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Characterization of two genes for the biosynthesis of the labdane diterpene Z ‐abienol in tobacco ( Nicotiana tabacum) glandular trichomes
Author(s) -
Sallaud Christophe,
Giacalone Cécile,
Töpfer Romy,
Goepfert Simon,
Bakaher Nicolas,
Rösti Sandrine,
Tissier Alain
Publication year - 2012
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/j.1365-313x.2012.05068.x
Subject(s) - trichome , nicotiana tabacum , diterpene , biology , nicotiana , biosynthesis , gene , farnesyl diphosphate synthase , botany , ectopic expression , solanaceae , biochemistry
Summary Leaves of tobacco ( Nicotiana tabacum ) are covered with glandular trichomes that produce sucrose esters and diterpenoids in varying quantities, depending on cultivar type. The bicyclic diterpene Z ‐abienol is the major labdanoid present in some oriental tobacco cultivars, where it constitutes a precursor of important flavours and aromas. We describe here the identification and characterization of two genes governing the biosynthesis of Z ‐abienol in N. tabacum . As for other angiosperm labdanoid diterpenes, the biosynthesis of Z ‐abienol proceeds in two steps. NtCPS2 encodes a class‐II terpene synthase that synthesizes 8‐hydroxy‐copalyl diphosphate, and NtABS encodes a kaurene synthase‐like (KSL) protein that uses 8‐hydroxy‐copalyl diphosphate to produce Z ‐abienol. Phylogenetic analysis indicates that NtABS belongs to a distinct clade of KSL proteins that comprises the recently identified tomato ( Solanum habrochaites ) santalene and bergamotene synthase. RT‐PCR results show that both genes are preferentially expressed in trichomes. Moreover, microscopy of NtCPS2 promoter‐GUS fusion transgenics demonstrated a high specificity of expression to trichome glandular cells. Ectopic expression of both genes, but not of either one alone, driven by a trichome‐specific promoter in transgenic Nicotiana sylvestris conferred Z ‐abienol formation to this species, which does not normally produce it. Furthermore, sequence analysis of over 100 tobacco cultivars revealed polymorphisms in NtCPS2 that lead to a prematurely truncated protein in cultivars lacking Z ‐abienol, thus establishing NtCPS2 as a major gene controlling Z ‐abienol biosynthesis in tobacco. These results offer new perspectives for tobacco breeding and the metabolic engineering of labdanoid diterpenes, as well as for structure–function relationship studies of terpene synthases.