The involvement of amino acids in latex lipid synthesis in Euphorbia lathyris seedlings (original) (raw)
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Distinct triterpene synthases in the laticifers of Euphorbia lathyris
Scientific Reports
Euphorbia lathyris was proposed about fifty years ago as a potential agroenergetic crop. The tremendous amounts of triterpenes present in its latex has driven investigations for transforming this particular biological fluid into an industrial hydrocarbon source. The huge accumulation of terpenes in the latex of many plant species represent a challenging question regarding cellular homeostasis. In fact, the enzymes, the mechanisms and the controllers that tune the amount of products accumulated in specialized compartments (to fulfill ecological roles) or deposited at important sites (as essential factors) are not known. Here, we have isolated oxidosqualene cyclases highly expressed in the latex of Euphorbia lathyris. this triterpene biosynthetic machinery is made of distinct paralogous enzymes responsible for the massive accumulation of steroidal and non-steroidal tetracyclic triterpenes. More than eighty years after the isolation of butyrospermol from shea butter (Heilbronn IM, Moffet GL, and spring Fs J. Chem. Soc. 1934, 1583), a butyrospermol synthase is characterized in this work using yeast and in folia heterologous expression assays. Euphorbia lathyris is a herbaceous plant native to the Mediterranean area and widespread in temperate regions. Its vascular tissues show a dense network of accompanying laticifers 1. This is also the case of all Euphorbia species and other plants from the Euphorbiaceae family, including the popular rubber tree Hevea brasiliensis 2 , and species in several plant families like the Moraceae, Apocynaceae, and Papaveraceae 3,4. A major trait of these plants is the massive accumulation of specialized metabolites in the laticifers, like for instance the tremendous amounts of the alkaloids morphine and codeine found in the laticifers of opium poppy 5,6. Laticifers in many Euphorbiaceae grow and form anastomoses throughout the plant from the embryo stage 7,8. The cytoplasm of these coenocytic cells is a milky fluid called latex that contains heavy loads of isoprenoids: cis-1,4-polyisoprene otherwise known as natural rubber in H. brasiliensis, and very often high concentrations of diterpenes or triterpenes in Euphorbia species 9,10. The biosynthetic pathway of E. lathyris seed oil diterpenoids (called euphorbia factors) has been fully described from the casbene precursor to the bioactive macrocyclic diterpene derivatives such as ingenol 11. The genus Euphorbia, comprising almost 2000 species, has been in the focus of chemotaxonomic studies from the early sixties. This was mostly inspired by the challenge of establishing phylogenetic relationships based on architectural, morphological, anatomical, physiological and chemical traits available at that time. Whether the observed chemical diversification acts as a driving force in morphological evolution remains unclear so far. At least the coincidental occurrence of a very specific triterpene signature with a particular set of morphological traits was observed in some cases, despite of the complex geographical distribution of Euphorbia species. These pioneering phytochemical surveys have revealed the triterpene (C 30 H 50 O) skeletal diversity in the latex of Euphorbia species 12. Euphorbia lathyris contains mostly lanostane (lanosterol, cycloartenol and their C24-methylated derivatives), euphane (butyrospermol, euphol), and hopane (hopenol-B) derivatives in its latex 13,14. The presence of a triterpene of the bacterial-type, namely, hopenol-B, was also found in Euphorbia supina 15. In reality, the chemical composition of the latex of Euphorbia lathyris is much more diverse 16 , and several studies have reported the presence of other substances than terpenoids like for instance L-dopa 17. Preceding the more recent interest in Euphorbia spp for pharmacologically active compounds including terpenoids 11 , Euphorbia lathyris was proposed about fifty years ago as a potential agroenergetic crop by Melvin Calvin 18,19. In fact, Euphorbia lathyris latex is
Serine in sterol synthesis in Euphorbia lathyris seedlings
Phytochemistry, 1986
During germination scrine was shown to lx taken up from the endosperm by the cotykdons and partly translocatcd to the hypocotyl. Up to 1.7Y/, of the ["CJscrinc taken up by the seedling was involved in phytostcrol synthais in which the hypocotyl was the most &ctive plant part. Compared with [ 14C]sucrose. scrinc was found to bc 12 times more effective in this synthesis. From the data obtained it could bc caRtdated that this ammo acid may yield about 7 Ye of ail the free stcrols in an ctiolatcd seedling of Eupburbia futhyris, making this substrate a suitable marker in steroi synthesis in endospermous szcdlings.
Plant physiology, 1989
A latex pellet fraction from Euphorbia lathyris incorporates mevalonate into triterpenols and their fatty acid esters. Conditions for improved incorporation were determined. CaCl(2) or CaCl(2) plus MnCl(2) stimulated biosynthesis, and the metal ion chelator, ethylenebis(oxyethylenenitrilo)tetraacetic acid (EGTA) enhanced stimulation. Ethylenediaminetetraacetic acid was almost as effective as EGTA, but phthalic acid and citric acid were relatively poor stimulators. The concentration of the Ca(2+)-EGTA complex was directly measured, and the incorporation data are best fitted by a curve that shows that the receptor for the complex is saturable. In the presence of the metal-chelate complex, the addition of fructose, 1,6-bisphosphate plus aldolase (triose-P) or malate provided additional stimulation. Incorporation was maximum at 40 micromolar R-mevalonate, and inhibition occurred at higher concentrations. The apparent K(m) for R-mevalonate was 15 micromolar. Under improved reaction condi...
