The maize fused leaves1 (fdl1) gene controls organ separation in the embryo and seedling shoot and promotes coleoptile opening.

The fdl1-1 mutation, caused by an Enhancer/Suppressor mutator (En/Spm) element insertion located in the third exon of the gene, identifies a novel gene encoding ZmMYB94, a transcription factor of the R2R3-MYB subfamily. The fdl1 gene was isolated through co-segregation analysis, whereas proof of gene identity was obtained using an RNAi strategy that conferred less severe, but clearly recognizable specific mutant traits on seedlings. Fdl1 is involved in the regulation of cuticle deposition in young seedlings as well as in the establishment of a regular pattern of epicuticular wax deposition on the epidermis of young leaves. Lack of Fdl1 action also correlates with developmental defects, such as delayed germination and seedling growth, abnormal coleoptile opening and presence of curly leaves showing areas of fusion between the coleoptile and the first leaf or between the first and the second leaf. The expression profile of ZmMYB94 mRNA-determined by quantitative RT-PCR-overlaps the pattern of mutant phenotypic expression and is confined to a narrow developmental window. High expression was observed in the embryo, in the seedling coleoptile and in the first two leaves, whereas RNA level, as well as phenotypic defects, decreases at the third leaf stage. Interestingly several of the Arabidopsis MYB genes most closely related to ZmMYB94 are also involved in the activation of cuticular wax biosynthesis, suggesting deep conservation of regulatory processes related to cuticular wax deposition between monocots and dicots.

Ultrastructural changes, pigment responses and bioaccumulation in the microalga Phaeodactylum tricornutum Bohlin exposed to BPA analogues.

As well-known, microalgae have a pivotal role in aquatic environments, being the primary producer. In this study, we investigated the effects of Bisphenol A (BPA) analogues on cell ultrastructure, reactive oxygen species (ROS) production and photosynthetic pigment responses in the diatom Phaeodactylum tricornutum. Microalgae were exposed during both exponential and stationary growth phases to an environmental relevant concentration (300 ng/L) of three differing BPA analogues (BPAF, BPF, and BPS) and their mixture (100 ng/L of each compound). Bioaccumulation of such compounds in microalgae was also analysed. During the stationary growth phase, a significant increase in the percentage of cells with hydrogen peroxide production was recorded after exposure to both BPS and MIX. Conversely, no significant effects on total chlorophylls and carotenoids were observed. During exponential growth phase we observed that control cultures had chloroplasts with well-organized thylakoid membranes and a central pyrenoid. On the contrary, the culture cells treated with BPA analogues and MIX showed chloroplasts characterized by evident dilation of thylakoid membranes. The presence of degeneration areas in the cytoplasm was also recorded. During the stationary growth phase, control and culture cells were characterized by chloroplasts with a regular thylakoid system, whereas BPA analogues-exposed cells were characterized by a deep degradation of the cytoplasm but showed chloroplasts without evident alterations of the thylakoid system. Lipid bodies were visible in treated microalgae. Lastly, microalgae bioaccumulated mainly BPS and BPF, alone or in the MIX. Overall, results obtained revealed that BPA analogues can affect some important biochemical and ultrastructure features of microalgae, promoting ROS production. Lastly, the capability of microalgae to bioaccumulate bisphenols suggest a potential ecotoxicological risk for filter-feeders organisms.

Brassica juncea and the Se-hyperaccumulator Stanleya pinnata exhibit a different pattern of chromium and selenium accumulation and distribution while activating distinct oxidative stress-response signatures.

Soils high in chromium and selenium exist in some countries, like China, India and the US. In the forms of chromate and selenate, these elements can compete during uptake by plants and lead to secondary effects on the absorption of the essential nutrient sulfur. In this study, we evaluated the potential of Brassica juncea and the Se-hyperaccumulator Stanleya pinnata to take-up and store chromium and selenium when applied individually or jointly, the effect on sulfur content, and the plant antioxidant responses. The aim is to advise the best use of these species in phytotechnologies. Plants were grown hydroponically with 50 µM chromate, 50 µM selenate and equimolar concentrations of both elements (50 µM chromate + 50 µM selenate). Our results suggest that B. juncea and S. pinnata possess transport systems with different affinity for chromate and selenate. The joint application of chromate and selenate restricted the accumulation of both elements, but the reduction of selenate uptake by chromate was more evident in B. juncea. On the other hand, selenate decreased chromium accumulation in B. juncea, whereas in S. pinnata such effect was evident only in roots. B. juncea plants stored more chromium and selenium than S. pinnata due to the higher biomass produced, but less selenium when treated with both elements. Chromate and selenate decreased sulfur accumulation in both species, but B. juncea was more sensitive to their toxicity when applied individually, as revealed by increased lipid peroxidation, hydrogen peroxide content in roots and antioxidant enzyme activity. This species can still be efficient for chromium and selenium phytoextraction as these elements in soil are less available than in hydroponics. In soils high in both elements, or low in selenium, S. pinnata is preferred for selenium phytoextraction and the biomass could be used for crop biofortification due its negligible chromium content.

