Dynamic modulation of nodulation factor receptor levels by phosphorylation-mediated functional switch of a RING-type E3 ligase during legume nodulation

The precise control of receptor levels is crucial for initiating cellular signaling transduction in response to specific ligands;however,such mechanisms regulating nodulation factor(NF)receptor(NFR)-mediated perception of NFs to establish symbiosis remain unclear.In this study,we unveil the pivotal role of the NFR-interacting RING-type E3 ligase 1(NIRE1)in regulating NFR1/NFR5 homeostasis to optimize rhizobial infection and nodule development in Lotus japonicus.We demonstrated that NiRE1 has a dual function in this regulatory process.It associates with both NFR1 and NFR5,facilitating their degradation through K48-linked polyubiquitination before rhizobial inoculation.However,following rhizobial inoculation,NFR1 phosphorylates NIRE1ata conserved residue,Tyr-109,inducing a functional switch in NIRE1,which enables NIRE1tomediateK63-linkedpolyubiquitination,thereby stabilizing NFR1/NFR5 in infected root cells.The introduction of phospho-dead NIRE1Y1osF leads to delayed nodule development,underscoring the significance of phosphorylation at Tyr-1o9 in orchestrating symbiotic processes.Conversely,expression of the phospho-mimic NIRE1Y0E results in the formation of spontaneous nodules in L.japonicus,further emphasizing the critical role of the phosphorylation-dependent functional switch in NiRE1.In summary,these findings uncover a fine-tuned symbiotic mechanism that a single E3 ligase could undergo a phosphorylationdependent functional switch to dynamically and precisely regulate NF receptor protein levels.

NODULATION TRIO in Medicago truncatula:Unveiling the redundant roles of MtLYK2,MtLYK3,and MtLYK2bis

The legume–rhizobial symbiosis is a distinctive model in plant–microbe interactions.Despite being foreign invaders,rhizobia are acknowledged as“friends”by compatible host plants,leading to mutualistic endosymbiosis(Cao et al.,2017).

A novel secreted protein, NISP1, is phosphorylated by soybean Nodulation Receptor Kinase to promote nodule symbiosis

Nodulation Receptor Kinase(NORK) functions as a co-receptor of Nod factor receptors to mediate rhizobial symbiosis in legumes, but its direct phosphorylation substrates that positively mediate root nodulation remain to be fully identified.Here, we identified a GmNORK-Interacting Small Protein(GmNISP1) that functions as a phosphorylation target of GmNORK to promote soybean nodulation. GmNORKα directly interacted with and phosphorylated GmNISP1. Transcription of GmNISP1 was strongly induced after rhizobial infection in soybean roots and nodules. GmNISP1 encodes a peptide containing 90 amino acids with a “DY” consensus motif at its N-terminus.GmNISP1 protein was detected to be present in the apoplastic space. Phosphorylation of GmNISP1 by GmNORKα could enhance its secretion into the apoplast. Pretreatment with either purified GmNISP1 or phosphorylation-mimic GmNISP1~(12D) on the roots could significantly increase nodule numbers compared with the treatment with phosphorylation-inactive GmNISP1~(12A).The data suggested a model that soybean GmNORK phosphorylates GmNISP1 to promote its secretion into the apoplast, which might function as a potential peptide hormone to promote root nodulation.

Suppression of LjBAK1-mediated immunity by SymRK promotes rhizobial infection in Lotus japonicus.

An important question in biology is how organisms can associate with different microbes that pose no threat (commensals), pose a severe threat (pathogens) and those that are beneficial (symbionts). The root nodule symbiosis serves as important model system to address such questions in the context of plant-microbe interactions. It is now generally accepted that rhizobia have the abilities to actively suppress host immune responses during the infection process, analogous to the way in which plant pathogens can evade immune recognition. However, much remains to be elucidated with regard to the mechanisms by which the host recognizes the rhizobia as pathogens and how, subsequently, these pathways are suppressed to allow establishment of the nitrogen fixing symbiosis. In this study, we found that SymRK (Symbiosis Receptor-like Kinase) is required for rhizobial suppression of plant innate immunity in Lotus japonicus. SymRK associates with LjBAK1 (BRASSINOSTEROID INSENSITIVE 1-Associated receptor Kinase 1), a well characterized, positive regulator of plant innate immunity, and directly inhibits LjBAK1 kinase activity. Rhizobial inoculation enhances the association between SymRK and LjBAK1 in planta. LjBAK1 is required to regulate plant innate immunity and plays a negative role in mediating rhizobial infection in L. japonicus. The data indicate that the protein complex of SymRK-LjBAK1 serves as an intersection point between rhizobial symbiotic signaling pathways and innate immunity pathways, which provides an evidence that rhizobia might actively suppress the host's ability to mount a defense response in the legume-rhizobium symbiosis.

An MAP kinase interacts with LHK1 and regulates nodule organogenesis in Lotus japonicus

Symbiosis receptor-like kinase(SymRK) is a key protein mediating the legume-Rhizobium symbiosis.Our previous work has identified an MAP kinase kinase,SIP2,as a SymRK-interacting protein to positively regulate nodule organogenesis in Lotus japonicus,suggesting that an MAPK cascade might be involved in Rhizobium-legume symbiosis.In this study,LjMPK6 was identified as a phosphorylation target of SIP2.Stable transgenic L.japonicus with RNAi silencing of LjMPK6 decreased the numbers of nodule primordia(NP) and nodule,while plants overexpressing LjMPK6 increased the numbers of nodule,infection threads(ITs),and NP,indicating that LjMPK6 plays a positive role in nodulation.LjMPK6 could interact with a cytokinin receptor,LHK1 both in vivo and in vitro.LjMPK6 was shown to compete with LHP1 to bind to the receiver domain(RD) of LHK1 and to downregulate the expression of two LjACS(1-aminocyclopropane-1-carboxylic acid synthase) genes and ethylene levels during nodulation.This study demonstrated an important role of LjMPK6 in regulation of nodule organogenesis and ethylene production in L.japonicus.