Sterols and triterpenols in latex and cultured tissues of Euphorbia pulcherrima
Phytochemistry, 1982
Key Word Index-Euphohia pulchenima; Euphorbiaceae; tissue culture; sterols; triterpenols. Abatrad-The sterol and triterpenol constituents of Euphorbi, pufchewima latex and cultured callus tissues were examined by GLC and mass spectrometry. Latex extracts from different varieties contained sitosterol, B-amyrin, germanicol, cycloartenol, /3-amyrin acetate, and germanicol acetate. Capillary GC proflles of these varieties indicated that the triterpene content was essentially identical for examined latices. Cultured tissues derived from petioles and stem internodes synthesized only sitosterol in significant quantities, although trace amounts of several sterols that occur in latex were also detected in cultured tissues. This study supports the interpretation that the pattern of triterpene synthesis in the laticifer of the normal plant is a highly controlled and stable phenomenon among varieties of this species.
Lipolytic isoenzymes from Euphorbia latex
Plant Science, 2003
The activity and substrate specificity of latex lipases from Euphorbia species (E. characias , E. wulfenii , E. pinea , E. myrsinites and E. dendroides ) were investigated. High lipolytic activity was found only in E. characias and for the first time in E. wulfenii latex. For both species the lipolytic activity on various triglycerides, and under different temperature and pH conditions, in both crude latex and in partially purified enzymes was quantified. Optimised extraction and purification methods permitted the recovery of the enzymatic fraction with high lipolytic activity. This fraction is probably constituted by a pool of different lipolytic enzymes. Finally, lipolytic activity was also measured for E. characias and E. wulfenii during vegetative and reproductive stages. #
Revista Brasileira de Fisiologia Vegetal, 2000
Seed composition and reserve mobilization were investigated in wild poinsettia (Euphorbia heterophylla L.). Lipids, around 60% of seed dry mass, are the major reserve. Proteins, including albumins (49%), salt insoluble globulins (30%), salt-soluble globulins (21%) and prolamins (0.3%), comprise about a quarter of seed dry mass. Soluble sugars comprise about 3.6% of seed dry mass, sucrose being the predominant sugar. Starch was not detected in the endosperm of E. heterophylla. Lipid depletion starts after initial imbibition, and is completed between 72 and 96 hours. Protein fractions exhibit different degradation patterns, salt-soluble globulins being continuously degraded after the start of imbibition whereas salt insoluble fractions are degraded between 36 and 72 hours, and albumins between 60 and 84 hours. Globulin depletion is accompanied by an increase in free amino acids in the endosperm whereas intense albumin depletion is not. This result suggests that during albumin depletion there is a rapid transfer of amino acids to the growing embryo. Histochemical studies indicated that light accelerates protein degradation in the micropylar area of the seed. Soluble sugars increase in the embryo with no concomitant decrease in the endosperm, suggesting that sugars are mostly originated from the catabolism of lipids.
Occurrence of fatty acid lower-alkyl esters in euonymus fruits
Small amounts of a mixture of fatty acid lower alkyl esters (FALAEs) were obtained from chloro form extracts of the fruit arils and seeds of four euonymus species (Euonymus sp., Celastraceae). The FALAEs were the products of biosyntheses in the cell rather than experimental artifacts. By using GC MS, this mix ture was shown to contain a total of 19 individual FALAE species comprising four separate fractions, viz. methyl (FAMEs), ethyl (FAEEs), n propyl, and n butyl FA esters. Fruit FALAEs included mainly FAEEs and, to a lesser extent, FAMEs, while the n propyl and n butyl FA esters, which occurred less frequently, were found here for the first time as the plant products. The FALAE acid components included C 14 -C 18 saturated, mono , di , and trienoic FAs with the predominance of ubiquitous linoleic, oleic, palmitic, and, in some cases, also α linolenic acid. The indices of qualitative and quantitative composition of separate FALAE frac tions varied considerably depending on the plant species, fruit part (aril or seed), and the extent of fruit matu rity. It can be supposed that, in some euonymus species, FAMEs and FAEEs are formed at the expense of the same FA pool characteristic for a given species. As a whole, euonymus FALAEs and triacylglycerols seem to be synthesized from different FA pools. Discussed is the physiological significance of FALAE biosynthesis in plant metabolism, possible pathways of this biosynthesis, as well as the perspectives of further investigations of FALAEs of plant origin.
Journal of Experimental Botany, 2009
A comparative study was carried out on the dynamics of lipid accumulation in developing seeds of three lupine species. Lupine seeds differ in lipid content; yellow lupine (Lupinus luteus L.) seeds contain about 6%, white lupine (Lupinus albus L.) 7-14%, and Andean lupine (Lupinus mutabilis Sweet) about 20% of lipids by dry mass. Cotyledons from developing seeds were isolated and cultured in vitro for 96 h on Heller medium with 60 mM sucrose (+S) or without sucrose (-S). Each medium was additionally enriched with 35 mM asparagine or 35 mM NaNO 3 . Asparagine caused an increase in protein accumulation and simultaneously decreased the lipid content, but nitrate increased accumulation of both protein and lipid. Experiments with [1-14 C]acetate and [2-14 C]acetate showed that the decrease in lipid accumulation in developing lupine seeds resulted from exhaustion of lipid precursors rather than from degradation or modification of the enzymatic apparatus. The carbon atom from the C-1 position of acetate was liberated mainly as CO 2 , whereas the carbon atom from the C-2 position was preferentially used in anabolic pathways. The dominant phospholipid in the investigated lupine seed storage organs was phosphatidylcholine. The main fatty acid in yellow lupine cotyledons was linoleic acid, in white lupine it was oleic acid, and in Andean lupine it was both linoleic and oleic acids. The relationship between stimulation of lipid and protein accumulation by nitrate in developing lupine cotyledons and enhanced carbon flux through glycolysis caused by the inorganic nitrogen form is discussed.