Glucose enhances indolic glucosinolate biosynthesis without reducing primary sulfur assimilation

Abstract

The effect of glucose as a signaling molecule on induction of aliphatic glucosinolate biosynthesis was reported in our former study. Here, we further investigated the regulatory mechanism of indolic glucosinolate biosynthesis by glucose in Arabidopsis. Glucose exerted a positive influence on indolic glucosinolate biosynthesis, which was demonstrated by induced accumulation of indolic glucosinolates and enhanced expression of related genes upon glucose treatment. Genetic analysis revealed that MYB34 and MYB51 were crucial in maintaining the basal indolic glucosinolate accumulation, with MYB34 being pivotal in response to glucose signaling. The increased accumulation of indolic glucosinolates and mRNA levels of MYB34, MYB51, and MYB122 caused by glucose were inhibited in the gin2-1 mutant, suggesting an important role of HXK1 in glucose-mediated induction of indolic glucosinolate biosynthesis. In contrast to what was known on the function of ABI5 in glucose-mediated aliphatic glucosinolate biosynthesis, ABI5 was not required for glucose-induced indolic glucosinolate accumulation. In addition, our results also indicated that glucose-induced glucosinolate accumulation was due to enhanced sulfur assimilation instead of directed sulfur partitioning into glucosinolate biosynthesis. Thus, our data provide new insights into molecular mechanisms underlying glucose-regulated glucosinolate biosynthesis.

Figure 1. Relative expression levels of MYB34 (A), MYB51 (B), MYB122 (C), CYP79B2 (D), and CYP83B1 (E) in young seedlings treated with glucose or sorbitol for indicated times.

The expression level was measured in 10-day-old Arabidopsis seedlings treated with 3% glucose or sorbitol, and then the whole plants were collected 6, 12, 18, 24, and 36 h after treatment, respectively. Each data point represents the mean of five independent biological replicates per treatment (mean ± SE). Expression level of genes in water-treated seedlings was set to 1. The gene transcription levels upon three treatments were compared for each time point and values not sharing a common letter are significantly different at P < 0.05.

Figure 2. Total indolic glucosinolate content and relative expression levels of genes related to glucosinolate biosynthesis in double and triple myb mutant seedlings treated with glucose or sorbitol.

Effect of glucose on accumulation of total indolic glucosinolates (A) was measured in myb34myb122, myb51myb122, myb34myb51 double and myb34myb51myb122 triple mutants, as well as expression levels of MYB34 (B), MYB51(C), CYP79B2 (D), and CYP83B1 (E) in these mutants. Ten-day-old Arabidopsis seedlings were treated with 3% glucose or sorbitol, and then the whole plants were collected 3 days (A) or 18 h (B–E) after treatment. Each data point represents the mean of three to five (for qPCR assay) or six (for glucosinolate assay) independent biological replicates per treatment (mean ± SE). Expression level of genes in water-treated Col-0 seedlings was set to 1. Values not sharing a common letter are significantly different at P < 0.05.

Figure 3. Total indolic glucosinolate content and relative expression levels of MYB34, MYB51, and MYB122 in glucose- or sorbitol-treated gin2-1 seedlings.

Ten-day-old Arabidopsis seedlings were treated with 3% glucose or sorbitol, and then the whole plants were collected 3 days (A) or 18 h (B–D) after treatment. Each data point represents the mean of five (for qPCR assay) or six (for glucosinolate assay) independent biological replicates per treatment (mean ± SE). Values not sharing a common letter are significantly different at P < 0.05. Relative expression values are given compared with Ler seedlings treated by water.

Figure 4. Total indolic glucosinolate content and relative expression levels of MYB34, MYB51, and MYB122 in abi5-7 treated with glucose or sorbitol.

Ten-day-old Arabidopsis seedlings were treated with 3% glucose or sorbitol, and then the whole plants were collected 3 days (A) or 18 h (B–D) after treatment. Each data point represents the mean of five (for qPCR assay) or six (for glucosinolate assay) independent biological replicates per treatment (mean ± SE). Values not sharing a common letter are significantly different at P < 0.05. Relative expression values are given compared with Col-0 seedlings treated by water.

Figure 5. Relative expression levels of CYP79B2 (A) and CYP83B1 (B) in abi5-7 treated with glucose or sorbitol.

Ten-day-old Arabidopsis seedlings were treated with 3% glucose or sorbitol, and then the whole plantswere harvested 18 h after treatment. Each data point represents the mean of five independent biological replicates per treatment (mean ± SE). Values not sharing a common letter are significantly different at P < 0.05. Relative expression values are given compared with Col-0 seedlings treated by water.

Figure 6. Relative expression levels of genes related to sulfate metabolism in glucose- or sorbitol-treated seedlings.

Ten-day-old Arabidopsis seedlings were treated with 3% glucose or sorbitol, and then the whole plants were harvested 6 h (A,B) or 12 h (C,D) after treatment. Each data point represents the mean of five independent biological replicates per treatment (mean ± SE). Values not sharing a common letter are significantly different at P < 0.05. Relative expression values are given compared with seedlings treated by water.

Figure 7. Glucosinolate contents in glucose-treated seedlings grown at different concentrations of sulfate.

Arabidopsis seedlings (wild type Col-0) were cultured at different concentrations of sulfate for 10 days and subsequently treated with 3% glucose. The whole plants were harvested 3 days after glucose treatment. Total aliphatic and indolic glucosinolate contents were measured. Each data point represents the mean of six independent biological replicates per treatment (mean ± standard error). Values not sharing a common letter are significantly different at P < 0.05.

Figure 8. Cys and GSH contents in glucose-treated seedlings grown at different concentrations of sulfate.

Arabidopsis seedlings (wild type Col-0) were cultured at different concentrations of sulfate for 10 days and then treated with 3% glucose. The whole plants were harvested 3 days after treatment, and then cysteine and GSH contents were measured. Each data point represents the mean of five independent biological replicates per treatment (mean ± standard error). Values not sharing a common letter are significantly different at P < 0.05.

Figure 9. A model for glucose-regulated glucosinolate biosynthesis in Arabidopsis.

Glucose plays a key and positive role in both primary and secondary metabolism. Symbols with light green color represent sulfate metabolism pathway. Symbols with dark green color represent aliphatic glucosinolate biosynthesis and regulation. Symbols with blue color represent indolic glucosinolate biosynthesis and regulation. Multiple enzymatic steps are indicated with interrupted arrows. GLC, glucose; Met, methionine; Trp, tryptophan; Desulfo-GS, Desulfo-glucosinolates; ^{– – – –>} possible activation.