A CEP Peptide Receptor-Like Kinase Regulates Auxin Biosynthesis and Ethylene Signaling to Coordinate Root Growth and Symbiotic Nodulation in Medicago truncatula

Abstract

Because of the large amount of energy consumed during symbiotic nitrogen fixation, legumes must balance growth and symbiotic nodulation. Both lateral roots and nodules form on the root system, and the developmental coordination of these organs under conditions of reduced nitrogen (N) availability remains elusive. We show that the Medicago truncatula COMPACT ROOT ARCHITECTURE2 (MtCRA2) receptor-like kinase is essential to promote the initiation of early symbiotic nodulation and to inhibit root growth in response to low N. C-TERMINALLY ENCODED PEPTIDE (MtCEP1) peptides can activate MtCRA2 under N-starvation conditions, leading to a repression of YUCCA2 (MtYUC2) auxin biosynthesis gene expression, and therefore of auxin root responses. Accordingly, the compact root architecture phenotype of cra2 can be mimicked by an auxin treatment or by overexpressing MtYUC2, and conversely, a treatment with YUC inhibitors or an MtYUC2 knockout rescues the cra2 root phenotype. The MtCEP1-activated CRA2 can additionally interact with and phosphorylate the MtEIN2 ethylene signaling component at Ser 643 and Ser 924 , preventing its cleavage and thereby repressing ethylene responses, thus locally promoting the root susceptibility to rhizobia. In agreement with this interaction, the cra2 low nodulation phenotype is rescued by an ein2 mutation. Overall, by reducing auxin biosynthesis and inhibiting ethylene signaling, the MtCEP1/MtCRA2 pathway balances root and nodule development under low-N conditions.

The Regulation of Nodule Number in Legumes Is a Balance of Three Signal Transduction Pathways

“…Currently, peptides, miRNAs, and hormones are all known to mediate nitrogen demand signaling under low nitrate conditions by binding to the receptors or by affecting gene expression in root and or shoot [ 59 , 61 , 62 ] (see Figure 2 b). A root generated peptide from the C-TERMINALLY ENCODED PEPTIDE (CEP) family, MtCEP1, is the only well-studied peptide signal of M. truncatula nitrogen demand signaling, exhibit negative effects on lateral root formation and positive effect on nodulation under low N conditions [ 61 ] ( Figure 2 b).…”

“…The CEP1-CRA2 dependent enhanced expression of miR2111 in shoots (see AON section below) was shown to target the mRNAs of the genes TOO MUCH LOVE 1 ( TML1 ) and TML2 in roots, lowering the transcript levels of both genes presumably to maintain susceptibility to symbiotic nodulation [ 59 ] ( Figure 2 b). A separate study showed that CEP1-activated MtCRA2 phosphorylates MtEIN2 (ETHYLENE INSENSITIVE 2), preventing its cleavage and repressing an ethylene response, thus promoting the root susceptibility to rhizobia [ 62 ] ( Figure 2 b). Exogenous ethylene treatment during the first 24 to 48 h of rhizobial inoculation is enough to suppress nodulation in wild type plants [ 65 ].…”

“…The ethylene-mediated suppression of nodulation in most hypernodulating AON mutants contrasted with the EIN mutants suggests an independent role of ethylene and AON in controlling nodulation [ 65 , 68 ]. As opposed to maintaining susceptibility (described in nitrogen dependent control of nodulation, also see Figure 2 b), rhizobial inoculation leads to rapid upregulation of ethylene biosynthesis that promotes MtEIN2 cleavage, activating the ethylene pathway to inhibit rhizobia infection [ 62 ]. Taken together, the data suggest that ethylene controls the number of nodules through MtEIN2 locally by controlling susceptibility to rhizobial infection independent of the AON pathway.…”

Multigene editing reveals that MtCEP1/2/12 redundantly control lateral root and nodule number in Medicago truncatula

“…C-terminally encoded peptides (CEPs) are small, secreted peptides containing 15 highly conserved amino acids in the mature state, which participate in root development and other multiple physiological activities, such as symbiotic nitrogen fixation and nitrate uptake ( Ohyama et al , 2008 ; Imin et al , 2013 ; Tabata et al , 2014 ). These peptides are derived from non-functional pre-propeptides, processed by proteolytic cleavage and the addition of post-translational modifications (PTMs), and then transferred to the apoplast, stem xylem, or possibly the endoplasmic reticulum (ER), where they function as ligands with their corresponding receptors to trigger downstream responses ( Okamoto et al , 2016 ; Patel et al , 2018 ; Zhu et al , 2020 ). Genes from the CEP family are present across seed plants and have evolved via duplication and variation, which has led to the diversification of CEP -associated regulatory networks ( Delay et al , 2013a ; Ogilvie et al , 2014 ).…”

SiCEP3, a C-terminally encoded peptides fromSetaria italica, promotes ABA import and signaling pathway

“…In Medicago , MtCEP1 is expressed in primary root tips, vascular tissue, and lateral organ primordia, particularly under nitrogen (N) limitations (Imin et al, 2013). The Medicago COMPACT ROOT ARCHITECTURE2 (MtCRA2) receptor-like kinase is essential for inhibiting root growth in response to low N, leading to a repression of MtYUC2 auxin biosynthesis gene expression (Zhu et al, 2020). Together, these data suggest a relationship between CEP peptide signaling and auxin.…”