CLE-HAR1 Systemic Signaling and NIN-Mediated Local Signaling Suppress the Increased Rhizobial Infection in the daphne Mutant of Lotus japonicus

Abstract

Legumes survive in nitrogen-limited soil by forming a symbiosis with rhizobial bacteria. During root nodule symbiosis, legumes strictly control the development of their symbiotic organs, the nodules, in a process known as autoregulation of nodulation (AON). The study of hypernodulation mutants has elucidated the molecular basis of AON. Some hypernodulation mutants show an increase in rhizobial infection in addition to developmental alteration. However, the relationship between the AON and the regulation of rhizobial infection has not been clarified. We previously isolated daphne, a nodule inception (nin) allelic mutant, in Lotus japonicus. This mutant displayed dramatically increased rhizobial infection, suggesting the existence of NIN-mediated negative regulation of rhizobial infection. Here, we investigated whether the previously isolated components of AON, especially CLAVATA3/ESR (CLE)-RELATED-ROOT SIGNAL1 (CLE-RS1), CLE-RS2, and their putative receptor HYPERNODULATION AND ABERRANT ROOT FORMATION1 (HAR1), were able to suppress increased infection in the daphne mutant. The constitutive expression of LjCLE-RS1/2 strongly reduced the infection in the daphne mutant in a HAR1-dependent manner. Moreover, reciprocal grafting analysis showed that strong reduction of infection in daphne rootstock constitutively expressing LjCLE-RS1 was canceled by a scion of the har1 or klavier mutant, the genes responsible for encoding putative LjCLE-RS1 receptors. These data indicate that rhizobial infection is also systemically regulated by CLE-HAR1 signaling, a component of AON. In addition, the constitutive expression of NIN in daphne har1 double-mutant roots only partially reduced the rhizobial infection. Our findings indicate that the previously identified NIN-mediated negative regulation of infection involves unknown local signaling, as well as CLE-HAR1 long-distance signaling.

Inhibition of legume nodulation by Pi deficiency is dependent on the autoregulation of nodulation (AON) pathway

“…We also provide evidence indicating that the AON pathway not only reduces the number of nodules in the Pi‐deficient plant, but also restricts the number of rhizobial infections, which was reflected in a reduction in the number of rhizobia‐induced root hair deformation events (Figure 4) (Isidra‐Arellano et al , 2018). Our data support early observations indicating that CLE‐HAR1 signaling inhibits rhizobial infection in L. japonicus (Yoro et al , 2019b). The fact that Pi deficiency induces the expression of critical components of the AON pathway, and also that the reduction of both rhizobia‐induced root hair deformation events and number of nodules in Pi‐deficient plants is NARK‐dependent, is supportive of our hypothesis that the Pi deficit itself triggers the AON pathway and nodule limitation.…”

“…The knockout mutants nark/sunn/har1 , tml and transgenic plants overexpressing the miRNA miR2111 show a supernodulation phenotype, indicating that these genetic components are crucial for maintaining an optimal number of nodules (Krusell et al , 2002; Nishimura et al , 2002; Searle et al , 2003; Schnabel et al , 2005; Takahara et al , 2013; Tsikou et al , 2018). However, the AON not only restricts nodule development, but also subsequent rhizobial infections (Yoro et al , 2019b). The cytokinin receptor LHK1 acts synergistically with the receptor HAR1 to regulate root susceptibility to rhizobial infection in L. japonicus ; the interplay of local and systemic pathways is therefore critical for modulating the optimal number of nodules (Miri et al , 2019).…”

Progress in the Self-Regulation System in Legume Nodule Development-AON (Autoregulation of Nodulation)

“…Evidence indicating the systemic signal control of nodule formation and activity was first found in the early 1950s. Using nodule removal, grafting, and split-root experiments, Kosslak et al showed that host plants regulate and limit the number of nodules by integrating the stem and root signals [ 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 ]. This systemic regulation of nodulation is called autoregulation of nodulation (AON) [ 10 ].…”

“…In 2018, Tsikou and other studies found that high soil NO 3 − concentration can inhibit rhizobial infection and legume nodulation, since NO 3 − uptake from soil is more cost-effective for plants than nitrogen-fixation via symbiosis with rhizobia, as a source of nitrogen [ 5 ]. The AON system also inhibits rhizobial infection by weakening the plant’s perception of the rhizobia signals [ 5 , 6 , 7 ]. Therefore, we have expanded the current research on the AON system to include NO 3 − signaling, as it inhibits nodule initiation and development.…”