The NAC family transcription factor GmNAC42–1 regulates biosynthesis of the anticancer and neuroprotective glyceollins in soybean

Background Glyceollins are isoflavonoid-derived pathogen-inducible defense metabolites (phytoalexins) from soybean (Glycine max L. Merr) that have important roles in providing defense against pathogens. They also have impressive anticancer and neuroprotective activities in mammals. Despite their potential usefulness as therapeutics, glyceollins are not economical to synthesize and are biosynthesized only transiently and in low amounts in response to specific stresses. Engineering the regulation of glyceollin biosynthesis may be a promising approach to enhance their bioproduction, yet the transcription factors (TFs) that regulate their biosynthesis have remained elusive. To address this, we first aimed to identify novel abiotic stresses that enhance or suppress the elicitation of glyceollins and then used a comparative transcriptomics approach to search for TF gene candidates that may positively regulate glyceollin biosynthesis. Results Acidity stress (pH 3.0 medium) and dehydration exerted prolonged (week-long) inductive or suppressive effects on glyceollin biosynthesis, respectively. RNA-seq found that all known biosynthetic genes were oppositely regulated by acidity stress and dehydration, but known isoflavonoid TFs were not. Systemic acquired resistance (SAR) genes were highly enriched in the geneset. We chose to functionally characterize the NAC (NAM/ATAF1/2/CUC2)-family TF GmNAC42–1 that was annotated as an SAR gene and a homolog of the Arabidopsis thaliana (Arabidopsis) indole alkaloid phytoalexin regulator ANAC042. Overexpressing and silencing GmNAC42–1 in elicited soybean hairy roots dramatically enhanced and suppressed the amounts of glyceollin metabolites and biosynthesis gene mRNAs, respectively. Yet, overexpressing GmNAC42–1 in non-elicited hairy roots failed to stimulate the expressions of all biosynthesis genes. Thus, GmNAC42–1 was necessary but not sufficient to activate all biosynthesis genes on its own, suggesting an important role in the glyceollin gene regulatory network (GRN). The GmNAC42–1 protein directly bound the promoters of biosynthesis genes IFS2 and G4DT in the yeast one-hybrid (Y1H) system. Conclusions Acidity stress is a novel elicitor and dehydration is a suppressor of glyceollin biosynthesis. The TF gene GmNAC42–1 is an essential positive regulator of glyceollin biosynthesis. Overexpressing GmNAC42–1 in hairy roots can be used to increase glyceollin yields \u3e 10-fold upon elicitation. Thus, manipulating the expressions of glyceollin TFs is an effective strategy for enhancing the bioproduction of glyceollins in soybean

Switchgrass (Panicum virgatum L.) polyubiquitin gene (PvUbi1 and PvUbi2) promoters for use in plant transformation

<p>Abstract</p> <p>Background</p> <p>The ubiquitin protein is present in all eukaryotic cells and promoters from ubiquitin genes are good candidates to regulate the constitutive expression of transgenes in plants. Therefore, two switchgrass (<it>Panicum virgatum </it>L.) ubiquitin genes (<it>PvUbi1 </it>and <it>PvUbi2</it>) were cloned and characterized. Reporter constructs were produced containing the isolated 5' upstream regulatory regions of the coding sequences (i.e. <it>PvUbi1 </it>and <it>PvUbi2 </it>promoters) fused to the <it>uidA </it>coding region (<it>GUS</it>) and tested for transient and stable expression in a variety of plant species and tissues.</p> <p>Results</p> <p><it>PvUbi1 </it>consists of 607 bp containing <it>cis</it>-acting regulatory elements, a 5' untranslated region (UTR) containing a 93 bp non-coding exon and a 1291 bp intron, and a 918 bp open reading frame (ORF) that encodes four tandem, head -to-tail ubiquitin monomer repeats followed by a 191 bp 3' UTR. <it>PvUbi2 </it>consists of 692 bp containing <it>cis</it>-acting regulatory elements, a 5' UTR containing a 97 bp non-coding exon and a 1072 bp intron, a 1146 bp ORF that encodes five tandem ubiquitin monomer repeats and a 183 bp 3' UTR. <it>PvUbi1 </it>and <it>PvUbi2 </it>were expressed in all examined switchgrass tissues as measured by qRT-PCR. Using biolistic bombardment, <it>PvUbi1 </it>and <it>PvUbi2 </it>promoters showed strong expression in switchgrass and rice callus, equaling or surpassing the expression levels of the CaMV <it>35S, 2x35S, ZmUbi1</it>, and <it>OsAct1 </it>promoters. GUS staining following stable transformation in rice demonstrated that the <it>PvUbi1 </it>and <it>PvUbi2 </it>promoters drove expression in all examined tissues. When stably transformed into tobacco (<it>Nicotiana tabacum</it>), the <it>PvUbi2+3 </it>and <it>PvUbi2+9 </it>promoter fusion variants showed expression in vascular and reproductive tissues.</p> <p>Conclusions</p> <p>The <it>PvUbi1 </it>and <it>PvUbi2 </it>promoters drive expression in switchgrass, rice and tobacco and are strong constitutive promoter candidates that will be useful in genetic transformation of monocots and dicots.</p

Le Courrier

16 décembre 18221822/12/16 (A0,N350)

Reference genome sequence of the model plant Setaria

International audienc