Pattern-Triggered Immunity  and Effector-Triggered Immunity: crosstalk and cooperation of PRR and NLR-mediated plant defense pathways during host–pathogen interactions

Abstract

The elucidation of the molecular basis underlying plant-pathogen interactions is imperative for the development of sustainable resistance strategies against pathogens. Plants employ a dual-layered immunological detection and response system wherein cell surface-localized Pattern Recognition Receptors (PRRs) and intracellular Nucleotide-Binding Leucine-Rich Repeat Receptors (NLRs) play pivotal roles in initiating downstream signalling cascades in response to pathogen-derived chemicals. Pattern-Triggered Immunity (PTI) is associated with PRRs and is activated by the recognition of conserved molecular structures, known as Pathogen-Associated Molecular Patterns. When PTI proves ineffective due to pathogenic effectors, Effector-Triggered Immunity (ETI) frequently confers resistance. In ETI, host plants utilize NLRs to detect pathogen effectors directly or indirectly, prompting a rapid and more robust defense response. Additionally epigenetic mechanisms are participating in plant immune memory. Recently developed technologies like CRISPR/Cas9 helps in exposing novel prospects in plant pathogen interactions. In this review we explore the fascinating crosstalk and cooperation between PRRs and NLRs. We discuss epigenomic processes and CRISPR/Cas9 regulating immune response in plants and recent findings that shed light on the coordination of these defense layers. Furthermore, we also have discussed the intricate interactions between the salicylic acid and jasmonic acid signalling pathways in plants, offering insights into potential synergistic interactions that would be harnessed for the development of novel and sustainable resistance strategies against diverse group of pathogens.

Introduction

Plant diseases are one of the main barriers restricting agricultural production globally (Jones and Dangl 2006). As per FAO (2019), plant diseases cost the global economy of around $220 billion per annum. Thus, importance must be given to the prompt management of plant diseases. Host genetic resistance deployment is seen to be the most successful management tactic. It is crucial to understand the molecular underpinnings of plant-pathogen interaction in order to develop sustainable resistance against pathogens (Kumar et al. 2021). Animals and humans respond to pathogens in different ways than plants. They have circulatory systems, as a result, they are able to relocate to the infection site and assist in protecting the targeted cells. Plants lack circulatory system; thus, each cell needs to be able to respond independently. Another important difference between plant and animal immunity is that whereas plants cannot adapt and build immunity, animals exposed to infection may (Gupta 2020). Instead, the ability to identify and combat specific diseases is hardwired into their genomes. The ability of plants to resist or repel biological attacks from pathogens is known as plant immunity (Ngou et al. 2022a, b). Two defense mechanisms are employed by plants to fend off diseases. The first is known as the "Pre-formed defense system," which uses preformed physical barriers and antimicrobial components (Kumar et al. 2021). The second one is induced defence system, which has two layers called PTI and ETI or PRR and NLR mediated plant immunity respectively. PTI, which is the first layer, is triggered by PRRs sensing PAMPs and ETI, which is the second layer, and is triggered by NLRs perceiving effectors (Jacob et al. 2013).

PRR-mediated plant immunity: perception, signal transduction and defence response

Perception of pathogens in the first line of defence depends on the interactions between host receptors and pathogen elicitors known as PAMPs or MAMPS (Microbe-associated molecular patterns). PAMPs are the conserved molecular structures shared by groups of related pathogens that are essential for the survival of these pathogens including flagellin, chitin, peptidoglycans, etc. (Kumar et al. 2011). PAMPs are recognized by plasma membrane bound proteins known as PRRs classified as RLKs (Receptor like kinases) and RLPs (Receptor like proteins) (Couto and Zipfel 2016a). Both RLKs and RLPs have an extracellular PAMP binding region, a transmembrane region, and in the case of RLKs, a cytoplasmic kinase domain that RLPs do not have (Jose et al. 2020). RLPs instead, have a condensed cytoplasmic area. Since PAMP binding domain mediate the recognition of PAMPs, RLKs and RLPs can be grouped into sub-families according to the type of PAMP binding region they have (Dievart et al. 2020). The most common PAMP binding region found in RLKs and RLPs is the leucine-rich repeat (LRR), yet, another seven PAMP binding domains have been reported, including lysin motifs (LysM), lectin, wall-associated kinases (WAK), S-locus domain, malectin-like, proline-rich, and cysteine-rich repeat (De Azevedo et al. 2021). The first PRR encoding gene reported was Cf-9 in 1994, followed by other PRR genes including Xa21, FLS2, EFR etc., that were also reported (Yu et al. 2021). Upon PAMP recognition by PRRs, PAMPs act like molecular glue and binds with and unites the extracellular PAMP binding domains of these PRRs and their co-receptors (Table 1). The extracellular PAMP binding domains of these PRRs and their co-receptors form heterodimeric complexes, which brings their kinase domain into close contact and trigger a sequence of auto and trans- phosphorylation processes (Roudaire et al. 2021). In this way, receptor complex is activated.

