The Ups and Downs of Plant NLR Expression During Pathogen Infection

Abstract

Plant Nucleotide binding-Leucine rich repeat (NLR) proteins play a significant role in pathogen detection and the activation of effector-triggered immunity. NLR regulation has mainly been studied at a protein level, with large knowledge gaps remaining regarding the transcriptional control of NLR genes. The mis-regulation of NLR gene expression may lead to the inability of plants to recognize pathogen infection, lower levels of immune response activation, and ultimately plant susceptibility. This highlights the importance of understanding all aspects of NLR regulation. Three main mechanisms have been shown to control NLR expression: epigenetic modifications, cis elements which bind transcription factors, and post-transcriptional modifications. In this review, we aim to provide an overview of these mechanisms known to control NLR expression, and those which contribute toward successful immune responses. Furthermore, we discuss how pathogens can interfere with NLR expression to increase pathogen virulence. Understanding how these molecular mechanisms control NLR expression would contribute significantly toward building a complete picture of how plant immune responses are activated during pathogen infection-knowledge which can be applied during crop breeding programs aimed to increase resistance toward numerous plant pathogens.

Keywords: NB-LRR; NLR; NLR expression; cis elements; epigenetics; pathogen infection; transcriptional regulatinon.

Figure 1

Schematic representation of NLR protein domains and NLR activation, together with NLR protein examples. (A) Different structures of NLR proteins identified in plants, with NLR protein examples listed on the right of each schematic. (B) Models of Nucleotide binding-Leucine rich repeat (NLR) protein recognition of pathogen Avirulence (Avr) proteins. (1) Direct recognition of pathogen Avr protein through binding to LRR domains of NLR proteins. Avr recognition is followed by the exchange of ADP with ATP at the Nucleotide binding (NB) domain, which activates the protein and downstream immune responses. (2) Pathogen Avr proteins may bind to guard proteins which are under the surveillance of NLR proteins. Once Avr binding is recognized the NLR protein is activated through the binding of ATP, ultimately leading to immune response activation. (3) Avr recognition by NLR dimer pairs occurs when an Avr proteins binds to a sensor NLR. Structural changes of the sensor NLR induced by the Avr activates a helper NLR, subsequently leading to immune response activation. The binding of ATP to the sensor NLR is not required for NLR function (ADP—Adenosine diphosphate; ATP—Adenosine triphosphate; CC—Coiled coil; CC_R—Coiled coil domain with integrated RPW8 domain; LRR—Leucine rich repeat; NB—Nucleotide binding; TIR—Toll/interleukin-1 receptor).

Figure 2

Schematic illustration of the main regulatory mechanisms of plant NLR expression. (A) Before pathogen infection, a heterochromatin structure is maintained by histone methylation marks H3K9me2 and H3K9me3, which suppresses Nucleotide binding-Leucine rich repeat (NLR) expression. Histone deacetylases (HDAC) also contribute to a heterochromatin structure. The H3K9me3 mark is also associated with DNA methylation of CG, CHG, and CHH sites, established by the RNA-directed DNA methylation pathway. De novo DNA methylation is guided by small interfering RNA (siRNA) in association with Argonaute 4 (AGO4), Domains rearranged methyltransferase 2 (DRM2), and RNA-dependent DNA methylation 1 (RDM1) proteins. DNA methylation is further maintained by DNA methyltransferase 1 (MET1), CMT3 (Chromomethylase 3), and CMT2/DRM2. (B) Following pathogen infection, a euchromatin structure is adopted which allows for the activation of NLR expression. Histone marks H3K3me3 and H3K36me2/3 are established by Histone methyl transferases (HMT), H3K9ac by Histone acetyltransferases (HAT), and H2Bub1 by Histone monoubiquitination 1 and 2 (HUB1/2). Repressor of silencing 1 (ROS1) and DEMETER enzymes (DME) antagonize the RNA-directed DNA methylation pathway, and lower levels of DNA methylation is observed. (C) A euchromatin structure allows for Transcription factors (TFs) to bind to cis elements within NLR promoter sequences located upstream from the Transcription start site (TSS). Most TFs are activated following stress hormone (H) detection, but may also be activated by pathogen effectors (E). The four most common cis elements identified within NLR promoter sequences include W-boxes, ABRE, MYB, and MYC elements. (D) Following NLR expression, alternative splicing (AS) patterns may contribute to different NLR mRNA isoforms, and thus, different levels of NLR proteins. AS may produce mRNAs containing (1) different exons, (2) different Untranslated regions (UTRs), (3) or a retained intron which may code for a stop codon, producing a truncated NLR protein following translation. (E) MicroRNA (miRNA) molecules in association with AGO1 can downregulate NLR expression by binding to NLR mRNAs to either block mRNA translation or cause mRNA degradation. Phased secondary RNA (phasiRNA) molecules can also be produced when diced NLR mRNAs are reverse transcribed by RNA-directed RNA polymerase 6 (RDR6) and diced by Dicer-like 4 (DCL4). These phasiRNA molecules may then target more NLR mRNA molecules to further contribute to NLR suppression (DCL3—Dicer-like 3; Pol IV—RNA polymerase IV).