Morpho-physiological responses by Isochrysis galbana Parke to different concentrations of oxytetracycline.

The pollution of aquatic bodies by pharmaceutical compounds is an emerging environmental problem, with little explored consequences. Oxytetracycline (OTC) is an antibiotic used for treatment of infections caused by a variety of microorganisms and it is widely employed in medicine, livestock husbandry and aquaculture. This pharmaceutical compound may cause deleterious effects on non-target aquatic organisms as microalgae. The objective of this study was to evaluate the effects of OTC on growth, pigment content and morpho-physiology of the microalga Isochrysis galbana Parke. The results highlighted that OTC exposure inhibited the growth of I. galbana in cultures treated with OTC 5.0 and 10.0 mg/L after 3 days and in cultures treated with OTC 5.0, 7.5 and 10.0 mg/L after 5 days. Effects of OTC on cells ultrastructure and physiology consisted in large cytoplasmic lipid inclusions and in a decrease of photosynthetic pigments amount.

Selenium biofortification in the 21st century: status and challenges for healthy human nutrition.

Background: Selenium (Se) is an essential element for mammals and its deficiency in the diet is a global problem. Plants accumulate Se and thus represent a major source of Se to consumers. Agronomic biofortification intends to enrich crops with Se in order to secure its adequate supply by people. Scope: The goal of this review is to report the present knowledge of the distribution and processes of Se in soil and at the plant-soil interface, and of Se behaviour inside the plant in terms of biofortification. It aims to unravel the Se metabolic pathways that affect the nutritional value of edible plant products, various Se biofortification strategies in challenging environments, as well as the impact of Se-enriched food on human health. Conclusions: Agronomic biofortification and breeding are prevalent strategies for battling Se deficiency. Future research addresses nanosized Se biofortification, crop enrichment with multiple micronutrients, microbial-integrated agronomic biofortification, and optimization of Se biofortification in adverse conditions. Biofortified food of superior nutritional quality may be created, enriched with healthy Se-compounds, as well as several other valuable phytochemicals. Whether such a food source might be used as nutritional intervention for recently emerged coronavirus infections is a relevant question that deserves investigation.

Bioactivity of Size-Fractionated and Unfractionated Humic Substances From Two Forest Soils and Comparative Effects on N and S Metabolism, Nutrition, and Root Anatomy of Allium sativum L.

Humic substances (HS) are powerful natural plant biostimulants. However, there is still a lack of knowledge about the relationship between their structure and bioactivity in plants. We extracted HS (THE1-2) from two forest soils covered with Pinus mugo (1) or Pinus sylvestris (2). The extracts were subjected to weak acid treatment to produce size-fractionated HS (high molecular size, HMS1-2; low molecular size, LMS1-2). HS were characterized for total acidity, functional groups, element and auxin (IAA) contents, and hormone-like activity. HS concentrations ranging from 0 to 5 mg C L-1 were applied to garlic (Allium sativum L.) plantlets in hydroponics to ascertain differences between unfractionated and size-fractionated HS in the capacity to promote mineral nutrition, root growth and cell differentiation, activity of enzymes related to plant development (invertase, peroxidase, and esterase), and N (nitrate reductase, glutamine synthetase) and S (O-acetylserine sulphydrylase) assimilation into amino acids. A positive linear dose-response relationship was determined for all HS in the range 0-1 mg C L-1, while higher HS doses were less effective or ineffective in promoting physiological-biochemical attributes of garlic. Bioactivity was higher for size-fractionated HS according to the trend LMS1-2>HMS1-2>THE1-2, with LMS2 and HMS2 being overall more bioactive than LMS1 and HMS1, respectively. LMS1-2 contained more N, oxygenated functional groups and IAA compared to THE1-2 and HMS1-2. Also, they exhibited higher hormone-like activities. Such chemical properties likely accounted for the greater biostimulant action of LMS1-2. Beside plant growth, nutrition and N metabolism, HS stimulated S assimilation by promoting the enrichment of garlic plantlets with the S amino acid alliin, which has recognized beneficial properties in human health. Concluding, this study endorses that i) treating THE with a weak acid produced sized-fractionated HS with higher bioactivity and differing in properties, perhaps because of novel molecular arrangements of HS components that better interacted with garlic roots; ii) LMS from forest soils covered with P. mugo or P. sylvestris were the most bioactive; iii) the cover vegetation affected HS bioactivity iv); HS stimulated N and S metabolism with relevant benefits to crop nutritional quality.

