Genome‐wide functional analysis of hot pepper immune receptors reveals an autonomous NLR clade in seed plants

Abstract

Plants possess hundreds of intracellular immune receptors encoding nucleotide-binding domain and leucine-rich repeat (NLR) proteins. Autoactive NLRs, some cases a specific domain of NLR, induce plant cell death in the absence of pathogen infection. In this study, we identified a group of NLRs (ANLs; ancient and autonomous NLRs) carrying autoactive coiled-coil (CC A) domains in pepper (Capsicum annuum L.) by genome-wide transient expression analysis. The CC A-mediated cell death mimics hypersensitive cell death triggered by interaction between NLR and pathogen effectors. Sequence alignment and mutagenesis analyses revealed that the intact α1 helix of CC A is critical for both CC A-and ANL-mediated cell death. The cell death induced by CC A does not require NRG1/ADR1 or NRC type helper NLRs, suggesting ANLs may function as singleton NLRs. We also found that CC A localize in the plasma membrane as Arabidopsis singleton NLR ZAR1. Extended studies revealed that autoactive CC A s are well conserved in other Solanaceae plants as well as a monocot plant, rice. Further phylogenetic analyses revealed that ANLs are present in all tested seed plants (spermatophytes). Our studies not only uncover the autonomous NLR clade in plants but also provide powerful resources for dissecting the underlying molecular mechanism of NLR-mediated cell death in plant.

RefPlantNLR: a comprehensive collection of experimentally validated plant NLRs

“…Phylogenetic analyses would help assign NLRs to subclades and provide a basis for generating hypotheses about the function and mode of action of novel NLRs which phylogenetically cluster with experimentally validated NLRs. This type of phylogenetic information can be combined with other features such as genetic clustering and has, for instance, proven valuable in previous work on rice and solanaceous NLRs (Białas et al ., 2017; Wu et al ., 2017) and for defining the CC G10 -NLR class (Lee et al ., 2020).…”

“…In flowering plants (angiosperms), NLRs form three main monophyletic groups with distinct N-terminal domain fusions: the TIR-NLR subclade containing an N-terminal Toll/interleukin-1 receptor (TIR) domain, the CC-NLR-subclade containing an N-terminal Rx-type coiled-coil (CC) domain and the CC R -NLR subclade containing an N-terminal RPW8-type CC (CC R ) domain (Shao et al ., 2016). Additionally, Lee et al . (2020) have recently proposed that the G10-subclade of NLRs is a monophyletic group containing a distinct type of CC (here referred to as CC G10 ; CC G10 -NLR).…”

Comparative analysis of de novo genomes reveals dynamic intra-species divergence of NLRs in pepper

“…In particular, G1 and G2, the largest subgroups in pepper NLRs [ 21 ], exhibited variable copy numbers among all accessions (Supplemental Table 10 ). On the other hand, GT, which includes TIR-NLR (TNL), and G10, referred to as the ancient and autonomous NLR (ANL) [ 22 ], had a moderate number of NLR subgroups and moderate CNVs. Moreover, GR, which includes RPW8-type helper NLR (RNL), and G8, which contains the NLRs required for cell death (NRC) helper group [ 23 ], had small numbers of NLRs and the fewest CNVs.…”

Plant NLR targets P-type ATPase for executing plasma membrane depolarization leading to calcium influx and cell death

“…Previously, we reported an Autonomous and Ancient NLR group (ANL) comprising CC domains conferring HR-mimic cell death in seed plants 7 . To take experimental advantages of small size and simple structure, CC A 309, an autoactive CC domain in pepper ANL, was adopted to identify novel immune components underlying NLR-mediated cell death.…”