Table 1.

Structures of plant immune receptors or their domains

Receptor	Type: cell-surface	Plant host	Ligand	Co-receptor	References
FLS2	LRR-RLK	Arabidopsis thaliana	flg22	BAK1	(Liu et al. 2012)
CERK1	LysM-RLK	Arabidopsis thaliana	PGN	LYM3/1	(Hohmann and Hothorn 2019)
SOBIR1	LRR-RLK	Arabidopsis thaliana	Not applicable (–)	LRR-RLP, BAK1	(Lal et al. 2018)
BIR3	Pseudokinase	Arabidopsis thaliana	–	BRI1/S.E.RK1	(Hohmann et al. 2018)
BIK1	RLCK	Arabidopsis thaliana	–	BAK1, FLS2	(Liu et al. 2016)
CEBiP	LysM-RLP	Oryza sativa	Chitin	OsCERK1	(Maekawa et al. 2011)
MLA10 CC	CC-NLR	Hordeum vulgare	–	–	(De La Concepcion et al. 2018)
Pikp-1 HMA	CC-NLR	O. sativa	AVR-PikD,	Pikp-2	(Wang et al. 2019a, b)
ZAR1	CC-NLR	Arabidopsis thaliana	Avr-AC	RKS1	(Couto et al. 2016)
RRS1 WRKY	TIR-NLR	Arabidopsis thaliana	PopP2	RPS4	(Zhang et al. 2017)

The primary signaling component of PTI, RLCK (Receptor like cytoplasmic kinase), is subsequently swiftly phosphorylated and activated by this activated receptor complex. It has been demonstrated that the LRR-RLKs BAK1-LIKE1/SERK4 (BKK1/SERK4) and Brassinosteroid insensitive 1 (BRI1)-associated kinase 1 (BAK1)/Somatic embryogenesis receptor kinase 3 (SERK3) (BAK1/SERK3) are co-receptors of RLKs (Ma et al. 2018a, b). These co-receptors function for a variety of RLKs that regulate immunity, in addition to the growth and development of plants (Zhang et al. 2016). However, in order to send the signal to downstream components, LRR-RLPs and LysM-RLPs, which lack a cytoplasmic kinase domain—associate with RLKs prior to PAMP binding. Co-receptors are then recruited to form an active receptor complex only after PAMP binding has occurred (Jamieson et al. 2018). Thus, LRR-RLPs and LysM-RLPs bind to the LysM-RLK Chitin elicitor receptor 1/LysM-containing receptor-like kinase 1 (CERK1/LYK1) and the LRR-RLK Suppressor of BAK 1 interacting receptor like kinase-1/Evershed (SOBIR1/EVR), respectively (Miya et al. 2007). These co-receptors are crucial for PRR-mediated PTI and are all highly conserved in terrestrial plants (Liebrand et al. 2014). Signal is transferred across cell by a process known as signal transduction. It involves protein phosphorylation, which is catalysed by protein kinases and ultimately results in cellular response (Marks et al. 2017). A signaling pathway is created when a PAMP binds to a receptor and causes a metabolic cascade. The PAMP-receptor complex perceives a signal in the apoplast that is amplified into the cell interior by the interaction of receptor, co-receptor and RLCK (Lin et al. 2013). Trans-phosphorylation of calcium channels leading to Ca²⁺ burst is caused by phosphorylation of RLCKs such as Botrytis induced kinase 1(BIK-1), Brassinosteroid signalling kinase 1 (BSK1), BSK3, BSK5, avrPphB Sensitive 1 (PBS1), PBS1-LIKE 1 (PBL1), and PBL2. The increase in cytoplasmic Ca²⁺ is sensed by Ca²⁺ sensors like CDPKs (Calcium dependent protein kinases). The activated CDPKs together with RLCKs phosphorylates and activates the plasma membrane localised NADPH oxidase respiratory burst oxidase protein D (RbohD) (Thor et al. 2020). RbohD catalyse the transfer of electrons from cytoplasmic NADPH to molecular oxygen in the apoplast, resulting in the production of membrane impermeable apoplastic superoxide (O^2_), superoxide dismutase converts superoxide into membrane permeable H₂O₂, resulting in reactive oxygen species (ROS) burst in cytoplasm (Mansoor et al. 2022a, b). ROS causes Ca²⁺ influx channels to drive more Ca²⁺ inside cells, so elevates cytoplasmic Ca²⁺ concentration (Bogeski et al. 2011). Ca²⁺ has significant impact on initiation and amplification of ROS production while ROS is required for amplification of Ca²⁺ concentration. Also RLCKs promotes the phosphorylation and activation of MAPK (Mitogen activated protein kinase) cascade to transduce signal to nucleus. CDPKs together with MAPK activation coordinates the transcriptional reprogramming of defence-related genes through activation of various transcription factors resulting in upregulation of defence related genes, but down regulation of growth mediated by phytohormones (Li et al. 2016a, b).

