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Phosphorylation of rhizobial effector NopZ by soybean NORK promotes association with NENA and enhances nodulation

ABSTRACT: Legume-rhizobial symbiosis, involving the intracellular accommodation of rhizobia within host cells for nitrogen fixation, represents a unique model among plant-microbe interactions. While this symbiosis requires sophisticated regulatory networks, direct host control over symbiont proteins remains largely elusive. Here, we demonstrate that soybean Nodulation Receptor Kinase (GmNORK) interacts with and directly phosphorylates the Sinorhizobium fredii HH103 effector NopZ. This host-mediated phosphorylation promotes NopZ nuclear/perinuclear enrichment and is associated with enhanced NopZ interaction with the nucleoporin GmNENA at the nuclear envelope/nuclear pore complex, establishing a key signaling step that links membrane-proximal symbiotic signaling to the nuclear pore. Expression of phosphomimetic NopZ variants enhances soybean nodulation, whereas GmNENA-silencing in soybean roots reduces nodule numbers. Our findings reveal a previously unknown model for plant-microbe interactions, where the direct phosphorylation of a bacterial effector by a host receptor kinase provides an essential regulatory mechanism to direct effector localization and promote the symbiotic program.

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ORGANISM(S): Escherichia Coli

SUBMITTER: Yutao Lei

LAB HEAD: Yangrong Cao

PROVIDER: PXD076268 | Pride | 2026-04-10

REPOSITORIES: Pride

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			Action	DRS
	20210514_Z-1.index	Other
	20210514_Z-1.raw	Raw
	checksum.txt	Txt

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Project description:Soybean (Glycine max) root nodulation is a symbiotic process that requires complex molecular and cellular coordination. The phloem plays a crucial role not only in nutrient transport but also in long-distance signaling that regulates nodulation. However, the molecular mechanisms underlying phloem-specific regulation during nodulation remain poorly characterized. Here, we developed a phloem-specific Translating Ribosome Affinity Purification sequencing (TRAP-seq) system to investigate the translational dynamics of phloem-associated genes during nodulation. Using a phloem-specific promoter (Glyma.01G040700) combined with the GAL4-UAS amplification system, we successfully captured the translatome of soybean root phloem at early (72 hours post-inoculation, hpi) and late (21 days post-inoculation, dpi) nodulation stages. Differential expression analysis revealed dynamic translational reprogramming, with 2,628 differentially expressed genes (DEGs) at 72 hpi and 8,422 DEGs at 21 dpi. Gene ontology and pathway enrichment analyses showed stage-specific regulatory shifts, including early activation of ethylene and defense pathways and late-stage enhancement of nutrient transport and vascular development. Transcription factor analysis identified GmbHLH121 as a key phloem-specific regulator of nodulation. Functional validation using RNAi knockdown and overexpression experiments demonstrated that GmbHLH121 negatively regulates nodule formation, likely acting downstream of or independently from early nodulation signaling pathways. Additionally, we uncovered dynamic regulation of cell wall-modifying enzymes (PME and PMEI) in the phloem, implicating their role in modulating plasmodesmata permeability and facilitating symplastic connectivity during nodulation. Our findings highlight the critical role of phloem-mediated translational regulation in coordinating root nodulation, emphasizing the phloem as an active regulatory hub for long-distance signaling and symbiotic efficiency. This study provides a foundational framework for future research on phloem-specific gene regulation and its impact on legume symbiosis.

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Phosphorylation of rhizobial effector NopZ by soybean NORK promotes association with NENA and enhances nodulation

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