Effects on Plant Growth and Reproduction of a Peach R2R3-MYB Transcription Factor Overexpressed in Tobacco.

In plants, anthocyanin production is controlled by MYB and bHLH transcription factors. In peach, among the members of these families, MYB10.1 and bHLH3 have been shown to be the most important genes for production of these pigments during fruit ripening. Anthocyanins are valuable molecules, and the overexpression of regulatory genes in annual fast-growing plants has been explored for their biotechnological production. The overexpression of peach MYB10.1 in tobacco plants induced anthocyanin pigmentation, which was particularly strong in the reproductive parts. Pigment production was the result of an up-regulation of the expression level of key genes of the flavonoid biosynthetic pathway, such as NtCHS, NtCHI, NtF3H, NtDFR, NtANS, and NtUFGT, as well as of the proanthocyanidin biosynthetic pathway such as NtLAR. Nevertheless, phenotypic alterations in transgenic tobacco lines were not only limited to anthocyanin production. Lines showing a strong phenotype (type I) exhibited irregular leaf shape and size and reduced plant height. Moreover, flowers had reduced length of anther's filament, nondehiscent anthers, reduced pistil length, aborted nectary glands, and impaired capsule development, but the reproductive parts including androecium, gynoecium, and petals were more pigmented that in wild type. Surprisingly, overexpression of peach MYB10.1 led to suppression of NtMYB305, which is required for floral development and, of one of its target genes, NECTARIN1 (NtNCE1), involved in the nectary gland formation. MYB10.1 overexpression up-regulated JA biosynthetic (NtAOS) and signaling (NtJAZd) genes, as well as 1-aminocyclopropane-1-carboxylate oxidase (NtACO) in flowers. The alteration of these hormonal pathways might be among the causes of the observed floral abnormalities with defects in both male and female gametophyte development. In particular, approximately only 30% of pollen grains of type I lines were viable, while during megaspore formation, there was a block during FG1 (St3-II). This block seemed to be associated to an excessive accumulation of callose. It can be concluded that the overexpression of peach MYB10.1 in tobacco not only regulates flavonoid biosynthesis (anthocyanin and proanthocyanidin) in the reproductive parts but also plays a role in other processes such as vegetative and reproductive development.

Evidences for interaction of PsbS with photosynthetic complexes in maize thylakoids.

The PsbS subunit of Photosystem II (PSII) has received much attention in the past few years, given its crucial role in photoprotection of higher plants. The exact location of this small subunit in thylakoids is also debated. In this work possible interaction partners of PsbS have been identified by immunoaffinity and immunoprecipitation, performed with mildly solubilized whole thylakoid membrane. The interacting proteins, as identified by mass spectrometry analysis of the immunoaffinity eluate, include CP29, some LHCII components, but also components of Photosystem I, of the cytochrome b(6)f complex as well as of ATP synthase. These proteins can be co-immunoprecipitated by using highly specific anti-PsbS antibodies and, vice-versa, PsbS is co-immunoprecipitated by antisera against components of the interacting complexes. We also find that PsbS co-migrates with bands containing PSII, ATP synthase and cytochrome b(6)f as well as with LHCII-containing bands on non-denaturing Deriphat PAGE. These results suggest multiple location of PsbS in the thylakoid membrane and point to an unexpected lateral mobility of this PSII subunit. As revealed by immunogold labelling with antibody against PsbS, the protein is associated either with granal membranes or prevalently with stroma lamellae in low or high-intensity light-treated intact leaves, respectively. This finding is consistent with the capability of PsbS to interact with complexes located in stroma lamellae, even though the exact physiological condition(s) under which these interactions may take place remain to be clarified.

Evidence for PSII donor-side damage and photoinhibition induced by cadmium treatment on rice (Oryza sativa L.).

The effects of cadmium (from 7.5 to 75 microM) on chloroplasts of rice were studied at the structural and biochemical level. Loss of pigments, reduction of thylakoids and decrease in oxygen evolution and Fv/Fm ratio occur in leaves following cadmium treatment. However, the amount of photosystem II reaction center proteins and that of its light harvesting complex is not affected, indicating that cadmium does not adversely influence the structural organization of this photosystem. In thylakoids isolated from cadmium-treated plants a loss in the capability to reduce 2,6-dichlorophenolindophenol is observed, which is partially restored if diphenylcarbazide is used as an electron donor, indicating that cadmium affects water splitting activity. In thylakoids isolated from control plants and treated with cadmium, diphenylcarbazide preserves most of the photosystem II activity lost after incubation with cadmium; most of the S(2) multiline electron paramagnetic resonance signal from the manganese cluster is lost, whereas the TyrD(+) and other signals are retained. Light-induced photosystem II damage, in vitro, is promoted by Cd-treatment as deduced from the mobility shift of the D1 protein observed by immunoblot.

