GmTNRP1, associated with rhizobial type‐III effector NopT, regulates nitrogenase activity in the nodules of soybean (Glycine max)

Abstract

Soybean is a rich source of protein and oil for humans. It forms a symbiotic association with rhizobia to fix atmospheric nitrogen for its growth and development.Rhizobial type-III effectors act as important signaling molecules in symbiosis.However, they are poorly studied. In this study, mutation of TtsI in Sinorhizobium fredii HH103 reduced nodule numbers and dry weights in SN14 (cultivated soybean) and ZYD00006 (wild soybean). Moreover, it reduced nitrogenase activity in SN14 but not in ZYD00006. Based on the differences in nitrogenase activity between SN14 and ZYD00006 inoculated with HH103 and the TtsI mutant, two conditional quantitative trait loci associated with nitrogenase activity were identified in chromosome segment substitution lines derived from SN14 and ZYD00006.Using transcriptome analysis, single-nucleotide polymorphism (SNP) analysis, and qRT-PCR, Glyma.13G150000 (GmTNRP1) was identified as a candidate gene in soybean that was associated with nitrogenase activity, along with NopT (a type-III effector). In addition, GmTNRP1 (an LRR-RK family protein located on the cell membrane) was observed to be induced by NopT to negatively regulate nitrogenase activity. In transgenic hairy roots with GmTNRP1 silencing, no significant difference was observed in terms of nitrogenase activity between nodules inoculated with the NopT mutant and those inoculated with HH103 (wild type).However, the nitrogenase activity in the nodules decreased after overexpression of GmTNRP1. SNP located in the GmTNRP1 promoter affected its expression in response to NopT. Haplotype analysis revealed that SN14 belonged to Hap1 and ZYD00006 belonged to Hap2, which explained the different results in the two varieties. To the best of our knowledge, this is the first study to identify a soybean gene associated with a rhizobial type-III effector in the nodules. GmTNRP1 can negatively regulate nitrogenase activity in the nodules. This study enhanced the

NopAA and NopD Signaling Association-Related Gene GmNAC27 Promotes Nodulation in Soybean (Glycine max)

“…T3Es can exert their functions when secreted into plants, whereupon host proteins can respond to or interact with these effectors. Both genetic and RNA-seq analyses have been utilized to identify the downstream response pathways and interacting proteins associated with these T3Es [21,24,57,58]. Here, RNA-seq analyses revealed that NopAA can induce substantial numbers of DEGs enriched in the MAPK pathway, in line with prior data related to rhizobia infection.…”

“…With the exception of the complete inhibition of Rj2-soybean nodulation in response to NopP in USDA122, there have not been any reports of any one T3E completely determining whether or not nodulation can occur [56]. In previous studies, our team has utilized HH103 T3E gene insertion mutants and a variety of soybean genetic resources to assess nodulation dynamics, revealing soybean varietyspecific effects of these T3Es [21,24,44]. Both NopAA and NopD were previously found to be likely contributors to the nodulation process.…”

“…Electroporation was used to transform A. tumefaciens EHA105 with the CaMV35S-GmNAC27-GFP plasmid, after which the A. tumefaciens culture was adjusted to an OD 600 of 0.2 using infiltration buffer (10 mM MgCl 2 , 10 mM MES-KOH pH 5.6, 150 µM acetosyringone) and injected into the top leaves of tobacco plants. After 48 h, a Zeiss LSM 700 confocal laser scanning microscope (Zeiss, Oberkochen, Germany) was utilized to assess RFP and GFP fluorescence [21].…”

“…To date, a subset of Nop proteins have been identified and subject to biochemical and functional characterization. For example, the T3SS apparatus of Sinorhizobium has been shown to consist of at least three members of this protein family: NopA, NopB, and NopX [21]. The Bradyrhizobium USDA110 NopP protein is capable of interacting with the R protein GmNNL1 and promoting the induction of immune responses in hosts [22].…”

NopC/T/L Signal Crosstalk Gene GmPHT1-4

“…Analogous to NopL, NopT's transient expression in Nicotiana benthamiana results in leaf necrosis and elicits a hypersensitive response (HR) [29]. Additionally, NopT induction of GmTNRP1, an LRR-RK family protein at the cell membrane, serves to downregulate nitrogenase activity [30].…”