cis-Jasmone, or (Z)-jasmone, is well known as a component of plant volatiles, and its release can be induced by damage, for example during insect herbivory. Using the olfactory system of the lettuce aphid to investigate volatiles from plants avoided by this insect, (Z)-jasmone was found to be electrophysiologically active and also to be repellent in laboratory choice tests. In field studies, repellency from traps was demonstrated for the damson-hop aphid, and with cereal aphids numbers were reduced in plots of winter wheat treated with (Z)-jasmone. In contrast, attractant activity was found in laboratory and wind tunnel tests for insects acting antagonistically to aphids, namely the seven-spot ladybird and an aphid parasitoid. When applied in the vapor phase to intact bean plants, (Z)-jasmone induced the production of volatile compounds, including the monoterpene (E)-␤-ocimene, which affect plant defense, for example by stimulating the activity of parasitic insects. These plants were more attractive to the aphid parasitoid in the wind tunnel when tested 48 h after exposure to (Z)-jasmone had ceased. This possible signaling role of (Z)-jasmone is qualitatively different from that of the biosynthetically related methyl jasmonate and gives a long-lasting effect after removal of the stimulus. Differential display was used to compare mRNA populations in bean leaves exposed to the vapor of (Z)-jasmone and methyl jasmonate. One differentially displayed fragment was cloned and shown by Northern blotting to be up-regulated in leaf tissue by (Z)-jasmone. This sequence was identified by homology as being derived from a gene encoding an ␣-tubulin isoform.

Alate and apterous virginoparae ofAphis fabae Scop, and alate virginoparae ofBrevicoryne brassicae (L.), walking in a linear track olfactometer, were attracted by odor from leaves of their host plants.A. fabae responded to odor from undamaged but not damaged bean leaves. Gynoparae (autumn migrants) ofA. fabae, however, did not respond to their host plant (spindle,Euonymus europaeus) odor. Odors of certain nonhost plants masked the attractiveness of the host plant leaves, but tansy (Tanacetum vulgare) and summer savory (Satureja hortensis) volatiles repelledB. brassicae andA. fabae, respectively. 3-Butenyl isothiocyanate attractedB. brassicae andLipaphis erysimi (Kalt.), the latter species being more sensitive in both behavioral and electrophysiological studies. Isothiocyanate receptors were found on the antennae ofA. fabae, which was repelled by these compounds, 4-pentenyl isothiocyanate being the most active.

The major class of glucosinolates in Arabidopsis thaliana (L.) Heynh. are biosynthesized from methionine involving a three-step chain-elongation cycle. Each passage through the cycle results in the net addition of a single methylene group, with up to six cycles of elongation occurring in A. thaliana. The first reaction of the cycle is catalyzed by a methylthioalkylmalate synthase (MAMS), which condenses a omega-methylthio-2-oxoalkanoic acid with acetyl-CoA. Here we have demonstrated that MAM1, one of two similar genes in the A. thaliana ecotype Columbia, encodes a MAMS catalyzing the condensing reactions of the first two elongation cycles but not those of further cycles. The Columbia ecotype is dominated by compounds that have undergone only two elongation cycles. The A. thaliana MAM1 protein exhibits basic sequence similarity to other previously described enzymes catalyzing the condensation of 2-oxo acids and acetyl-CoA, such as isopropylmalate synthase (EC 2.3.3.13), an enzyme of leucine biosynthesis, and homocitrate synthase (EC 2.3.3.14). It also shares similar properties with them, including the catalytic requirements for a divalent metal ion and an adenine nucleotide. However, the MAM1 protein does not show activity with the substrates of any of these other enzymes, and was chromatographically separable from isopropylmalate synthase in extracts of A. thaliana. Thus, MAM1 is exclusively an enzyme of secondary metabolism, distinct from primary metabolic enzymes catalyzing similar reactions.