Comparative effects of selenate and selenite on selenium accumulation, morphophysiology, and glutathione synthesis in Ulva australis.

The capacity of Ulva australis Areschoug to tolerate and accumulate selenium (Se) supplied in the form of selenate or selenite was investigated. The macroalga was provided for 3 and 7 days with concentrations of selenate (Na2SeO4) or selenite (Na2SeO3) ranging from 0 to 400 µM. U. australis exhibited the highest ability to accumulate selenium when fed with 100 µM selenate and 200 µM selenite after 7 days, and accumulation values were respectively 25 and 36 ppm Se. At the same concentrations, stimulation of the synthesis of chlorophylls and carotenoids was observed. Elevated doses of selenate or selenite decreased Se accumulation inside algal cells, perhaps through repression of membrane transporters. This effect was more pronounced in thalli cultivated with selenate. There were no morphological and ultrastructural alterations in thalli exposed to Se. However, selenite induced the increase of the oxidized fraction of glutathione (GSSG), perhaps because of its capacity to bind the thiol group of reduced glutathione (GSH). In conclusion, this study highlights the capacity of U. australis to resist to very high concentrations of selenite and selenate, which are normally toxic to other organisms. Also, the lack of bioconcentration in U. australis indicates that this alga does not facilitate delivery of Se in the food chain and remains safe for consumption when it grows in water bodies contaminated with Se. Its potential for the removal of excess Se from water bodies appears limited.

Accumulation of selenium in Ulva sp. and effects on morphology, ultrastructure and antioxidant enzymes and metabolites.

The impact of selenium (Se) on Ulva sp., a green macroalga naturally growing in the Venice Lagoon, was investigated. The alga was provided for 10 days with concentrations of selenate (Na(2)SeO(4)) ranging from 0 to 100 µM. Se accumulation in the algal biomass was linearly related to the selenate dose and this relationship was not affected by the high sulfate concentration measured in the seawater. The amount of Se measured in the alga was always relatively low and not hazardous to algal consumers. However, Se induced the formation of hydrogen peroxide (H(2)O(2)) in Ulva sp. and, as a result, the activity of antioxidant enzymes (superoxide dismutase, SOD, and catalase, CAT) and the amount of antioxidant metabolites (phenols, flavonoids and carotenoids) increased, even when selenate was supplied to the macroalga at low concentration (2.5 µM). This indicated that different components of the antioxidant defence system played a pivotal role in overcoming oxidative damage by Se in the macroalga, and explained the lack of morphological and ultrastructural alterations in Ulva sp. exposed to selenate.

Involvement of p53 in specific anti-neuroectodermal tumor activity of aloe-emodin.

Previously, we have identified aloe-emodin (AE) as a new type of anticancer agent, with activity that is based on apoptotic cell death promoted by a neuroectodermal tumor-specific drug uptake. We attempt to clarify the intracellular target of AE and the apoptosis-signaling pathway activated by AE in neuroblastoma cell lines. Two-photon excitation microscopy and spectroscopic titrations documented that AE is highly concentrated in susceptible cells and binds to DNA. One of the most important mediators of apoptotic response to genotoxic stimuli, such as anticancer agents, is the p53 tumor suppressor gene. To evaluate the role played by p53 in AE-induced apoptosis a p53 mutant cell line, which lacks transcriptional activity of p53 targeted genes, was tested. AE displayed a reduced growth inhibitory and pro-apoptotic activity in p53 mutant cells (SK-N-BE(2c)) with respect to the p53 wild-type line (SJ-N-KP). This effect was not caused by a reduced drug uptake in the mutant neuroblastoma cell line but was related to a different apoptotic cell phenotype. Whereas SJ-N-KP cells were susceptible to a p53 transcription-dependent pathway of apoptosis, SK-N-BE(2c) cells underwent apoptosis with up-regulation of p53 expression but not of p53-target genes. After AE treatment p53 translocates to the mitochondria inter-membrane space in both neuroblastoma cell lines. Due to its high accumulation in neuroectodermal tumor cells AE could also kill tumor cells harboring p53 mutant genes. This property would further contribute to AE specific anti-tumor activity and might be exploitable in the clinic.