INVESTIGATING THE SORGOLEONE BIOSYNTHETIC …

ANALYSIS OF SORGOLEONE BIOSYNTHESIS IN …

The effects of JA and MeJa on the levels of transcript of five genes involved in sorgoleone biosynthesis were monitored. Levels of transcript of the sorgoleone biosynthetic genes, DES2, DES3, ARS1, ARS2, and OMT3 increased after exposure to either JA or MeJa at 5.0 μM. The extent to which the levels of transcript were affected by either JA or MeJa was dependent on the length of time the plants were exposed to jasmonates.

PKS activity responsible for sorgoleone biosynthesis was ..

The effects of JA and MeJa on the levels of transcript of five genes involved in sorgoleone biosynthesis were monitored. Levels of transcript of the sorgoleone biosynthetic genes, DES2, DES3, ARS1, ARS2, and OMT3 increased after exposure to either JA or MeJa at 5.0 μM. The extent to which the levels of transcript were affected by either JA or MeJa was dependent on the length of time the plants were exposed to jasmonates.

To determine the concentrations of JA and MeJa that promote maximum sorgoleone biosynthesis, sorghum seeds were imbibed with solution containing different concentrations (0, 0.5, 1.0, 5.0, 10, 50, 100, 250, and 500 μM) of either JA or MeJa for 1 week. To examine how gene expression and sorgoleone accumulation in response to MeJa and JA changed over time, 5-day-old sorghum seedlings were treated once with 5.0 μM JA or MeJa, and then their roots were harvested for analysis at multiple time points (0, 3, 6, 12, 24, 36, 48, 60, and 72 h after treatment). Roots samples were stored in sealed clear polyethylene plastic bags at −80 °C until use. There were four replicates for each time point, and the entire experiment was repeated twice.


Sorgoleone biosynthesis is likely ..

Sorgoleone biosynthesis appears to occur exclusively in root hair cells, which in sorghum appear as cytoplasmically dense cells filled with numerous osmiophilic deposits (Czarnota et al. ). These deposits presumably are associated with the rhizosecretion of sorgoleone, which can constitute as much as 85 % of the exudate dry weight in some cultivars (Bertin et al.; Czarnota et al., ; Duke ; Rimando et al.). The cell-specific localization and prolific output of the sorgoleone biosynthetic pathway renders the use of expressed sequence tag (EST) analysis an obvious method to isolate genes encoding sorgoleone biosynthetic enzymes (Baerson et al.). Labeling studies performed by Fate and Lynn () first demonstrated that biosynthesis proceeds through the action of an alkylresorcinol synthase (ARS), a novel type III polyketide synthase that acts on fatty acyl-CoA starter units (Baerson et al.). Subsequently, both the predicted 5-n-pentadecatrienyl resorcinol and a 3-methyl ether derivative of sorgoleone were identified in sorghum root extracts, indicating that dihydroxylation of the resorcinol ring is preceded by O-methylation at the 3-hydroxyl position (Baerson et al.; Dayan et al.; Fate and Lynn ). Once released into the rhizosphere, the resulting chemically unstable hydroquinone rapidly oxidizes to the bioactive sorgoleone benzoquinone, where it may persist in soil for extended periods (Czarnota et al.; Einhellig and Souza ; Gimsing et al.; Netzly et al.). Synthesis of sorgoleone from the available palmitoleoyl-CoA would, therefore, likely require, at minimum, the participation of a Δ12 and Δ15 fatty acid desaturase (DES), an ARS, 3-O-methyltransferase (OMT), and cytochrome P450 (Baerson et al.) (Fig. ).

SbOMT3 is also involved in the biosynthesis of sorgoleone

Sorgoleone biosynthesis appears to occur exclusively in root hair cells, which in sorghum appear as cytoplasmically dense cells filled with numerous osmiophilic deposits (Czarnota et al. ). These deposits presumably are associated with the rhizosecretion of sorgoleone, which can constitute as much as 85 % of the exudate dry weight in some cultivars (Bertin et al.; Czarnota et al., ; Duke ; Rimando et al.). The cell-specific localization and prolific output of the sorgoleone biosynthetic pathway renders the use of expressed sequence tag (EST) analysis an obvious method to isolate genes encoding sorgoleone biosynthetic enzymes (Baerson et al.). Labeling studies performed by Fate and Lynn () first demonstrated that biosynthesis proceeds through the action of an alkylresorcinol synthase (ARS), a novel type III polyketide synthase that acts on fatty acyl-CoA starter units (Baerson et al.). Subsequently, both the predicted 5-n-pentadecatrienyl resorcinol and a 3-methyl ether derivative of sorgoleone were identified in sorghum root extracts, indicating that dihydroxylation of the resorcinol ring is preceded by O-methylation at the 3-hydroxyl position (Baerson et al.; Dayan et al.; Fate and Lynn ). Once released into the rhizosphere, the resulting chemically unstable hydroquinone rapidly oxidizes to the bioactive sorgoleone benzoquinone, where it may persist in soil for extended periods (Czarnota et al.; Einhellig and Souza ; Gimsing et al.; Netzly et al.). Synthesis of sorgoleone from the available palmitoleoyl-CoA would, therefore, likely require, at minimum, the participation of a Δ12 and Δ15 fatty acid desaturase (DES), an ARS, 3-O-methyltransferase (OMT), and cytochrome P450 (Baerson et al.) (Fig. ).

Probing allelochemical biosynthesis in sorghum root hairs

This study investigated the roles of jasmonates in the regulation of sorgoleone accumulation and the expression of genes involved in sorgoleone biosynthesis in sorghum roots. Both methyl jasmonate (MeJa) and jasmonic acid (JA) substantially promoted root hair formation, secondary root development, root weight, and sorgoleone accumulation in sorghum roots. Sorgoleone content varied widely depending on the concentration of JA or MeJa and the duration of their application. Root weight and sorgoleone accumulation were highest after the application of JA or MeJa at a concentration of 5.0 μM, and then declined with increasing concentrations of jasmonates. At 5.0 μM, JA and MeJa increased sorgoleone content by 4.1 and 3.4-fold, respectively. Transcript accumulation was apparent for all genes, particularly for the O-methyltransferase 3 gene, which increased in expression levels up to 8.1-fold after a 36-h exposure to MeJa and 3.5-fold after a 48-h exposure to JA. The results of this study pave the way for more effective biosynthesis of sorgoleone, an important and useful allelochemical obtained from a variety of plant species.