ABSTRACT
Legumes improve their mineral nutrition through nitrogen-fixing root nodule symbioses with soil rhizobia. Rhizobial infection of legumes is regulated by a number of transcription factors, including ERF Required for Nodulation1 (ERN1). Medicago truncatula plants defective in ERN1 are unable to nodulate, but still exhibit early symbiotic responses including rhizobial infection. ERN1 has a close homolog, ERN2, which shows partially overlapping expression patterns. Here we show that ern2 mutants exhibit a later nodulation phenotype than ern1, being able to form nodules but with signs of premature senescence. Molecular characterization of the ern2-1 mutation reveals a key role for a conserved threonine for both DNA binding and transcriptional activity. In contrast to either single mutant, the double ern1-1 ern2-1 line is completely unable to initiate infection or nodule development. The strong ern1-1 ern2-1 phenotype demonstrates functional redundancy between these two transcriptional regulators and reveals the essential role of ERN1/ERN2 to coordinately induce rhizobial infection and nodule organogenesis. While ERN1/ERN2 act in concert in the root epidermis, only ERN1 can efficiently allow the development of mature nodules in the cortex, probably through an independent pathway. Together, these findings reveal the key roles that ERN1/ERN2 play at the very earliest stages of root nodule development.
Subject(s)
Medicago truncatula/metabolism , Medicago truncatula/microbiology , Plant Diseases/microbiology , Plant Proteins/metabolism , Plant Roots/microbiology , Rhizobium/physiology , Symbiosis , Transcription Factors/metabolism , Alleles , Amino Acid Sequence , Base Sequence , Gene Expression Regulation, Plant , Medicago truncatula/genetics , Medicago truncatula/ultrastructure , Mutation/genetics , Mycorrhizae/physiology , Nitrogen Fixation , Organogenesis/genetics , Plant Epidermis/genetics , Plant Epidermis/microbiology , Plant Proteins/chemistry , Plant Proteins/genetics , Plant Roots/genetics , Plant Roots/ultrastructure , Promoter Regions, Genetic/genetics , Protein Binding , Root Nodules, Plant/metabolism , Root Nodules, Plant/microbiology , Root Nodules, Plant/ultrastructure , Signal Transduction/genetics , Symbiosis/genetics , Transcription Factors/chemistry , Transcription, GeneticABSTRACT
Rhizobial bacteria activate the formation of nodules on the appropriate host legume plant, and this requires the bacterial signaling molecule Nod factor. Perception of Nod factor in the plant leads to the activation of a number of rhizobial-induced genes. Putative transcriptional regulators in the GRAS family are known to function in Nod factor signaling, but these proteins have not been shown to be capable of direct DNA binding. Here, we identify an ERF transcription factor, ERF Required for Nodulation (ERN), which contains a highly conserved AP2 DNA binding domain, that is necessary for nodulation. Mutations in this gene block the initiation and development of rhizobial invasion structures, termed infection threads, and thus block nodule invasion by the bacteria. We show that ERN is necessary for Nod factor-induced gene expression and for spontaneous nodulation activated by the calcium- and calmodulin-dependent protein kinase, DMI3, which is a component of the Nod factor signaling pathway. We propose that ERN is a component of the Nod factor signal transduction pathway and functions downstream of DMI3 to activate nodulation gene expression.
