Wounding and methyl jasmonate increase cyanogenic glucoside concentrations in Sorghum bicolor via upregulation of biosynthesis

Abstract

The cyanogenic glucoside, dhurrin, present in Sorghum bicolor is thought to have multiple functions, including in defence against herbivory. The hormone methyl jasmonate (MeJA) is also induced by herbivory and is key to instigating defence processes in plants. To investigate whether dhurrin is induced in response to herbivore attack and also to the associated presence of MeJA, sorghum plants were either wounded or exogenous MeJA was applied. We show that specific wounding (pin board and perforation) and the application of MeJA increases dhurrin concentration in leaves and sheath tissue 12 h after treatment. Quantitative PCR shows that the expression of two genes, SbCYP79A1 and SbUGT85B1, involved in the synthesis of dhurrin are significantly induced by exogenous MeJA and by wounding. Analysis of 2 kb of sequence upstream of the start codon of SbCYP79A1 identifies several cis‐acting elements that have been linked to MeJA induction. A promoter deletion series, coupled to GFP, and transiently expressed in Nicotiana benthamiana suggests that there are potentially three sequence motifs (~−925 to −976) involved in the binding of transcription factors that result in increased expression of SbCYP79A1 and the synthesis of dhurrin in response to MeJA.

The Putative GATA Transcription Factor SbGATA22 as a Novel Regulator of Dhurrin Biosynthesis

“…Relative expression levels of nitrate reductase (Sb07g022750) were also measured for the nitrogen experiment. Previous research normalizing gene expression to two reference genes [25] has shown no significant differences when using both Ubiquitin and Actin (Sb01g010030) in conjunction or when using only Ubiquitin [12,28]. An EpMotion 5075 Robot (Eppendorf, Hamburg, Germany) was used to set up the 384 well plates, and qPCR reactions were performed using a Light Cycler 480II (Roche, Basel, Switzerland) with cycling parameters as follows: initial denaturation: 10 min, 95 • C; denaturation: 15 s, 95 • C; annealing: 15 s, 66/63 • C; extension: 15 s, 72 • C (acquired at end of extension), for 40 cycles.…”

“…Currently, little is known about the molecular regulation of dhurrin biosynthesis, although genome-wide demethylation has been found to affect SbCYP79A1 expression in sorghum [25]. The majority of sorghum studies have focused on dhurrin regulation at the physiological level or analyzed gene expression without identifying the drivers of this expression [18,[26][27][28]. The manipulation of dhurrin production has involved knocking out genes of the biosynthetic pathway rather than altering regulation [29,30].…”

“…In Lotus japonicus, methyl jasmonate has been shown to induce the expression of genes involved in the synthesis of the CNglc lotaustralin, and a bHLH transcription factor has been identified that controls this response [32]. While dhurrin is also believed to be regulated at the transcriptional level in sorghum [18,20] and methyl jasmonate has been shown to induce SbCYP79A1 expression and increase dhurrin concentration in leaves [28], to date, no direct regulator of the dhurrin biosynthetic genes has been reported.…”