Multi-strategy engineering greatly enhances provitamin A carotenoid accumulation and stability in Arabidopsis seeds

Staple grains with low levels of provitamin A carotenoids contribute to the global prevalence of vitamin A deficiency and therefore are the main targets for provitamin A biofortification. However, carotenoid stability during both seed maturation and postharvest storage is a serious concern for the full benefits of carotenoid biofortified grains. In this study, we utilized Arabidopsis as a model to establish carotenoid biofortification strategies in seeds. We discovered that manipulation of carotenoid biosynthetic activity by seed-specific expression of Phytoene synthase (PSY) increases both provitamin A and total carotenoid levels but the increased carotenoids are prone to degradation during seed maturation and storage, consistent with previous studies of provitamin A biofortified grains. In contrast, stacking with Orange (ORHis), a gene that initiates chromoplast biogenesis, dramatically enhances provitamin A and total carotenoid content and stability. Up to 65- and 10-fold increases of β-carotene and total carotenoids, respectively, with provitamin A carotenoids composing over 63% were observed in the seeds containing ORHis and PSY. Co-expression of Homogentisate geranylgeranyl transferase (HGGT) with ORHis and PSY further increases carotenoid accumulation and stability during seed maturation and storage. Moreover, knocking-out of β-carotene hydroxylase 2 (BCH2) by CRISPR/Cas9 not only potentially facilitates β-carotene accumulation but also minimizes the negative effect of carotenoid over production on seed germination. Our findings provide new insights into various processes on carotenoid accumulation and stability in seeds and establish a multiplexed strategy to simultaneously target carotenoid biosynthesis, turnover, and stable storage for carotenoid biofortification in crop seeds

Clp protease and OR directly control the proteostasis of phytoene synthase, the crucial enzyme for carotenoid biosynthesis in Arabidopsis

Ajuts: This work was supported by Agriculture and Food Research Initiative competitive award no. 2016-67013-24612 from the USDA National Institute of Food and Agriculture and by the HarvestPlus research consortium (2014H6320.FRE)Phytoene synthase (PSY) is the crucial plastidial enzymein the carotenoid biosynthetic pathway. However, its post-translational regulation remains elusive. Likewise, Clp protease constitutes a central part of the plastid protease network, but its substrates for degradation are not well known. In this study, we report that PSY is a substrate of the Clp protease. PSY was uncovered to physically interact with various Clp protease subunits (i.e., ClpS1, ClpC1, and ClpD). High levels of PSY and several other carotenogenic enzyme proteins overaccumulate in the clpc1, clpp4, and clpr1-2 mutants. The overaccumulated PSY was found to be partially enzymatically active. Impairment of Clp activity in clpc1 results in a reduced rate of PSY protein turnover, further supporting the role of Clp protease in degrading PSY protein.On the other hand, the ORANGE (OR) protein, a major post-translational regulator of PSY with holdase chaperone activity, enhances PSY protein stability and increases the enzymatically active proportion of PSY in clpc1, counterbalancing Clp-mediated proteolysis in maintaining PSY proteinhomeostasis. Collectively, these findings provide novel insights into the quality control of plastid-localized proteins and establish a hitherto unidentified post-translational regulatory mechanism of carotenogenic enzymes in modulating carotenoid biosynthesis in plants

Characterization of cassava ORANGE proteins and their capability to increase provitamin A carotenoids accumulation

Cassava (Manihot esculenta Crantz) biofortification with provitamin A carotenoids is an ongoing process that aims to alleviate vitamin A deficiency. The moderate content of provitamin A carotenoids achieved so far limits the contribution to providing adequate dietary vitamin A levels. Strategies to increase carotenoid content focused on genes from the carotenoids biosynthesis pathway. In recent years, special emphasis was given to ORANGE protein (OR), which promotes the accumulation of carotenoids and their stability in several plants. The aim of this work was to identify, characterize and investigate the role of OR in the biosynthesis and stabilization of carotenoids in cassava and its relationship with phytoene synthase (PSY), the rate-limiting enzyme of the carotenoids biosynthesis pathway. Gene and protein characterization of OR, expression levels, protein amounts and carotenoids levels were evaluated in roots of one white (60444) and two yellow cassava cultivars (GM5309-57 and GM3736-37). Four OR variants were found in yellow cassava roots. Although comparable expression was found for three variants, significantly higher OR protein amounts were observed in the yellow varieties. In contrast, cassava PSY1 expression was significantly higher in the yellow cultivars, but PSY protein amount did not vary. Furthermore, we evaluated whether expression of one of the variants, MeOR_X1, affected carotenoid accumulation in cassava Friable Embryogenic Callus (FEC). Overexpression of maize PSY1 alone resulted in carotenoids accumulation and induced crystal formation. Co-expression with MeOR_X1 led to greatly increase of carotenoids although PSY1 expression was high in the co-expressed FEC. Our data suggest that posttranslational mechanisms controlling OR and PSY protein stability contribute to higher carotenoid levels in yellow cassava. Moreover, we showed that cassava FEC can be used to study the efficiency of single and combinatorial gene expression in increasing the carotenoid content prior to its application for the generation of biofortified cassava with enhanced carotenoids levels