So, PRR signalling pathway is tightly regulated as the excessive activation of PRRs leads to auto-immunity and growth inhibition (Albrecht et al. 2012). Plants secrete some proteins like U-box domain- containing protein 12 (PUB12) and PUB13 that mediate the polyubiquitination of PRR FLS2 (Flagellin sensitive 2), which leads to the endocytosis and degradation of this protein (Lu et al. 2011). It is also necessary to manage how PRRs and their co-receptors are activated. RLK known as BAK1-interacting receptor (BIR)-like kinase 1 binds to and sequesters BAK1 to stop auto-activation of BAK1-associated PRRs (Hohmann et al. 2018). Multiple protein phosphatases also control the phosphorylation state of PRRs. Poltergeist-like 4 (PLL4) and PLL5 connect with PRR EFR (Elongation factor receptor) in Arabidopsis and inhibit the reaction caused by PAMP elf18 (Holton et al. 2015). BAK1’s level of phosphorylation is negatively regulated by Protein phosphatase 2A (Segonzac et al. 2014). To avoid extended activation, downstream signalling components are also controlled in addition to PRRs. As a key signalling component in the PRR-signalling pathway, the Arabidopsis RLCK BIK1 is controlled by a variety of methods. Extra-large protein 2 (XLG2) inhibits the proteasome mediated degradation of BIK1 in conjugation with other heterotrimeric G proteins (Liang et al. 2016). Phosphorylation status of BIK1 is also negatively regulated by the protein phosphatase PP2C38 (Fig. 1) (Couto et al. 2016). Other PRR-signalling elements must also be controlled in addition to RLCKs. RbohD is ubiquitinated by E3 ubiquitin ligase PIRE (PBL13 interacting RING domain E3 ligase), which leads to proteasome-mediated degradation (Lee et al. 2020).

Fig. 1.

Sequence of events initiated by PRR recognition involved in plant local immunity

The phosphorylation of MPK3 and MPK6 is negatively regulated by PP2C phosphatases PP2C5 and AP2C1 (Brock et al. 2010). Auto-inhibited Ca²⁺-ATPases 8(ACA8) and its closest homolog ACA10, associates with FLS2 and participates in fine regulation of ca²⁺ level during PAMP responses. One of the important membranes associated defence responses includes the production of ROS: ROS is directly toxic to microbes. ROS cause membrane phospholipids to undergo lipid peroxidation, resulting in mixture of lipid hydroperoxides (Mansoor et al. 2022a, b). These are poisonous, damage cell membranes of pathogens and ultimately cause cell death. Phytoalexins are produced as a result of transcriptional reprogramming triggered by PTI (Chang 2012). For example, in case of pea and Monilinia fructicola (Non-pathogenic) interaction pisatin is produced, and in case of french beans and Phytophthora infestans interaction phaseolin is produced (El Hadrami et al. 2009). Primary entry points for pathogen invasion are stomata. The turgor pressure of the guard cell, which is in turn regulated by ion flux through ion channels anchored in the guard cell membrane, controls the opening and closing of the stomatal aperture (stomatal movement). Stomata closure regulation is a crucial component of PTI process at pre-invasive stage. Abscisic acid (ABA), a chemical signal that is perceived by ABA receptors (ABAR), Pyrabactin resistance 1 (PYR1), and Pyrabactin resistance like (PYL), is produced more readily in response to pathogen attack (Parwez et al. 2022). By interacting with ABA insensitive 1, a negative regulator of ABA signalling, the ABA receptor complex releases its inhibition of downstream targets, activating protein kinase open stomata 1 (OS1) in the process. ABI1 interacts with PUB12 and PUB13, two box ubiquitin E3 ligases, although these enzymes only destroy it after it interacts with ABAR in the presence or absence of ABA. Protein phosphorylation closely controls NADPH oxidase function, and ABA-activated OST1 phosphorylates NADPH oxidase directly, causing a ROS burst (Han et al. 2019). Following ABA signaling, ROS then cause an increase in cytosolic Ca² + level. Basically, for stomatal closure, guard cells need increased cytoplasmic Ca²⁺ concentration and this can be achieved by ABA dependent pathway or the way we discussed previously. Ca²⁺ in guard cells will then interact with K⁺ influx carrier and deactivates it, as a result K⁺ ion can’t be transported inside guard cells. Ca²⁺ interacts with chloride (Cl⁻), export channels and allow chloride ions to export out of guard cells. It also interacts with proton pump and inhibits function of proton pump. As a result, proton can’t be pumped out so there is high concentration of protons inside guard cells and making guard cells inside acidic and outside alkaline. K⁺ efflux channels activate, taking K⁺ outside. So osmotic imbalance is created and as a resulted water will move outside the guard cells through water channels. Guard cells become flaccid and stomata are closed.

