Isolation and Characterization of Mutants of Thiophene Synthesis in Tagetes erecta

Abstract

Two mutants of Tagetes erecfa displaying aberrant thiophene composition were identified by screening more than 300 plants from a mutagenized M, population using high-performance liquid chromatography analysis of root extracts. Both mutants, which may have originated from the same mutational event, contained high amounts of the C,, monothiophene 2-(but-3-en-I -ynyl)-5-(penta-1,3-diynyl)-thiophene that was previously not found in T. erecfa and also high amounts of two C,, bithienyls that were absent or present at low concentrations in the wild type. The mutant phenotype was also expressed in 21 Agrobacterium rhizogenes transformed root clones derived from both mutants. Feeding experiments with root cultures derived from one mutant and from the wild type indicated that the monothiophene accumulating in the mutant is the common precursor for all bithienyl thiophenes in wild-type and mutant Tagefes erecfa. These experiments also showed that one mutant is deficient in demethylation of the monothiophene.Although mutants have been widely applied to investigate metabolic pathways in microorganisms and fungi, they have been used relatively rarely for this purpose in plants. Most metabolic mutants of plants affect primary metabolic processes, such as carbon assimilation (Somerville and Ogren, 1979) and metabolism of starch (Caspar et al., 1985), lipids (Browse et al., 1985;Kunst et al., 1989), and amino acids (Haughn and Somerville, 1986;Kreps and Town, 1991;Frankard et al., 1992;Wright et al., 1992;Wu and King, 1994), whereas only a few mutants of secondary metabolic pathways are known (Haughn et al., 1991;Chapple et al., 1992; Rathjen and Robinson, 1992). This situation exists in spite of the fact that chemically induced loss-of-function mutations can be obtained relatively easily (Haughn and Somerville, 1987).Tugetes species (marigolds) produce, mainly in the roots, aromatic sulfur-containing compounds known as thiophenes, which are toxic to nematodes when ingested Bijloo, 1958, 1959). Co-occurrence patterns and precursor-feeding experiments in related species have led to a generally accepted biogenetic scheme, as depicted in Figure 1 (Bohlmann et al., 1973;Bohlmann and Zdero, 1985). Oleic acid is converted into PYE via repeated steps of desaturation and chain shortening. PYE is then converted into the thiophenes that accumulate in Tagetes * Corresponding author; fax 31-80-553450.species. (Bohlmann and Berger, 1965;Bohlmann and Hinz, 1965;Schulte et al., 1965;Bohlmann et al., 1966). The key step in this conversion is the addition of H,S or its biochemical equivalent to conjugated triple bonds and subsequent ring formation, which is probably a two-step reaction (Bohlmann et al., 1973). In addition to the formation of two or three thiophene rings, remova1 of a terminal methyl group and modification of a vinyl group are necessary to obtain the various thiophenes that finally accumulate (Fig. 1).Not much is known about the order in which these reactions take place in Tugetes or about the specificity of the enzymes involve...

Fungal responsive fatty acid acetylenases occur widely in evolutionarily distant plant families

“…These include acetylenic thiophenes such as those associated with the insecticidal and nematicidal properties of Tagetes species (e.g. marigold) (Jacobs et al ., 1995; Towers et al ., 1997).…”

Thiophene acetylenes and furanosesquiterpenes from Xanthopappus subacaulis and their antibacterial activities

“…The other thirteen known compounds were identified as xanthopappin B (4) (Tian et al, 2006), 5-(1,2-dihydroxyethyl)-2-(E)-hept-5-ene-1,3-diynylthiophene (6) (Tian et al, 2006), trichothecolone (13) (Loukaci et al, 2000), dibenzylbutyrolactone (14) (Ward et al, 1996), 6b,11a,20(R)-trihydroxypregna-4-en-3-one (15) (Farooq et al, 1994), rosenonolactone (16) (Loukaci et al, 2000), (23Z)-cycloart-23-ene-3b, 25-dihydroxy (17) (Kitajima et al, 1998), cytochalasin B (18) (Aldridge et al, 1967), luteolin (He and Wang, 1997), and oleic acid (23) (Jacobs et al, 1995). Their structures were confirmed by comparison of their 1 H NMR, 13 C NMR and MS data with the corresponding authentic sample or literature data.…”

“…On the basis of the above work, a plausible biosynthetic pathway for thiophene acetylenes 1-10 is proposed (Scheme 1). Oleic acid is transformed into the key precursor A via repeated steps of desaturation and chain-shortening (Jacobs et al, 1995). Precursor A is then converted into the thiophene acetylenes (Ia and Ib) that accumulate in X. subacaulis; the key step in this conversion is the addition of H 2 S or its biochemical equivalent to conjugated triple bonds, which is most likely a two-step reaction .…”

Tagetes erecta

“…Two mutants of Tagetes erecta were found to contain high amounts in the root extracts of the C13 monothiophene 2-(but-3-en-1-ynyl)-5-(penta-1,3-diynyl)-thiophene that was previously not found in T. erecta and also high amounts of two C13 bithienyls that were absent or present at low concentrations in the wild type (Jacobs et al 1995 ). The mutant phenotype was also expressed in 21 Agrobacterium rhizogenes transformed root clones derived from both mutants.…”