Dynamic changes of the Prf/Pto tomato resistance complex following effector recognition

Abstract

In both plants and animals, nucleotide-binding leucine-rich repeat (NLR) immune receptors play critical roles in pathogen recognition and activation of innate immunity. In plants, NLRs recognise pathogen-derived effector proteins and initiate effector-triggered immunity (ETI). However, the molecular mechanisms that link NLR-mediated effector recognition and downstream signalling are not fully understood. By exploiting the well-characterised tomato Prf/Pto NLR resistance complex, we identified the 14-3-3 proteins TFT1 and TFT3 as interacting partners of both the NLR complex and the protein kinase MAPKKKα. Moreover, we identified the helper NRC proteins (NLR-required for cell death) as integral components of the Prf /Pto NLR recognition complex. Notably our studies revealed that TFTs and NRCs interact with distinct modules of the NLR complex and, following effector recognition, dissociate facilitating downstream signalling. Thus, our data provide a mechanistic link between activation of immune receptors and initiation of downstream signalling cascades.

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

“…tomato (Pst) is a model system for understanding the molecular basis of bacterial pathogenesis and plant immunity (Martin, 2012;Oh & Martin, 2011a;Pedley & Martin, 2003). The tomato CNL protein Prf has been particularly well characterized for its role in tomato immunity to bacterial speck disease (Balmuth & Rathjen, 2007;Gutierrez et al, 2010;Kud et al, 2013;Mucyn et al, 2006;Ntoukakis et al, 2013;Pedley & Martin, 2003;Rathjen et al, 1999;Salmeron et al, 1996;Saur et al, 2015;Sheikh et al, 2023). Prf forms an immune complex with an intracellular serine/threonine kinase protein Pto, which can recognize and bind to the effectors AvrPto and AvrPtoB from race 0 strains of Pst such as DC3000 (Balmuth & Rathjen, 2007;Ntoukakis et al, 2013;Saur et al, 2015).…”

“…A screen for M3Ka-interacting proteins in tomato identified a 14-3-3 protein, Tft7, which positively regulates Prf/Pto-mediated PCD in tomato and also interacts with Mkk2, which acts downstream of M3Ka (Oh et al, 2010, Oh & Martin, 2011b. Recently, two other 14-3-3 proteins, Tft1 and Tft3, were identified as interacting partners of the M3Ka protein and the Prf/Pto complex (Sheikh et al, 2023). Another tomato M3Ka-interacting protein, Mai1, is a receptor-like cytoplasmic kinase, that acts upstream of M3Ka and positively contributes to PCD and disease resistance associated with Prf/Pto activation (Roberts et al, 2019).…”

“…Tomato has six characterized Nrc helper proteins (Nrc1, Nrc2, Nrc3, Nrc4a, Nrc4b, and Nrc4c) mediating NTI responses along with multiple sensor NLR proteins Sheikh et al, 2023;Sueldo et al, 2015;Wu et al, 2016Wu et al, , 2017Wu et al, , 2020Wu & Kamoun, 2021). In addition, NrcX appears to act as a modulator of other Nrc proteins (Adachi et al, 2023).…”

“…Reduction of Nrc2 and Nrc3 gene expression by VIGS in N. benthamiana and tomato significantly compromised Prf-/Pto-mediated resistance to Pst expressing AvrPto and AvrPtoB (Wu et al, 2016;Wu & Kamoun, 2021). Recently, there has been significant progress in elucidating the mechanism underlying the interaction between these helper NLRs and the Prf/Pto complex during the NTI response (Sheikh et al, 2023). Nrc2 and Nrc3 were found to directly interact with Prf in unchallenged leaves.…”

“…Nrc2 and Nrc3 were found to directly interact with Prf in unchallenged leaves. Upon effector recognition, Nrc2 and Nrc3 dissociate from the Prf/Pto complex, triggering downstream responses that ultimately lead to PCD and the restriction of pathogen growth (Sheikh et al, 2023). Here, to further investigate the role of Nrc2 and Nrc3 in Prf-/Ptomediated immunity, we used CRISPR/Cas9 to generate tomato lines with loss-of-function mutations in Nrc2 and Nrc3.…”