bHLH05 Is an Interaction Partner of MYB51 and a Novel Regulator of Glucosinolate Biosynthesis in Arabidopsis

Abstract

By means of yeast (Saccharomyces cerevisiae) two-hybrid screening, we identified basic helix-loop-helix transcription factor05 (bHLH05) as an interacting partner of MYB51, the key regulator of indolic glucosinolates (GSLs) in Arabidopsis (Arabidopsis thaliana). Furthermore, we show that bHLH04, bHLH05, and bHLH06/MYC2 also interact with other R2R3-MYBs regulating GSL biosynthesis. Analysis of bhlh loss-of-function mutants revealed that the single bhlh mutants retained GSL levels that were similar to those in wild-type plants, whereas the triple bhlh04/05/06 mutant was depleted in the production of GSL. Unlike bhlh04/06 and bhlh05/06 mutants, the double bhlh04/05 mutant was strongly affected in the production of GSL, pointing to a special role of bHLH04 and bHLH05 in the control of GSL levels in the absence of jasmonic acid. The combination of two specific gain-of-function alleles of MYB and bHLH proteins had an additive effect on GSL levels, as demonstrated by the analysis of the double MYB34-1D bHLH05D94N mutant, which produces 20-fold more indolic GSLs than bHLH05D94N and ecotype Columbia-0 of Arabidopsis. The amino acid substitution D94N in bHLH05D94N negatively affects the interaction with JASMONATE-ZIM DOMAIN protein, thereby resulting in constitutive activation of bHLH05 and mimicking jasmonic acid treatment. Our study revealed the bHLH04, bHLH05, and bHLH06/MYC2 factors as novel regulators of GSL biosynthesis in Arabidopsis.

Changing a conserved amino acid in R2R3‐MYB transcription repressors results in cytoplasmic accumulation and abolishes their repressive activity in Arabidopsis

“…These results reveal the importance of this SID for SAD2 interaction. Previously, the Asp → Asn mutation in the JAZ interaction domain of MYC2 and MYC3 was found to abolish their binding to most JAZ proteins (Fernández‐Calvo et al ., ; Frerigmann et al ., ; Goossens et al ., ). Here, we show that the Asp → Asn mutation (D261N) in MYB4 significantly abolishes the interaction with SAD2.…”

Transport of defense compounds from source to sink: lessons learned from glucosinolates

“…Several studies have shown a direct link between jasmonic acid (JA) signaling and glucosinolate biosynthesis [74][75][76]. Expression of AtGTR1 is induced by JA treatment [19], which suggests an involvement in plant response to biotic stress.…”

Regulation of plant secondary metabolism and associated specialized cell development by MYBs and bHLHs

“…Furthermore, similar transcription factor complexes have been shown to regulate cell differentiation and cell fate specification in eukaryotes. For example, the interactions between MYB and bHLH family proteins have been shown to activate or repress different gene modules in glucosinolate biosynthesis in Arabidopsis (Figures 4 and 5A; Schweizer et al, 2013; Frerigmann et al, 2014a), and in skeletal muscle differentiation in animals (Kaspar et al, 2005). In addition, depending on the plant lineage, the interactions between MYB, bHLH and WD40 domain proteins have been shown to activate all or a set of biosynthetic genes in anthocyanin and proanthocyanidin biosynthesis (Figure 3, 5A; Goff et al, 1992; Grotewold et al, 2000; Aharoni et al, 2001; Zimmermann et al, 2004; Quattrocchio et al, 2006), as well as the cell differentiation of single-celled trichomes, non-hair root epidermal cells (atrichoblasts) in the embryonic stem and mucilage secretory cells in the outer seed coat (Figure 5B; Payne et al, 2000; Bernhardt et al, 2003; Esch et al, 2003; Zhang et al, 2003; Gonzalez et al, 2009; Li et al, 2009).…”