Transformation of Saccharomyces cerevisiae with a cDNA encoding a sterol C‐methyltransferase from Arabidopsis thaliana results in the synthesis of 24‐ethyl sterols

Husselstein, Tania; Gachotte, Daniel; Desprez, Thierry; Bard, Martin; Benveniste, Pierre

doi:10.1016/0014-5793(96)00089-0

Cited by 55 publications

(40 citation statements)

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“…The sterols of erg6 Ntsmt1-1 were identified as follows. GC-MS of acetates of lanosterol (VI), zymosterol (IX), 5A-cholesta-7,24-dien-3β-ol (X), fecosterol (XVI), 5A-stigmasta-8, Z-24(24 1 )-dien-3β-ol (XVIII) and ergosterol (XIX) were similar to those described previously [16]. GC-MS of acetates of 4,4,14A-trimethyl-5A-ergosta-8,24(24 1 )-dien-3β-ol (eburicol, XI), 4,4,14A-trimethyl-5A-stigmasta-8, Z-24(24 1 )-dien-3β-ol (XIIZ), 4,4-dimethyl-5A-ergosta-8,24(24 1 )-dien-3β-ol (XIV) and 5A-ergosta-7,24(24 1 )-dien-3β-ol (episterol, XVII), were similar to those described in [17].…”

Section: Methodssupporting

confidence: 81%

“…This comparison confirms that in both tobacco and rice there are two types of SMTs. The first ones (encoded by Ntsmt1-1, Ntsmt1-2 and Ossmt1-1) are more than 80% identical with already identified SMTs from R. communis [17], G. max [16] and Z. mays [18]. The second ones (encoded by Ntsmt2-1 and Ossmt2-1) are 67 to 80% identical with already identified SMTs from A. thaliana cDNAs Atsmt2-1 (previously named cDNA 411) and a Lanosterol ϭ 4,4,14A-trimethyl-5A-cholesta-8,24-dien-3β-ol (VI); eburicol ϭ 4,4,14A-trimethyl-5A-ergosta-8,24(24 1 )-dien-3β-ol (XI); zymosterol ϭ 5A-cholesta-8,24-dien-3β-ol (IX); fecosterol ϭ 5A-ergosta-8,24(24 1 )-dien-3β-ol (XVI); episterol ϭ 5A-ergosta-7,24(24 1 )-dien-3β-ol (X); ergosterol ϭ 5A-ergosta-5,7,E-22-trien-3β-ol (XX).…”

Section: Resultsmentioning

confidence: 55%

“…Both sterols were methylated, but the catalytic efficiency of the expressed SMT was 17-times higher with 24-methylene lophenol than with cycloartenol [17]. These results [16,17] provide evidence that the A. thaliana cDNA Atsmt2-1 (and most probably the smt2 cDNAs in general) encode(s) a 24-methylene lophenol C24 1 methyltransferase, thus catalyzing the second methylation step of plant sterol biosynthesis.…”

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confidence: 75%

“…2). Previous reports showed the presence in various plants of either one cDNA [15,16,18] or two cDNAs [17] belonging to only one cDNA family. Tobacco belongs to solanaceae (dicotyledones), whereas rice belongs to poaceae (monocotyledones) and they are quite remote taxons.…”

Section: Discussionmentioning

confidence: 99%

“…The deduced sequences from cDNAs of the first family (smt1) are closer to that of ERG6 than those of the second family (smt2) [17,18]. An Arabidopsis thaliana cDNA belonging to the second family has been thoroughly studied [16,17]. Previously called cDNA 411 and now Atsmt2-1, it was used to transform the yeast mutant erg6, the SMT gene of which is disrupted [11].…”

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confidence: 99%

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Two families of sterol methyltransferases are involved in the first and the second methylation steps of plant sterol biosynthesis

