application of these elements affects plant physiology and, particularly, N utilization in leaf crops. The main objective of the present study was to determine whether the NO3- accumulation and the nitrogen use efficiency was affected by the application of different doses of Zn in Lactuca sativa plants. RESULTS: Zn doses in the range 80-100 µmol L-1 produced an increase in Zn concentration provoking a decrease of NO3- concentration and increase of the nitrate reductase, glutamine synthetase and aspartate aminotransferase activities, as well as the photorespiration processes. As result, we observed an increase in reduced N, total N concentration and N utilization efficiency. Consequently, at a dose of 80 µmol L-1 of Zn, the amino acid concentration increased significantly. CONCLUSION: Adequate Zn fertilization is an important critical player in lettuce, especially at a dose of 80 µmol L-1 of Zn, because it could result in an increase in the Zn concentration, a reduction of NO3- levels and an increase the concentration of essential amino acids, with all of them having beneficial properties for the human diet. © 2016 Society of Chemical Industry.

Abscisic acid and ethephon regulation of cellulase in the endosperm cap and radicle during lettuce seed germination.

The purpose of this study was to investigate the role of cellulase in endosperm cap weakening and radicle elongation during lettuce (Lactuca sativa L.) seed germination. The application of abscisic acid (ABA) or ethephon inhibits or promotes germination, respectively, by affecting endosperm cap weakening and radicle elongation. Cellulase activities, and related protein and transcript abundances of two lettuce cellulase genes, LsCEL1 and LsCEL2, increase in the endosperm cap and radicle prior to radicle protrusion following imbibition in water. ABA or ethephon reduce or elevate, respectively, cellulase activity, and related protein and transcript abundances in the endosperm cap. Taken together, these observations suggest that cellulase plays a role in endosperm cap weakening and radicle elongation during lettuce seed germination, and that the regulation of cellulase in the endosperm cap by ABA and ethephon play a role in endosperm cap weakening. However, the influence of ABA and ethephon on radicle elongation may not be through their effects on cellulase.

Roles of MPBQ-MT in Promoting α/γ-Tocopherol Production and Photosynthesis under High Light in Lettuce.

2-methyl-6-phytyl-1, 4-benzoquinol methyltransferase (MPBQ-MT) is a vital enzyme catalyzing a key methylation step in both α/γ-tocopherol and plastoquinone biosynthetic pathway. In this study, the gene encoding MPBQ-MT was isolated from lettuce (Lactuca sativa) by rapid amplification of cDNA ends (RACE), named LsMT. Overexpression of LsMT in lettuce brought about a significant increase of α- and γ-tocopherol contents with a reduction of phylloquinone (vitamin K1) content, suggesting a competition for a common substrate phytyl diphosphate (PDP) between the two biosynthetic pathways. Besides, overexpression of LsMT significantly increased plastoquinone (PQ) level. The increase of tocopherol and plastoquinone levels by LsMT overexpression conduced to the improvement of plants' tolerance and photosynthesis under high light stress, by directing excessive light energy toward photosynthetic production rather than toward generation of more photooxidative damage. These findings suggest that the role and function of MPBQ-MT can be further explored for enhancing vitamin E value, strengthening photosynthesis and phototolerance under high light in plants.

Selected biochemical properties of polyphenol oxidase in butter lettuce leaves (Lactuca sativa L. var. capitata) elicited with dl-ß-amino-n-butyric acid.

The study concentrated on changes in certain biochemical parameters of polyphenol oxidase (PPO) from lettuce leaves caused by dl-ß-amino-n-butyric acid (BABA) elicitation. PPO from control plants demonstrated the highest affinity toward catechol, whereas PPO from BABA-elicited lettuce showed the highest affinity to 4-methylcatechol. The optimum temperature for enzymes from control plants was 35°C, whereas from plants elicited with 1mM BABA this was 25°C. PPO from plants elicited with BABA was also more sensitive to the tested inhibitors than PPO from control plants. l-Cysteine was the most effective inhibitor. Native gel stained for PPO activity in control samples showed two isoforms. However, in BABA-treated lettuce three bands visualising PPO activity were observed. The information obtained in this study will be valuable for the development of treatment technology and storage conditions to control undesirable browning reactions in elicited lettuce.

Involvement of reactive oxygen species in endosperm cap weakening and embryo elongation growth during lettuce seed germination.

Endosperm cap (CAP) weakening and embryo elongation growth are prerequisites for the completion of lettuce seed germination. Although it has been proposed that the cell wall loosening underlying these processes results from an enzymatic mechanism, it is still unclear which enzymes are involved. Here it is shown that reactive oxygen species (ROS), which are non-enzymatic factors, may be involved in the two processes. In Guasihong lettuce seeds imbibed in water, O2·(-) and H2O2 accumulated and peroxidase activity increased in the CAP, whereas its puncture force decreased. In addition, in the radicle, the increase in embryo growth potential was accompanied by accumulation of O2·(-) and an increase in peroxidase activity. Imbibing seeds in 0.3% sodium dichloroisocyanurate (SDIC) reduced endosperm viability and the levels of O2·(-), H2O2, and peroxidase activity in the CAP, whereas the decrease in its puncture force was inhibited. However, in the embryo, SDIC did not affect the accumulation of O2·(-), peroxidase activity, and the embryo growth potential. As a result, SDIC caused atypical germination, in which the endosperm ruptured at the boundary between the CAP and lateral endosperm. ROS scavengers and ROS generation inhibitors inhibited the CAP weakening and also decreased the embryo growth potential, thus decreasing the percentage of seed germination. Exogenous ROS and ROS generation inducers increased the percentage of CAP rupture to some extent, and the addition of H2O2 to 0.3% SDIC enabled some seeds to undergo typical germination.