Project description:A transcriptome of Cluster II Frankia in nitrogen-fixing root-nodule symbiosis with the host plant, Datisca glomerata, was obtained by Illumina sequencing and mapping to the corresponding published genome (NCBI Bioproject PRJNA46257). Major metabolic pathways detected in Cluster II Frankia in symbiosis with Datisca glomerata were comparable to those described as up-regulated in the Frankia alni-Alnus glutinosa symbiosis (N Alloisio et al, MPMI 23(5):593-607, 2010): nitrogenase biosynthesis, tricarboxylic acid cycle, respiratory-chain related functions, oxidation protection, and terpenoid biosynthesis. These functions are consistent with the primary activities of Frankia in root nodules, e.g. to carry out the energetically-demanding fixation of atmospheric dinitrogen to ammonium, and to maintain internal reducing conditions. Expression of genes coding for amino-acid biosynthetic pathways, including arginine as reported previously (AM Berry et al. Funct Plant Biol 38, 645–652, 2011) was detected. A striking difference from other Frankia strains, revealed in the transcriptome of the Cluster II Frankia in symbiosis, was the expression of homologs of rhizobial nodulation genes, nodA, nodB and nodC.
Project description:Plant-derived nodule-specific Cys-rich peptides (NCRs) play essential roles towards microsymbionts control. To pinpoint whether DgDef1 — a nodule-specific defensin from the actinorhizal plant Datisca glomerata — fulfills similar functions, the peptide was characterized. DgDef1 affects growth of several bacteria including Sinorhizobium meliloti. In this experiment, it was analyzed if DgDef1 without its CTPP domain (DgDef1delta SP delta CTPP) triggers a cellular response in S. meliloti. The S. meliloti strain Rm1021 was challenged in 10 mL TY medium (OD600 = 0.5) with 25 µg/mL of DgDef1 delta SP delta CTPP at 30°C, 150 rpm for 1h. The negative control was treated with acetonitrile:water (1:3) instead of DgDef1 delta SP delta CTPP. Assays were performed in four replicates. RNA was isolated and material from biological replicates were combined in equal amounts. This resulted in two RNA pools representing DgDef1 treated and untreated Rm1021 cells. The pooled RNAs were used for RNAseq in order to identify differentially expressed genes.