Bouvier-Navé¹,

Husselstein²,

Benveniste³

1998

European Journal of Biochemistry

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Two methyl transfers are involved in the biosynthesis of 24-methyl and 24-ethyl sterols, which play major roles in plant growth and development. The first methyl transfer applies to cycloartenol, the second to 24-methylene lophenol. About ten cDNA clones encoding S-adenosyl-L-methionine (AdoMet) sterol methyltransferases (SMTs) have been isolated so far from various plants. According to their deduced amino acid sequences, they were classified in two families, smt1 and smt2; in addition, smt2 cDNAs were shown to encode a 24-methylene lophenol C24 1 methyltransferase [Bouvier-Navé, P., Husselstein, T., Desprez, T. & Benveniste, P. (1997) Eur. J. Biochem. 246, 518Ϫ529]. We now report the comparison of two cDNAs isolated from Nicotiana tabacum, Ntsmt1-1 which belongs to the first SMT cDNA family and Ntsmt2-1 which belongs to the second. Both cDNAs were expressed in the yeast null mutant erg6, deficient in SMT. Whereas erg6 is devoid of 24-alkyl sterols, erg6 Ntsmt1-1 contained a majority of 24-methylene sterols and erg6 Ntsmt2-1, a majority of 24-ethylidene sterols, indicating distinct functions for the expression products of these cDNAs. In the presence of AdoMet, delipidated microsomes from erg6 Ntsmt1-1 efficiently converted cycloartenol into 24-methylene cycloartanol, but did not produce any 24-ethylidene lophenol upon incubation with 24-methylene lophenol. This demonstrates that cDNA Ntsmt1-1 (and most probably the other plant SMT cDNAs of the first family) encode(s) a cycloartenol C24 methyltransferase. In contrast, delipidated microsomes of erg6 Ntsmt2-1 were shown to methylate preferentially 24-methylene lophenol, as expected from an SMT encoded by an smt2 cDNA. In summary, among various cDNAs isolated from N. tabacum, one (Ntsmt1-1) belongs to the first family of plant SMT cDNAs according to its deduced amino acid sequence and was shown to encode a cycloartenol C24 methyltransferase, whereas another (Ntsmt2-1) belongs to the second family and was shown to encode a 24-methylene lophenol C24 1 methyltransferase. Meanwhile, two cDNAs were isolated from Oriza sativa and shown to belong to smt1 and to smt2 families, respectively. These data disclose the coexistence, in a given plant species, of two distinct SMTs, each catalyzing one step of methylation in the sterol biosynthesis pathway.Keywords : plant sterol methyltransferase; yeast transformation; complementation analysis ; cycloartenol; 24-methylene lophenol.Sterols from higher plants and fungi differ from vertebrate sterols by the presence of an extra alkyl group at C24 [1,2]. The alkylation of the side chain of sterols is catalyzed by Sadenosyl-L-methionine (AdoMet) sterol C24 methyltransferases (SMTs). The presence in higher plants of 24-ethyl sterols corresponds to two distinct methyl transfers from AdoMet [1,2]. In agreement with the chemical structures of intermediates of plant sterol biosynthesis, it was shown by substrate-specificity studies that cycloartenol (I) is the substrate of the first methylation reaction, resulting in 24-methylene cycloartan...

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Section: Methodssupporting

confidence: 81%

Section: Resultsmentioning

confidence: 55%

mentioning

confidence: 75%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Two families of sterol methyltransferases are involved in the first and the second methylation steps of plant sterol biosynthesis

Bouvier-Navé¹,

Husselstein²,

Benveniste³

1998

European Journal of Biochemistry

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Biochemistry of Terpenoids: Monoterpenes, Sesquiterpenes, Diterpenes, Sterols, Cardiac Glycosides and Steroid Saponins

Gershenzon

Kreis

Roberts³

et al. 2018

Annual Plant Reviews Online

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Biochemistry of Sterols, Cardiac Glycosides, Brassinosteroids, Phytoecdysteroids and Steroid Saponins

Kreis

Müller‐Uri

2018

Annual Plant Reviews Online

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Phytosterols are synthesized via the mevalonate pathway of terpenoid formation and arise from the initial cyclization of 3 S ‐squalene‐2,3‐epoxide. Plant steroids are derived from sterols and comprise steroid saponins, steroid alkaloids, pregnanes, androstanes, estranes, ecdysteroids, withanolides and cardiac glycosides. The typical route of sterol and steroid biosynthesis follows the cycloartenol pathway, whereas the lanosterol route seems to be operative mainly in fungi and animals. It was demonstrated, however, that both sterol pathways can be operative in higher plants. Crucial steps in the conversion of cycloartenol/lanosterol to sterols are the events leading to the removal of the methyl groups at C‐4 and C‐14. Meanwhile, all steps in the sterol pathway have been elucidated and the respective enzymes/genes characterized. The biosynthetic pathway leading from phytosterol precursors to the cardiac glycosides – important compounds in the treatment of cardiac insufficiency in humans – was basically deduced from studies using radiolabelled precursors. The more recent identification and characterization of several enzymes/genes involved in pregnane and cardenolide metabolism, such as 3β‐hydroxysteroid dehydrogenase and progesterone 5β‐reductase, have further clarified the pathway. Brassinosteroids (BRs) are hydroxylated derivatives of cholestane and they are specific plant steroid hormones that are essential for normal plant development. The biosynthesis of BRs has mainly been studied in Arabidopsis thaliana . Many of the genes encoding biosynthetic enzymes have been cloned using mutants of Arabidopsis thaliana , pea, tomato and rice which revert to a wild‐type phenotype following treatment with exogenous BRs. Phytoecdysteroids are related in structure to the invertebrate steroid hormones. Their biological significance in plants is still under discussion. The understanding of the biosynthetic pathway(s) for phytoecdysteroids is very limited. Steroid saponins constitute a vast group of glycosides present almost exclusively in the monocotyledonous angiosperms, and occurring in only a few dicotyledonous families. As far as enzymatic and genetic aspects are concerned, the biosynthesis of steroid saponins (including the steroid alkaloids) has not been studied extensively. The withanolides are C 28 ‐steroids and biogenetically related to the steroid saponins in that they are derived from ergostane‐type sterols. These compounds appear to be specific for the Solanaceae and their biosynthesis has not yet been studied at the enzyme/gene level.

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Transformation of Saccharomyces cerevisiae with a cDNA encoding a sterol C‐methyltransferase from Arabidopsis thaliana results in the synthesis of 24‐ethyl sterols

Cited by 55 publications

References 28 publications

Two families of sterol methyltransferases are involved in the first and the second methylation steps of plant sterol biosynthesis

Two families of sterol methyltransferases are involved in the first and the second methylation steps of plant sterol biosynthesis

Biochemistry of Terpenoids: Monoterpenes, Sesquiterpenes, Diterpenes, Sterols, Cardiac Glycosides and Steroid Saponins

Biochemistry of Sterols, Cardiac Glycosides, Brassinosteroids, Phytoecdysteroids and Steroid Saponins

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