Unlike pathogen-caused PTI where ROS burst is much more robust, beneficial microbes-induced immune responses, have been shown to be transient and relatively mild as a result of their host immune manipulation mechanisms, which are carried out in order to develop beneficial relationships with their hosts. They only produce brief stimulation of defense genes without growth inhibition by causing a tiny oxidative burst by a process known as induced systemic resistance (ISR). It relies on signalling pathways activated by ethylene and jasmonic acid (Yuan et al. 2019). Ethylene receptor (ETR) is copper ion dependent. In absence of ethylene (ET), copper associates to ETR and copper transporter (CTR) that is signal amplifier and is also associated with ETR and there is phosphorylation of histidine kinase domain which is cytosolic domain of ETR. Then phosphorylation of CTR takes place and then phosphorylates ethylene insensitive 2 (EIN2) which is transcription factor stabilizer (Li et al. 2019). Phosphorylation of EIN2 activates EIN2 targeting protein 1(ETP1). ETP1 goes and associates with EIN2 and tag poly-ubiquitin to EIN2 to cytosolic site and as a result 26S proteasome mediated EIN2 degradation. EIN3 that is transcription factor in nucleus will also be degraded by association of F box protein. Inside nucleus, F box protein binds EIN3 and E2 of F box protein poly-ubiquitinate and destroys the EIN3 by involving 26S proteasome (Potuschak et al. 2003). So there is neither EIN2 nor EIN3 present for signalling and activating ethylene responsive genes. In the presence of ethylene, ET binds to ETR, as the result CTR is cleaved out of ETR. Now CTR goes and interact with EIN2. This interaction cleaves C-terminal end of EIN2. This cleaved C-terminal end is then transported to nucleus and binds with F box protein and as a result, F box protein is occupied and there is no F box to degrade EIN3 (Qiao et al. 2012). EIN3 binds to regulatory region of ET responsive genes and that response causes synthesis of ethylene responsive proteins (Pre et al. 2008). For defence purpose, JA forms conjugate with amino acid isolusine (Ile) JA-Ile. In case of high JA-Ile concentration, JA-Ile conjugate binds to F box protein (E3 ubiquitin ligase SCF^coi1) and activates it, then activated SCF^coi1 binds to JAZ (Jasmonic acid zink domain) that is repressor of JA responsive genes and cleaves JAZ through 26S proteasome (Thines et al. 2007) resulting in initiation and regulation JA responsive gene expression through their transcription factor MYC2/3 (Kazan and Manners 2013). While in the case of low concentration of JA-Ile, MYC2/3 transcription factor of JA responsive genes binds to JAZ because SCF^coi1 is in inactive form as it does not binds with JA-Ile conjugate. So JAZ is in active form and inhibits JA responsive gene expression. Recent studies suggest that ISR increases the activity of PAL (Phenylalanine ammonia-lyase), PPO (polyphenoloxidases), peroxidases (POX), glucanase and chitinase and accumulation of ROS (Wang et al. 2015a, b). PAL, PPO and POX led to an increase in tissue lignification (Mansoor et al. 2023). Local induction of PTI often results in more robust resistance to the subsequent attack in the distal plant tissues, phenomenon known as systemic acquired resistance (SAR) (Conrath et al. 2015). Increase in the cytosolic Ca²⁺ activates master transcription factor calmodulin binding protein 60 g (CBP60g) (Qian et al. 2021), which together with SAR deficient 1(SARD1) upregulate genes involved in SA synthesis and signalling. Increase in SA concentration results in change in redox of cell. Non-expressor of pathogenesis-related genes 1(NPR1) monomerization is induced by change in redox of cell by accumulation of SA, and accumulation of NPR1 monomers in nucleus is required for the expression of PR (Pathogenesis related) genes through their interaction with transcription factors (Dong 2004). Also, SA down regulates the expression of genes encoding catalase and ascorbate peroxidase, resulting in increase in H₂O₂ concentration. H₂O₂ acts as secondary messenger for expression of PR genes (Smirnoff and Arnaud 2019).