The Characterization and SCR Performance of Mn-Containing α-Fe2O3 Derived from the Decomposition of Siderite

In this work, a nano-structured iron-manganese oxide composite was prepared by calcining natural manganese-rich siderite at different temperatures (450, 500, 550, 600 °C, labeled as H450, H500, H550, H600, respectively), and their performances of selective catalytic reduction (SCR) of NO by NH3 were investigated. XRD, XRF, BET, XPS, SEM, and TEM were used to investigate the morphology, composition, and surface characteristics of the catalyst. The results showed that the decomposition of siderite occurred from 450 °C to around 550 °C during the calcination in air atmosphere; moreover, the siderite could be converted into nano-structured α-Fe2O3. The specific surface area of the material increased, and Mn2+ was transformed into Mn4+, which were beneficial to the SCR. Among these catalysts, H550 had the best SCR performance, with NO removal of 98% at a temperature window from 200 to 250 °C. The presence of water vapor and sulfur dioxide can inhibit the SCR performance of the catalysts, but this inhibition effect was not obvious for H550 at the optimum reaction temperature (250 °C). The findings presented in this study are significant toward the application of the Mn-rich siderite as a precursor in preparing the Fe-Mn oxides for catalytic de-NOx by SCR

Synthesis and evaluation of odour-active methionyl esters of fatty acids via esterification and transesterification of butter oil

10.1016/j.foodchem.2013.08.124Food Chemistry145796-801FOCH

Clp protease and OR directly control the proteostasis of phytoene synthase, the crucial enzyme for carotenoid biosynthesis in Arabidopsis

Ajuts: This work was supported by Agriculture and Food Research Initiative competitive award no. 2016-67013-24612 from the USDA National Institute of Food and Agriculture and by the HarvestPlus research consortium (2014H6320.FRE)Phytoene synthase (PSY) is the crucial plastidial enzymein the carotenoid biosynthetic pathway. However, its post-translational regulation remains elusive. Likewise, Clp protease constitutes a central part of the plastid protease network, but its substrates for degradation are not well known. In this study, we report that PSY is a substrate of the Clp protease. PSY was uncovered to physically interact with various Clp protease subunits (i.e., ClpS1, ClpC1, and ClpD). High levels of PSY and several other carotenogenic enzyme proteins overaccumulate in the clpc1, clpp4, and clpr1-2 mutants. The overaccumulated PSY was found to be partially enzymatically active. Impairment of Clp activity in clpc1 results in a reduced rate of PSY protein turnover, further supporting the role of Clp protease in degrading PSY protein.On the other hand, the ORANGE (OR) protein, a major post-translational regulator of PSY with holdase chaperone activity, enhances PSY protein stability and increases the enzymatically active proportion of PSY in clpc1, counterbalancing Clp-mediated proteolysis in maintaining PSY proteinhomeostasis. Collectively, these findings provide novel insights into the quality control of plastid-localized proteins and establish a hitherto unidentified post-translational regulatory mechanism of carotenogenic enzymes in modulating carotenoid biosynthesis in plants

New Synthesis of nZVI/C Composites as an Efficient Adsorbent for the Uptake of U(VI) from Aqueous Solutions

New nanoscale zerovalent iron/carbon (nZVI/C) composites were successfully prepared via heating natural hematite and pine sawdust at 800 °C under nitrogen conditions. Characterization by SEM, XRD, FTIR, and XPS analyses indicated that the as-prepared nZVI/C composites contained a large number of reactive sites. The lack of influence of the ionic strength revealed inner-sphere complexation dominated U­(VI) uptake by the nZVI/C composites. Simultaneous adsorption and reduction were involved in the uptake process of U­(VI) according to the results of XPS and XANES analyses. The presence of U–C/U-U shells demonstrated that innersphere complexation and surface coprecipitation dominated the U­(VI) uptake at low and high pH conditions, respectively. The uptake behaviors of U­(VI) by the nZVI/C composites were fitted well by surface complexation modeling with two weak and two strong sites. The maximum uptake capacity of U­(VI) by the nZVI/C composites was 186.92 mg/g at pH 4.0 and 328 K. Additionally, the nZVI/C composites presented good recyclability and recoverability for U­(VI) uptake in regeneration experiments. These observations indicated that the nZVI/C composites can be considered as potential adsorbents to remove radionuclides for environmental remediation