Lights, location, action: Shade avoidance signalling over spatial scales.

Plants growing in dense vegetation stands need to flexibly position their photosynthetic organs to ensure optimal light capture in a competitive environment. They do so through a suite of developmental responses referred to as the shade avoidance syndrome. Belowground, root development is also adjusted in response to aboveground neighbour proximity. Canopies are dynamic and complex environments with heterogenous light cues in the far-red, red, blue and UV spectrum, which can be perceived with photoreceptors by spatially separated plant tissues. Molecular regulation of plant architecture adjustment via PHYTOCHROME-INTERACTING FACTOR (PIF) transcription factors and growth-related hormones such as auxin, gibberellic acid, brassinosteroids and abscisic acid were historically studied without much attention to spatial or tissue-specific context. Recent developments and technologies have, however, sparked strong interest in spatially explicit understanding of shade avoidance regulation. Other environmental factors such as temperature and nutrient availability interact with the molecular shade avoidance regulation network, often depending on the spatial location of the signals, and the responding organs. Here, we aim to review recent advances in how plants respond to heterogenous light cues and integrate these with other environmental signals.

Nitrate-induced CLE35 signaling peptides inhibit nodulation through the SUNN receptor and miR2111 repression.

Legume plants form nitrogen (N)-fixing symbiotic nodules when mineral N is limiting in soils. As N fixation is energetically costly compared to mineral N acquisition, these N sources, and in particular nitrate, inhibit nodule formation and N fixation. Here, in the model legume Medicago truncatula, we characterized a CLAVATA3-like (CLE) signaling peptide, MtCLE35, the expression of which is upregulated locally by high-N environments and relies on the Nodule Inception-Like Protein (NLP) MtNLP1. MtCLE35 inhibits nodule formation by affecting rhizobial infections, depending on the Super Numeric Nodules (MtSUNN) receptor. In addition, high N or the ectopic expression of MtCLE35 represses the expression and accumulation of the miR2111 shoot-to-root systemic effector, thus inhibiting its positive effect on nodulation. Conversely, ectopic expression of miR2111 or downregulation of MtCLE35 by RNA interference increased miR2111 accumulation independently of the N environment, and thus partially bypasses the nodulation inhibitory action of nitrate. Overall, these results demonstrate that the MtNLP1-dependent, N-induced MtCLE35 signaling peptide acts through the MtSUNN receptor and the miR2111 systemic effector to inhibit nodulation.

Independent Regulation of Symbiotic Nodulation by the SUNN Negative and CRA2 Positive Systemic Pathways.

Plant systemic signaling pathways allow the integration and coordination of shoot and root organ metabolism and development at the whole-plant level depending on nutrient availability. In legumes, two systemic pathways have been reported in the Medicago truncatula model to regulate root nitrogen-fixing symbiotic nodulation. Both pathways involve leucine-rich repeat receptor-like kinases acting in shoots and proposed to perceive signaling peptides produced in roots depending on soil nutrient availability. In this study, we characterized in the M. truncatula Jemalong A17 genotype a mutant allelic series affecting the Compact Root Architecture2 (CRA2) receptor. These analyses revealed that this pathway acts systemically from shoots to positively regulate nodulation and is required for the activity of carboxyl-terminally encoded peptides (CEPs). In addition, we generated a double mutant to test genetic interactions of the CRA2 systemic pathway with the CLAVATA3/EMBRYO SURROUNDING REGION peptide (CLE)/Super Numeric Nodule (SUNN) receptor systemic pathway negatively regulating nodule number from shoots, which revealed an intermediate nodule number phenotype close to the wild type. Finally, we showed that the nitrate inhibition of nodule numbers was observed in cra2 mutants but not in sunn and cra2 sunn mutants. Overall, these results suggest that CEP/CRA2 and CLE/SUNN systemic pathways act independently from shoots to regulate nodule numbers.

Unraveling new molecular players involved in the autoregulation of nodulation in Medicago truncatula.