Effector triggered susceptibility (ETS)

The coexistence of plants and pathogens that infect them, shows that they have been evolving simultaneously. Virulence of pathogen and resistance of host are kept in dynamic equilibrium, allowing both of them to survive for a considerable period of time. Identification of pathogen effectors is another method of pathogen perception. Plants that do not identify them, are susceptible leading to ETS and if identified leads to ETI. It has been demonstrated that a variety of effectors target PRR signaling pathway to inhibit PTI and cause ETS. Unless otherwise specified, effectors described in this section come from P. syringae strains. AvrPtoB, an E3 ubiquitin ligase, degrade FLS2 activity (Maekawa et al. 2011). Activated BAK1 is particularly degraded by HopB1 (Li et al. 2016a, b). AvrPto targets SOBIR1 and FLS2–BAK1 complex by preventing their kinase activities (Wu et al. 2017). EFR is directly dephosphorylated by tyrosine phosphatase HopAO1 (Macho et al. 2014). As important immunological regulators, RLCKs are target for a variety of effectors. AvrAC from X. campestris uridylylates BIK1 and PBL2 (Wang et al. 2015a, b). HopZ1a acetylates RLCKs, and AvrPphB is cysteine protease that breaks down RLCKs such as BIK1, PBS1, and PBL1 (Bastedo et al. 2019). Effectors also target other downstream PRR signalling elements. ADP ribosyl- transferase HopF2 targets both BAK1 and MKK5 to inhibit PTI signalling (Zhou et al. 2014). HopAI1 inactivates MPK3, MPK4, and MPK6 via its phosphothreonine lyase activity (Zhang and Gassmann 2007). AvrRpt2 inhibits MPK4/11 activation (Eschen-Lippold et al. 2016). In order to reduce immunity, pathogens also target transcription factors involved in defence. For example, R. solanacearum effector PopP2 acetylates and suppresses transcription factors WRKY to inhibit immunity (Zhang et al. 2017). In addition, the Verticillium dahliae effector VdSCP41 inhibits SARD1 and CBP60g to facilitate its proliferation (Qin et al. 2018).

NLR-mediated plant immunity: perception, signal transduction

When effectors are recognized by resistance (R) proteins, ETI is activated (Alhoraibi et al. 2019). R proteins have two conserved domains, central nucleotide-binding (NB) and at C-terminal leucine-rich repeat (LRR) domains, known as NLRs and a variable N-terminus domain. NLRs are divided into two groups, depending on N-terminus; (1) Toll interleukin-1-receptor NLR (TNL); (2) coiled-coil NLR (CNL). NLRs selectively identify effectors, either directly through LRR domain or indirectly via host guardee/decoy protein, and quite frequently such recognition triggers hypersensitive reaction, which is a localized defense response at the point of pathogen ingress, resulting in a programmed cell/tissue death (localized necrotic lesions) (Zavaliey et al. 2020). During coevolution, some NLRs acquired unusual integrated decoy (ID) domains for pathogen recognition. Although duration and amplitude of ETI reactions are frequently significantly greater than those of PTI reactions, PRRs and NLRs both trigger similar immune responses (Peng et al. 2018a, b). According to gene-for-gene hypothesis; NLR can detect the presence of effector which then activates plant immunity (Biezen and Jones 1998). Direct or indirect detection models can be used to describe effector recognition (Cesari et al. 2014). LRR domains of NLRs are necessary for direct effector recognition. Pathogen effectors have evolved to evade direct binding to NLRs (Cesari 2018), whereas plant coevolution to re-establish direct effector detection appears to be progressing more slowly. Nonetheless, plants have developed indirect effector recognition mechanisms. As of yet, there are more diversified and many examples of indirectly recognition in plants than there are of direct recognition. The guard model is one of the indirect effector detection system in which NLRs sends modifications to the guardee protein, a host target protein (Jones and Dangl 2006). The Arabidopsis resistance to Pseudomonas syringae pv. maculicola 1 (RPM1)-interacting protein4(RIN4) is among the most well- known example of guard model. Since RIN4 mutation constitutively triggers resistance to Pseudomonas syringae 2 (RPS2) and is fatal, RIN4 is negative regulator of immune responses and is essential for good plant growth (Axtell and Staskawicz 2003) When RIN4 is targeted by Pseudomonas syringae, effector proteins Avr-Rpm1 or AvrRpt2, the NLR RPM1 or RPS2 are activated (Kim et al. 2005). Additionally, RIN4 being cleaved by AvrRpt2 causes NLR Malus x Robusta 5 to become active in apples (Prokchorchik et al. 2020). Since pathogen effectors have evolved to minimize targeting of guardee proteins, it is difficult for host plants to change functional guardee proteins to further boost pathogen detection. Plants, on the other hand, have evolved specialized detection system that allows them to identify modified decoys. Decoy concept confines pathogen effectors to host recognition system by having plant R proteins identify effector-mediated modifications of plant decoy protein that has very similar structure to the actual host target protein (Cesari 2018). For instance, botrytis-induced kinase 1 (BIK1), a component of PTI that can be targeted and cleaved by effector AvrPphB to compromise resistance, is structurally similar to plant decoy protein, avirulence protein Pseudomonas phaseolicola B (AvrPPHB)-susceptible 1 (PBS1) (Su et al. 2018). Instead, RPS5-mediated cell death is triggered when PBS1 is cleaved by AvrPphB (De Young et al. 2012). Plants may be able to broaden the range of pathogens they can recognize with a smaller number of NLR proteins by using indirect recognition systems. For instance, pair of NLR called resistance to Ralstonia solanacearum 1 (RRS1)/RPS4 can detect presence of effectors PopP2 from Ralstonia solanacearum, AvrRps4 from Pseudomonas syringae, and unidentified effector from Colletotrichum (Birker et al. 2009). Decoy domain, which evolved into NLR RRS1 during plant evolution and contains conserved amino acid sequence WRKYGQK seen in WRKY transcription factors, was involved in these recognition events. According to Sarris et al. (2015). WRKY decoy domain is crucial target for these effectors and initiates RRS1/RPS4-mediated immunity. For NLRs in effector recognition, this integrated decoy model offers self-monitoring activity (Cesari 2018). Analysis of canonical RRS1 and R-gene analog 5 (RGA5) homologs in other plant species demonstrates that it looks to be useful tool for plants in recognizing pathogen effectors. There were numerous NLRs with one or more integrated decoys and a variety of integrated decoy kinds (Cesari 2018). In light of pathogen evolution, integrated decoy model emphasizes structural evolution of plant NLRs.