Glucosinolates are natural plant products that have received rising attention due to their role in interactions between pests and crop plants and as chemical protectors against cancer. Glucosinolates are derived from amino acids and have aldoximes as intermediates. We report that cytochrome P450 CYP79F1 catalyzes aldoxime formation in the biosynthesis of aliphatic glucosinolates in Arabidopsis thaliana. Using recombinant CYP79F1 functionally expressed in Escherichia coli, we show that both dihomomethionine and trihomomethionine are metabolized by CYP79F1 resulting in the formation of 5-methylthiopentanaldoxime and 6-methylthiohexanaldoxime, respectively. 5-methylthiopentanaldoxime is the precursor of the major glucosinolates in leaves of A. thaliana, i.e. 4-methylthiobutylglucosinolate and 4-methylsulfinylbutylglucosinolate, and a variety of other glucosinolates in Brassica sp. Transgenic A. thaliana with cosuppression of CYP79F1 have a reduced content of aliphatic glucosinolates and a highly increased level of dihomomethionine and trihomomethionine. The transgenic plants have a morphological phenotype showing loss of apical dominance and formation of multiple axillary shoots. Our data provide the first evidence that a cytochrome P450 catalyzes the Nhydroxylation of chain-elongated methionine homologues to the corresponding aldoximes in the biosynthesis of aliphatic glucosinolates.

and the ‡ ‡IACR-Rothamsted, Harpenden, AL5 2JQ, United Kingdom CYP83B1 from Arabidopsis thaliana has been identified as the oxime-metabolizing enzyme in the biosynthetic pathway of glucosinolates. Biosynthetically active microsomes isolated from Sinapis alba converted p-hydroxyphenylacetaldoxime and cysteine into S-alkylated p-hydroxyphenylacetothiohydroximate, S-(p-hydroxyphenylacetohydroximoyl)-L-cysteine, the next proposed intermediate in the glucosinolate pathway. The production was shown to be dependent on a cytochrome P450 monooxygenase. We searched the genome of A. thaliana for homologues of CYP71E1 (P450ox), the only known oxime-metabolizing enzyme in the biosynthetic pathway of the evolutionarily related cyanogenic glucosides. By a combined use of bioinformatics, published expression data, and knock-out phenotypes, we identified the cytochrome P450 CYP83B1 as the oxime-metabolizing enzyme in the glucosinolate pathway as evidenced by characterization of the recombinant protein expressed in Escherichia coli. The data are consistent with the hypothesis that the oxime-metabolizing enzyme in the cyanogenic pathway (P450ox) was mutated into a "P450mox" that converted oximes into toxic compounds that the plant detoxified into glucosinolates.Glucosinolates are naturally occurring amino acid-derived S-glucosides of thiohydroximate-O-sulfonates. They co-occur with endogenous thioglucosidases called myrosinases that upon tissue damage hydrolyze glucosinolates into a wide range of degradation products such as e.g. isothiocyanates, nitriles, and thiocyanates. Glucosinolates (or rather their degradation products) are involved in plant defense and constitute characteristic flavor compounds and cancer-preventive agents in Brassica vegetables.The biosynthetic pathway from precursor amino acid to the core glucosinolate structure has been well studied, and many of the intermediates are known, including oximes, thiohydroximic acids, and desulfoglucosinolates (1, 2). Recently, it has been shown that cytochromes P450 belonging to the CYP79 family catalyze the conversion of amino acids to oximes (3-7). Little is known about the formation of thiohydroximic acids from oximes. The remaining part of the pathway for the core structure involves a UDP-glucose:thiohydroximic acid glucosyltransferase and a sulfotransferase (for review, see Ref.2).It has been proposed that aci-nitro compounds are intermediates in the conversion of oximes to thiohydroximic acids (8). This was supported by isolation of 1-nitro-2-phenylethane from Tropaeolum majus shoots and by in vivo conversion of phenylacetaldoxime into 1-nitro-2-phenylethane and of 1-nitro-2-[1,2-14 C]phenylethane into benzylglucosinolate (9). The aci-nitro is proposed to be conjugated with a sulfur donor to produce an S-alkyl thiohydroximate, possibly by a glutathione S-transferase (2). Biochemical studies indicate that the S-alkyl thiohydroximate is subsequently hydrolyzed to the thiohydroximic acid by a C-S lyase (10).Glucosinolates are related to cyanogenic glucosides because both...