Bouvier-Navé P, Husselstein T, Benveniste P
Institut de Biologie Moléculaire des Plantes, Département de Biologie Cellulaire et Moléculaire, Strasbourg, France.
Eur J Biochem. 1998 Aug 15;256(1):88-96. doi: 10.1046/j.1432-1327.1998.2560088.x.
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, smtl and smt2; in addition, smt2 cDNAs were shown to encode a 24-methylene lophenol C24 methyltransferase [Bouvier-Navé, P., Husselstein, T., Desprez, T. & Benveniste, P. (1997) Eur. J. Biochem. 246, 518-529]. 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 Ntsm1-1 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 methyltransferase. Meanwhile, two cDNAs were isolated from Oriza sativa and shown to belong to smtl 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.
24-甲基和24-乙基甾醇的生物合成涉及两次甲基转移，这两种甾醇在植物生长发育中起主要作用。第一次甲基转移作用于环阿屯醇，第二次作用于24-亚甲基环菠萝醇。到目前为止，已从多种植物中分离出约十个编码S-腺苷-L-甲硫氨酸（AdoMet）甾醇甲基转移酶（SMT）的cDNA克隆。根据其推导的氨基酸序列，它们被分为两个家族，即smtl和smt2；此外，smt2 cDNA显示编码一种24-亚甲基环菠萝醇C24甲基转移酶[Bouvier-Navé, P., Husselstein, T., Desprez, T. & Benveniste, P. (1997) Eur. J. Biochem. 246, 518 - 529]。我们现在报告从烟草中分离出的两个cDNA的比较，Ntsmt1-1属于第一个SMT cDNA家族，Ntsmt2-1属于第二个家族。这两个cDNA都在缺乏SMT的酵母缺失突变体erg6中表达。虽然erg6不含24-烷基甾醇，但erg6 Ntsmt1-1含有大部分24-亚甲基甾醇，而erg6 Ntsmt2-1含有大部分24-亚乙基甾醇，这表明这些cDNA的表达产物具有不同的功能。在存在AdoMet的情况下，来自erg6 Ntsm1-1的脱脂微粒体有效地将环阿屯醇转化为24-亚甲基环阿屯醇，但与24-亚甲基环菠萝醇一起孵育时不会产生任何24-亚乙基环菠萝醇。这表明cDNA Ntsmt1-1（很可能还有第一个家族的其他植物SMT cDNA）编码一种环阿屯醇C24甲基转移酶。相反，erg6 Ntsmt2-1的脱脂微粒体显示优先甲基化24-亚甲基环菠萝醇，正如smt2 cDNA编码的SMT所预期的那样。总之，在从烟草中分离出的各种cDNA中，一个（Ntsmt1-1）根据其推导的氨基酸序列属于植物SMT cDNA的第一个家族，并被证明编码一种环阿屯醇C24甲基转移酶，而另一个（Ntsmt2-1）属于第二个家族，并被证明编码一种24-亚甲基环菠萝醇C24甲基转移酶。同时，从水稻中分离出两个cDNA，分别显示属于smtl和smt2家族。这些数据揭示了在一个特定植物物种中两种不同的SMT共存，每种SMT催化甾醇生物合成途径中的一步甲基化反应。