NB domain has ATP binding properties and functions as switch to activate NLR. Upon NLR activation, TIR and CC domains serve as signaling regions for the downstream response (Axtell and Staskawicz 2003). There are two types of NLRs functionally. Sensor NLRs that can identify effectors for example CNLs and TNLs. Helper NLRs that transduce signal for example ADR1 (activated disease resistance protein 1) and NRG1 (N requirement gene 1) (Espinosa et al. 2003). When an effector is recognized, NB domain of NLRs, causes them to homo-, hetero-, or oligomerize, which is crucial for downstream signalling (Van den Ackerveken et al. 1994), creating a "resistosome" of NLRs. NLRs takes on the open-lid form in response to pathogen effectors. ADP-ATP exchange causes NLR to activate. NADase activity of plant TIR generates nicotinamide adenine dinucleotide (NAD) derivatives following the development of NLR resistosome (Mansfield et al. 1994). These, are thought to be signaling molecules that activate components of signaling chain farther down the line. EDS1, "helper NLRs" NRG1 and ADR1, and enhanced disease susceptibility 1 (EDS1) are necessary for NLR signaling. Following the activation of TNLs, the EP domain containing SAG101 (Senescence associated gene 101) and EDS1 associates with NRG1similarly PAD4 (Phytoalexin deficient 4) and EDS1 associates with ADR1.These associations lead to activation of these signalling compounds, but how that is done, is under investigation. Activation of signalling compounds in turn, activate downstream immune response such as defence related gene expression and hypersensitive response (HR). The ADR1 and NRG1 act as calcium channels, so activation of these helper NLRs induce Ca²⁺ influx and trigger downstream immune response. While as CNLs depend only on helper ADR1 to function, NRG1 and ADR1 mediate ETI. The response is similar in both PTI and ETI but ETI is stronger and faster and results in HR (Torres et al. 2002). According to molecular research, the central adenosine diphosphate/adenosine triphosphate (ADP/ATP)-binding and exchange (nucleotide-binding adaptor shared by APAF-1, certain R gene products, and CED-4; NB-ARC) domains of CNL and TNL mediates the conformational activation of these proteins (Bernoux et al. 2016). The structural characterisation of pre-activated monomeric and pathogen-activated forms of the Arabidopsis CNL receptor ZAR1 was made possible in 2019 (Wang et al. 2019a, b) thanks to cryo-EM techniques. The idea that TNL receptor signaling may be mediated by a similar enzymatic action was suggested by the finding that certain plant TIR domains are NADases that can produce ribosylated cyclic nucleotide products both in vitro and in vivo. Cryo-EM structures of two TNL resistosomes, Nicotiana benthamiana Recognition of XopQ 1 (Roq1) and Arabidopsis Recognition of Peronospora parasitica 1 (RPP1), were obtained (Schultink et al. 2017). Both resistosomes are immediately triggered by cognate pathogen effector binding to the LRR domains, shown how the formation of a TIR-domain NADase enzyme is caused by the biochemical process of TNL activation (Martin et al. 2020). Three plant-related discoveries appear to be essential to advancing the field. The first is that two types of resistosomes with distinct signaling features are formed by pathogen-activated sensor CNLs and TNLs. The second is that CNL resistosomes are Ca²⁺ permeable channels in plasma membranes. As a result, immunological signal transmission within and across cells may be defined by the Ca²⁺ channel activity of sensor and helper CNL-type NLRs. The third is that two EDS1-family/CCHeLo-NLR signaling branches are activated by a set of ribosylated nucleotides produced by TNL and TIR proteins, which link cell-surface and intracellular receptor systems to immune execution. Taken together, these discoveries offer a far more comprehensive understanding of the plant defense system and new insights into the development of disease-resistant crop varieties.

