Omics-based identification of Arabidopsis Myb transcription factors regulating aliphatic glucosinolate biosynthesis

Abstract

Understanding plant metabolism as an integrated system is essential for metabolic engineering aimed at the effective production of compounds useful to human life and the global environment. The ''omics'' approach integrates transcriptome and metabolome data into a single data set and can lead to the identification of unknown genes and their regulatory networks involved in metabolic pathways of interest. One of the intriguing, although poorly described metabolic pathways in plants is the biosynthesis of glucosinolates (GSLs), a group of bioactive secondary products derived from amino acids that are found in the family Brassicaceae. Here we report the discovery of two R2R3-Myb transcription factors that positively control the biosynthesis of GSLs in Arabidopsis thaliana by an integrated omics approach. Combined transcriptome coexpression analysis of publicly available, condition-independent data and the condition-specific (i.e., sulfur-deficiency) data identified Myb28 and Myb29 as candidate transcription factor genes specifically involved in the regulation of aliphatic GSL production. Analysis of a knockout mutant and ectopic expression of the gene demonstrated that Myb28 is a positive regulator for basal-level production of aliphatic GSLs. Myb29 presumably plays an accessory function for methyl jasmonate-mediated induction of a set of aliphatic GSL biosynthetic genes. Overexpression of Myb28 in Arabidopsis-cultured suspension cells, which do not normally synthesize GSLs, resulted in the production of large amounts of GSLs, suggesting the possibility of efficient industrial production of GSLs by manipulation of these transcription factors. A working model for regulation of GSL production involving these genes, renamed Production of Methionine-Derived Glucosinolate (PMG) 1 and 2, are postulated.coexpression ͉ functional genomics ͉ transcriptomics

A robust omics-based approach for the identification of glucosinolate biosynthetic genes

“…In our analysis, AtBCAT-3 (At3g49680) and AtB-CAT-4 (At3g19710) were also coexpressed with the Met-derived GSL biosynthetic genes of known function (Hirai et al 2007), suggesting the involvement of these genes in Met side-chain elongation. The function of these genes has recently been confirmed, and AtBCAT-3 was shown to function in both GSL and amino acid biosynthesis (Knill et al 2008;Schuster et al 2006).…”

“…In addition, UGT74C1 (At2g31790), which is assumed to be involved in Met-derived GSL biosynthesis on the basis of the coexpression analysis (Gachon et al 2005), is positively regulated by Myb28 (Hirai et al 2007). On the other hand, a putative Tyr aminotransferase gene mentioned above is not regulated by Myb28 (Hirai et al 2007), suggesting that it may encode a C-S lyase involved only in Trp-/Phe-derived GSL biosynthesis. The reason for this assumption was that the C-S lyase gene SUR1 had been originally misannotated as a Tyr aminotransferase.…”

“…In fact, MAM genes and IPMS (isopropylmalate synthase) genes, which are responsible for Met side-chain elongation and Leu synthesis, respectively, share sequence similarity with each other and with bacterial IPMS (de Kraker et al 2007;Field et al 2004;Kroymann et al 2001). All of the above-mentioned candidate genes are under the transcriptional regulation involving Myb28 (Hirai et al 2007). In addition, UGT74C1 (At2g31790), which is assumed to be involved in Met-derived GSL biosynthesis on the basis of the coexpression analysis (Gachon et al 2005), is positively regulated by Myb28 (Hirai et al 2007).…”

“…All of the above-mentioned candidate genes are under the transcriptional regulation involving Myb28 (Hirai et al 2007). In addition, UGT74C1 (At2g31790), which is assumed to be involved in Met-derived GSL biosynthesis on the basis of the coexpression analysis (Gachon et al 2005), is positively regulated by Myb28 (Hirai et al 2007). On the other hand, a putative Tyr aminotransferase gene mentioned above is not regulated by Myb28 (Hirai et al 2007), suggesting that it may encode a C-S lyase involved only in Trp-/Phe-derived GSL biosynthesis.…”

“…This analysis suggested that Myb28 and Myb29 may be transcription factors positively regulating Met-derived GSL biosynthesis, but not Trpderived GSL biosynthesis. Reverse-genetic and molecular biological experiments have proved Myb28 to be a key transcription factor that positively regulates Met-derived GSL biosynthesis and Myb29 to be a transcription factor probably involved in methyl jasmonate-mediated induction of GSL biosynthesis (Hirai et al 2007). Concurrently, several groups have independently found that Myb28, Myb29 and Myb76 (At5g07700) are the positive regulators of Met-derived GSL biosynthesis and that Myb51 (At1g18570) and Myb122 (At1g74080), as well as previously-characterized Myb34 (At5g60890), are the positive regulators of Trp-derived GSL biosynthesis (Beekwilder et al 2008;Gigolashvili et al 2007a-c;Sonderby et al 2007;Malitsky et al 2008).…”