Two CCAAT‐box‐binding transcription factors redundantly regulate early steps of the legume‐rhizobia endosymbiosis

Abstract

SUMMARYDuring endosymbiotic interactions between legume plants and nitrogen-fixing rhizobia, successful root infection by bacteria and nodule organogenesis requires the perception and transduction of bacterial lipo-chitooligosaccharidic signal called Nod factor (NF). NF perception in legume roots leads to the activation of an early signaling pathway and of a set of symbiotic genes which is controlled by specific early transcription factors (TFs) including CYCLOPS/IPD3, NSP1, NSP2, ERN1 and NIN. In this study, we bring convincing evidence that the Medicago truncatula CCAAT-box-binding NF-YA1 TF, previously associated with later stages of rhizobial infection and nodule meristem formation is, together with its closest homolog NF-YA2, also an essential positive regulator of the NF-signaling pathway. Here we show that NF-YA1 and NF-YA2 are both expressed in epidermal cells responding to NFs and their knock-down by reverse genetic approaches severely affects the NF-induced expression of symbiotic genes and rhizobial infection. Further over-expression, transactivation and ChIP-PCR approaches indicate that NF-YA1 and NF-YA2 function, at least in part, via the direct activation of ERN1. We thus propose a model in which NF-YA1 and NF-YA2 appear as early symbiotic regulators acting downstream of DMI3 and NIN and possibly within the same regulatory complexes as NSP1/2 to directly activate the expression of ERN1.

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

“…NSP1 has been shown to bind to NODULATION-RESPONSIVE ELEMENT cis-elements in the promoters of ENOD11, ERN1, and NIN, while NSP2 likely binds to this region indirectly by way of its interaction with NSP1 (Hirsch et al, 2009). Likewise, NIN binds to the promoters of NF-YA1, CLE-RS1, CLE-RS2, and NPL in L. japonicus, while NF-YA1 binds to the ERN1 promoter in M. truncatula and is believed to coregulate its expression together with NSP1 and NSP2 (Xie et al, 2012;Soyano et al, 2013Laloum et al, 2014). MERLIN predicts 4,393 regulator-target gene relationships, including several of these known associations.…”

“…Members of the heterotrimeric, nucleosome-like NF-Y family of TFs are targets of the NIN protein (Soyano et al, 2013) and mediate early infection and nodule development processes (Petroni et al, 2012;Laloum et al, 2013Laloum et al, , 2014. NF-YA subunits bind to conserved CCAAT motifs in promoters, where they recruit the histone 2A/2B-like NF-YC and NF-YB subunits (Nardini et al, 2013), which also bind DNA, albeit nonspecifically.…”

“…NF-YA subunits bind to conserved CCAAT motifs in promoters, where they recruit the histone 2A/2B-like NF-YC and NF-YB subunits (Nardini et al, 2013), which also bind DNA, albeit nonspecifically. NF-YA1 and NF-YA2 are paralogs that are essential for both infection and nodule meristematic persistence in M. truncatula (Combier et al, 2006;Laloum et al, 2014;Laporte et al, 2014), while silencing of NF-YC1 leads to the arrest of nodule development and infection in common bean (Phaseolus vulgaris; Zanetti et al, 2010). In addition to NF-YA1, we observed that two B-type NF-Ys (NF-YB16 and NF-YB18) exhibit a Nod factor-induced and ET-repressed transcriptional response by 6 hpi (Fig.…”

A Laser Dissection-RNAseq Analysis Highlights the Activation of Cytokinin Pathways by Nod Factors in the Medicago truncatula Root Epidermis

“…In the epidermis, NIN is necessary for the onset of rhizobium infection Fournier et al, 2015) but restricts the expression of EARLY NODULIN11 (ENOD11), a marker of the preinfection and infection steps Vernié et al, 2015). ENOD11 is up-regulated in response to NF by ETHYLENE RESPONSE FACTOR REQUIRED FOR NODULATION1 (ERN1; Andriankaja et al, 2007;Middleton et al, 2007) as well as NUCLEAR TRAN-SCRIPTION FACTOR Y (MtNF-YA1) and MtNF-YA2 (Laloum et al, 2014). It was shown earlier that two other transcriptional regulators, NODULATION SIG-NALING PATHWAY1 (NSP1) and NSP2, are essential for nodulation (Kaló et al, 2005;Smit et al, 2005) and form a complex binding the promoter of NIN, ERN1, and MtENOD11 (Hirsch et al, 2009).…”

“…root tissues (Vernié et al, 2015). NIN is required to initiate nodule formation in the cortex (Schauser et al, 1999) and to transcriptionally activate NF-YA1 (Soyano et al, 2013;Laloum et al, 2014), a transcription factor (TF) involved in various nodulation steps (Combier et al, 2006;Soyano et al, 2013;Laporte et al, 2014;Xiao et al, 2014). In the epidermis, NIN is necessary for the onset of rhizobium infection Fournier et al, 2015) but restricts the expression of EARLY NODULIN11 (ENOD11), a marker of the preinfection and infection steps Vernié et al, 2015).…”