ZIG ZAG model of plant immunity

The gene-for-gene hypothesis and recognition of elicitors are reconciled in zig zag model of plant immunity (Jones and dangl 2006). It depicts various stages of pathogen's interaction with its host. First, PAMPs are identified by their corresponding PRRs, which results in PTI. Second, effectors stop PAMP from being recognized, causing ETS. Thirdly, these effectors become a-virulence factor once the plant host activates a resistance gene leading to ETI. Eventually, pathogen, develops new effectors or/ and loses existing a-virulence factors to avoid ETI leading to ETS. This can continue, creating zig zag pattern between ETI and ETS (Cook et al. 2015). (Fig. 2).

Fig. 2.

The principles of plant immunity

Interaction between PTI and ETI in plant immunity

It has long been noticed, that PTI serves as the first line of defense against pathogens in an induced defense system. However, it appears that ETI didn't start working until an effector had stopped PTI. The conclusion that PTI had little impact on immune response during ETI resulted from this. Nonetheless, increasing data suggests that PTI and ETI signalling are interconnected. Most of transcriptional profiles created for PTI and ETI overlap (Tao et al. 2003). Additionally, Hatsugai et al. found that PTI inhibits an ETI signalling sector raise possibility that PTI regulates immunological signal pathways to fine tone plant defense and reduce unnecessary fitness costs (Hatsugai et al. 2017). Recent researches suggested that PRRs are necessary for the proper operation of ETI (Ngou et al. 2021). In response to PAMP flg22, effector AvrRpt2-mediated resistance was reduced in two PTI-related mutants, bbc and fec, but AvrRpt2-triggered immunity was boosted in Col-0 (Yuan et al. 2021). This suggested that ROS build up during ETI is caused by PTI signaling via PRR/co-receptors. It was also discovered that nicotinamide adenine dinucleotide phosphate oxidase activity was responsible for increased ROS in ETI. In fact, ROS generated by ETI are regulated by classic pathogen-triggered ROS gene respiratory burst oxidase homologue D (RBOHD) [22]. Furthermore, Pseudomonas syringae DC3000 that was expressing avrRpt2 caused well-known PTI-defective mutant, rbohd, to display a sensitive phenotype. These findings indicated that RBOHD serves as critical node connecting PTI and ETI (Wei et al. 2015). Detailing role of PTI protein kinase BIK1 in ETI reaction, RBOHD only generates ROS when it is phosphorylated by BIK1. By conditionally expressing effector Avr- Rps4 in Arabidopsis, Ngou et al., also discovered significant increase of PTI-responsive gene transcripts and PTI-related proteins (Ngou et al. 2021). Similar outcomes were obtained with bacterial treatment by Yuan et al., who found that AvrRpt2-triggered immunity enhanced activity of PTI components like brassinosteroid insensitive 1-associated receptor kinase 1, BIK1, mitogen-activated protein kinase 3, and mitogen-activated protein kinase 6, among others (Yuan et al. 2021). Together, these findings show that ETI signals via PTI and increases PTI response. PTI simultaneously improves ETI and is functionally crucial for ETI response. PTI and ETI work together synergistically to create strong defense against pathogen (Wei et al. 2015). PTI specifically produces anti-microbial chemicals, strengthens cell walls, and increases callose deposition to fight pathogens. ETI, meantime, sharpens PTI operation by enhancing PTI components (Peng et al. 2018a, b). These discoveries shed light on the interactions between immunological extracellular and intracellular receptors, giving us a comprehensive understanding of plant immunity. However, further research is needed to determine the mechanism through which ETI potentiates PTI. Component locations in PTI-TI crosstalk cannot be determined since the biochemical role of NLR downstream components, such as helper NLRs, EDS1, and non-race-specific disease resistance 1, is still unknown. While parts of early-diverging plants showed reduced conservation, the structural similarity of components linking PTI and ETI and, consequently, their conservation, is substantially linked with proximity in the evolutionary tree. The known functional coevolution of RNLs and EDS1/PAD4/SAG101 was represented in the comparative method. For example, in monocots, the absence of NRG1 was associated with the absence of SAG101. More significantly, we were able to begin speculating about the possible origins and recruitment of PTI and ETI as members of the plant immunity network by compiling and integrating the evolutionary conservation of signals and executors linking them. Therefore, the EDS1/PAD4/SAG101 module is a relatively new development, probably unique to seed plants, but early signaling elements like BAK1 and BIK1, along with their downstream targets for Ca²⁺ and ROS bursts, are older. However, since land plants first appeared, the network of SA synthesis, perception, and communication has remained mostly intact, indicating that SA's role in plant immunity is very old (Ramírez-Zavaleta et al. 2022).