The number of legume root nodules resulting from a symbiosis with rhizobia is tightly controlled by the plant. Certain members of the CLAVATA3/Embryo Surrounding Region (CLE) peptide family, specifically MtCLE12 and MtCLE13 in Medicago truncatula, act in the systemic autoregulation of nodulation (AON) pathway that negatively regulates the number of nodules. Little is known about the molecular pathways that operate downstream of the AON-related CLE peptides. Here, by means of a transcriptome analysis, we show that roots ectopically expressing MtCLE13 deregulate only a limited number of genes, including three down-regulated genes encoding lysin motif receptor-like kinases (LysM-RLKs), among which are the nodulation factor (NF) receptor NF Perception gene (NFP) and two up-regulated genes, MtTML1 and MtTML2, encoding Too Much Love (TML)-related Kelch-repeat containing F-box proteins. The observed deregulation was specific for the ectopic expression of nodulation-related MtCLE genes and depended on the Super Numeric Nodules (SUNN) AON RLK. Moreover, overexpression and silencing of these two MtTML genes demonstrated that they play a role in the negative regulation of nodule numbers. Hence, the identified MtTML genes are the functional counterpart of the Lotus japonicus TML gene shown to be central in the AON pathway. Additionally, we propose that the down-regulation of a subset of LysM-RLK-encoding genes, among which is NFP, might contribute to the restriction of further nodulation once the first nodules have been formed.

Nitrogen Systemic Signaling: From Symbiotic Nodulation to Root Acquisition.

Plant nutrient acquisition is tightly regulated by resource availability and metabolic needs, implying the existence of communication between roots and shoots to ensure their integration at the whole-plant level. Here, we focus on systemic signaling pathways controlling nitrogen (N) nutrition, achieved both by the root import of mineral N and, in legume plants, through atmospheric N fixation by symbiotic bacteria inside dedicated root nodules. We explore features conserved between systemic pathways repressing or enhancing symbiotic N fixation and the regulation of mineral N acquisition by roots, as well as their integration with other environmental factors, such as phosphate, light, and CO2 availability.

Compact Root Architecture 2 Promotes Root Competence for Nodulation through the miR2111 Systemic Effector.

Nitrogen-deprived legume plants form new root organs, the nodules, following a symbiosis with nitrogen-fixing rhizobial bacteria [1]. Because this interaction is beneficial for the plant but has a high energetic cost, nodulation is tightly controlled by host plants through systemic pathways (acting at long distance) to promote or limit rhizobial infections and nodulation depending on earlier infections and on nitrogen availability [2]. In the Medicago truncatula model legume, CLE12 (Clavata3/Embryo surrounding region 12) and CLE13 signaling peptides produced in nodulated roots act in shoots through the SUNN (Super Numeric Nodule) receptor to negatively regulate nodulation and therefore autoregulate nodule number [3-5]. Conversely, CEP (C-terminally Encoded Peptide) signaling peptides produced in nitrogen-starved roots act in shoots through the CRA2 (Compact Root Architecture 2) receptor to promote nodulation already in the absence of rhizobia [6-9]. We show in this study that a downstream shoot-to-root signaling effector of these systemic pathways is the shoot-produced miR2111 microRNA [10] that negatively regulates TML1 (Too Much Love 1) and TML2 [11] transcripts accumulation in roots, ultimately promoting nodulation. Low nitrogen conditions and CEP1 signaling peptides induce in the absence of rhizobia the production of miR2111 depending on CRA2 activity in shoots, thus favoring root competence for nodulation. Together with the SUNN pathway negatively regulating the same miR2111 systemic effector when roots are nodulated, this allows a dynamic fine-tuning of the nodulation capacity of legume roots by nitrogen availability and rhizobial cues.

The NIN transcription factor coordinates CEP and CLE signaling peptides that regulate nodulation antagonistically.

Legumes tightly regulate nodule number to balance the cost of supporting symbiotic rhizobia with the benefits of nitrogen fixation. C-terminally Encoded Peptides (CEPs) and CLAVATA3-like (CLE) peptides positively and negatively regulate nodulation, respectively, through independent systemic pathways, but how these regulations are coordinated remains unknown. Here, we show that rhizobia, Nod Factors, and cytokinins induce a symbiosis-specific CEP gene, MtCEP7, which positively regulates rhizobial infection. Via grafting and split root studies, we reveal that MtCEP7 increases nodule number systemically through the MtCRA2 receptor. MtCEP7 and MtCLE13 expression in rhizobia-inoculated roots rely on the MtCRE1 cytokinin receptor and on the MtNIN transcription factor. MtNIN binds and transactivates MtCEP7 and MtCLE13, and a NIN Binding Site (NBS) identified within the proximal MtCEP7 promoter is required for its symbiotic activation. Overall, these results demonstrate that a cytokinin-MtCRE1-MtNIN regulatory module coordinates the expression of two antagonistic, symbiosis-related, peptide hormones from different families to fine-tune nodule number.