Novel Pathogen Responsive Promoters and cis-Regulatory Elements in Soybean

Promoters are key regulators of gene expression at the transcriptional level. Promoter functionality is largely determined by the cis-elements generally located within the promoter regions. In soybean, only a few promoters have been well studied, and pathogen responsive promoters and their regulatory cis-elements have not been widely investigated. Here we studied induction of pathogen responsive genes and their promoter regions in soybean. Using previously-reported and in-house transcriptomic data, we identified 24 genes highly up-regulated after inoculation with Sclerotinia sclerotiorum, Fusarium virguliforme, Phytophthora sojae, and Pseudomonas syringae. These genes encode proteins such as ABC transporters, Asparaginase, nipped-B-like protein (Delangin) SCC2-related, aldo/keto reductase, O-methyltransferase (OMT), ammonium transporter, and dihydroflavonol reductase/cinnamoyl-CoA reductase (DFR/CCR). Full-length cDNAs for OMT and DFR/CCR genes were also cloned and overexpressed in Arabidopsis to gain a better understanding of their functionality. The promoter regions for 21 of the aforementioned genes were subsequently isolated, fused with GFP, and rapidly characterized using transiently-transformed lima bean cotyledons and stably-transformed soybean hairy roots. Most of these promoters gave very low to moderate basal GFP expression in transient expression and uninoculated soybean hairy roots, except for the OMT and Asparaginase promoters that gave relatively high GFP expression levels. We also conducted a computational analysis of the promoter sequences to predict over-represented cis-elements likely associated to pathogen responses. This study will increase our understanding of defense responsive genes and expand the toolbox of soybean pathogen-inducible promoters and cis-elements for potential use in both basic research and development of transgenic plants with enhanced resistance.