EFD Is an ERF Transcription Factor Involved in the Control of Nodule Number and Differentiation inMedicago truncatula

Abstract

Mechanisms regulating legume root nodule development are still poorly understood, and very few regulatory genes have been cloned and characterized. Here, we describe EFD (for ethylene response factor required for nodule differentiation), a gene that is upregulated during nodulation in Medicago truncatula. The EFD transcription factor belongs to the ethylene response factor (ERF) group V, which contains ERN1, 2, and 3, three ERFs involved in Nod factor signaling. The role of EFD in the regulation of nodulation was examined through the characterization of a null deletion mutant (efd-1), RNA interference, and overexpression studies. These studies revealed that EFD is a negative regulator of root nodulation and infection by Rhizobium and that EFD is required for the formation of functional nitrogen-fixing nodules. EFD appears to be involved in the plant and bacteroid differentiation processes taking place beneath the nodule meristem. We also showed that EFD activated Mt RR4, a cytokinin primary response gene that encodes a type-A response regulator. We propose that EFD induction of Mt RR4 leads to the inhibition of cytokinin signaling, with two consequences: the suppression of new nodule initiation and the activation of differentiation as cells leave the nodule meristem. Our work thus reveals a key regulator linking early and late stages of nodulation and suggests that the regulation of the cytokinin pathway is important both for nodule initiation and development.

Deletion-Based Reverse Genetics in Medicago truncatula

“…Sequencing of these amplicons revealed a 1,570-bp deletion entirely removing the exon 1 and the 3' end of exon 2. This deletion completely abolishes the production of the EFD transcript, as verified by quantitative reverse transcription PCR analysis (Vernie et al, 2008). Following screening of the 3D pools, we were able to recover two mutant plants from 50 seeds taken from the identified family, one heterozygous and one homozygous for the efd1-1 mutation.…”

“…To demonstrate the utility of the De-TILLING strategy for recovering mutations in small genes, we targeted the transcription factor EFD (for ethylene response factor required for nodule differentiation; Vernie et al, 2008) initially identified as being upregulated in M. truncatula nodules using microarray analyses (El Yahyaoui et al, 2004) and suppression subtractive hybridization approaches (Godiard et al, 2007). The coding region of this gene consisted of two exons of 85 and 503 bp.…”

A Phylogenetic Strategy Based on a Legume-Specific Whole Genome Duplication Yields Symbiotic Cytokinin Type-A Response Regulators

“…symbiosis (Gonzalez-Rizzo et al, 2006;Vernié et al, 2008). Although their exact molecular functioning remains elusive, we can now position MtRR9 and MtRR11 in the Nod factor signaling network.…”

Deep Sequencing of the Medicago truncatula Root Transcriptome Reveals a Massive and Early Interaction between Nodulation Factor and Ethylene Signals

“…Homologs with symbiotic function and lower expression levels than ERN1, namely ERN2 (Medtr6g029180) and ERN3 (Medtr8g085960), were also identified in our analysis but not found to be differentially expressed. Vernié et al (2008) characterized a fourth AP2/ERF that is required for nodule differentiation at later nodule stages, EFD (contig_56456_1). In agreement with previous studies, our data show that EFD was expressed in M. truncatula roots but that EFD expression levels do not change following inoculation.…”