Helper NLRs Nrc2 and Nrc3 act codependently with Prf/Pto and activate MAPK signaling to induce immunity in tomato

Authors: ZHANG, NING - Boyce Thompson Institute; GAN, JOYCE - Boyce Thompson Institute; CARNEAL, LAUREN - Boyce Thompson Institute; TOBON, JULIANA - Cornell University; Filiatrault, Melanie; MARTIN, GREGORY - Boyce Thompson Institute

Submitted to: The Plant Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/4/2023
Publication Date: 10/16/2023
Citation: Zhang, N., Gan, J., Carneal, L., Tobon, J., Filiatrault, M.J., Martin, G. 2023. Helper NLRs Nrc2 and Nrc3 act codependently with Prf/Pto and activate MAPK signaling to induce immunity in tomato. The Plant Journal. 117:7-22. https://doi.org/10.1111/tpj.16502.
DOI: https://doi.org/10.1111/tpj.16502

Interpretive Summary: Plants have a sophisticated immune system to defend themselves against pathogens. Plant intracellular immune receptors, primarily nucleotide-binding leucine-rich repeat proteins (NLRs) detect pathogen virulence proteins (effectors) and activate a defense response. Emerging studies indicate that some plant NLRs function in pairs or complex networks to recognize effectors and activate the plant immune responses. However, for many of these NLRs, the points at which they act to regulate plant immunity during infection are still largely unknown. In this study, the role of two helper NLRs on immunity in tomato to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) was investigated. Using tomato lines lacking several NLRs (Nrc1, Nrc2, Nrc3, and Nrc4a/c) and testing their response to different Pst strains, it was revealed that the helper NLRs Nrc2 and Ncr3 are involved in resistance to certain Pst strains and can act redundantly but also additively. Nrc2 and Nrc3 were found to alter plant cell death in plants that overexpress signaling proteins Prf and Pto, helping to identify where they might play a role in the plant defense signaling cascade. These studies provide insights into the mechanisms by which helper NLRs function in conjunction with sensor NLRs and shed light on the intricate interactions and signaling pathways involved in the plant defense response. Understanding the roles of these key proteins in plant immunity could inform the development of crops that are resistant to pathogens.

Technical Abstract: Plant intracellular immune receptors, primarily nucleotide-binding, leucine-rich repeat proteins (NLRs), can detect virulence proteins (effectors) from pathogens and activate NLR-triggered immunity (NTI). Recently, ‘sensor’ NLRs have been reported to function with ‘helper’ NLRs to activate immune responses. We investigated the role of two helper NLRs, Nrc2 (NLR required for cell death 2) and Nrc3, on immunity in tomato to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) mediated by the sensor NLR Prf and the Pto kinase. Loss-of-function mutations in both Nrc2 and Nrc3 completely compromised Prf/Pto-mediated NTI to Pst containing the cognate effectors AvrPto and AvrPtoB. An nrc3 mutant showed intermediate susceptibility between wild-type tomato plants and a Prf mutant, while an nrc2 mutant developed only mild disease symptoms. These observations indicate that Nrc2 and Nrc3 act additively to contribute to Prf/Pto-mediated immunity. We also examined at what point Nrc2/3 act in the Prf/Pto-mediated immune response. In the nrc2/3 mutant, the hypersensitive response (HR)-associated cell death induced by overexpression of the constitutive active proteins of AvrPtoB, Pto or Prf was abolished, but that induced by M3K' or Mkk2 was not. This indicates that Nrc2 and Nrc3 act upstream or independently of MAPK signaling and downstream of or together with Prf/Pto. Nrc2 and Nrc3 were not required for the HR triggered by Ptr1, another sensor NLR mediating Pst resistance, although they do appear to be involved in resistance to certain Pst race 1 strains.