Evidence for PTI and ETI concerted evolution in plants

Given that a recent phylogenomic analysis found a strong association between PRR and NLR gene counts across terrestrial plant species, PTI–ETI cooperation is probably widely applicable (Ngou et al. 2022a, b). Furthermore, the genomes of terrestrial plants that have adapted to aquatic, parasitic, or carnivorous environments typically contain less NLR and PRR genes (Baggs et al. 2020). Furthermore, it appears that pre-existing immunity modules were altered during the co-evolutionary pairing of functional (compatible) protein complexes between a sensor CNL HopZ-Activated resistance 1 (ZAR1) and co-functioning PTI-regulating HOPZ-ETI-DEFICIENT 1 (ZED1) cytoplasmic protein kinases (Gong et al. 2022). When combined, these findings point to a physiological foundation for the coordinated acquisition and loss of intracellular and cell surface receptor capacity during plant evolution and niche.

Role of CRISPR/Cas9 to enhance plant immunity mediated through PTI and ETI

Clustered Regularly Interspaced Short Palindromic Repeats is referred to as CRISPR/Cas9, and Cas9 is a protein that is connected to CRISPR. A repeat sequence of around 21–40 bp and spacer sequences of 25–40 bp are present in the CRISPR arrangement. According to Pourcel et al., spacer elements function as earlier invasions of foreign DNA particles that transfer resistance to phage infection, and those spacer elements have a common terminal sequence (Pourcel et al. 2005). Three steps make up the durable resistance created by CRISPR/Cas9: adaptation, expression, and interference. The adaptation stage involves cutting up foreign DNA into smaller pieces and incorporating those pieces into CRISPR displays. The production of crRNA, which directs endonucleases to cleave viral DNA by base pairing, results from the transcription of CRISPR loci (Yosef et al. 2012). For DNA interference in the type II CRISPR/Cas system, a single Cas9 protein is needed (Zaynab et al. 2020a, b). The Cas9 endonuclease contains minor RNAs such as tracrRNA and crRNA as well as multiple domains like as the HNH nuclease domain and the RuvC nuclease domain, which are located at the middle and amino terminus, respectively. Cas9 facilitates adaptation, produces crRNA by processing pre-crRNA, and generates tracrRNA-directed double-strand breaks (DSBs) in conjunction with particular double-stranded RNA (RNase III) (Jackson et al. 2014). Similar to CRISPR II, CRISPR III exhibited unique properties such as DNA and RNA cutting and association with the Cas10 nuclease. This kind of cleavage is reliant on changes in the transcription DNA sequence, which contains a strong transcription promoter (Zaynab et al. 2020a, b). When plants and fungi interact, pathogen effectors typically engage with plant R protein to initiate the ETI response, which in turn activates the plant defense mechanism. Historically, the easiest and most promising method of acquiring resistance in plants was to introduce the R gene (Idnurm et al. 2017). Furthermore, a mutation in the S gene that causes loss of function has no effect on the general health or development of the plant. Therefore, by employing CRISPRCas9 tools to target the host susceptibility genes, resistance against a number of fungal infections was established. Furthermore, a mutation in the S gene that causes loss of function has no effect on the general health or development of the plant. Therefore, by employing CRISPRCas9 tools to target the host susceptibility genes, resistance against a number of fungal infections was established. To start the infection process, fungi release enzymes that break down the cell wall of plants, making it easier for the pathogen to enter the host. Plants produce callose to strengthen their cell walls and secrete enzyme inhibitors in response to fungal pathogen invasion (Imam et al. 2016). Other defense mechanisms are also employed. Genes involved in callose deposition or inhibitors of enzymes that break down cell walls can be powerful targets for GE to build resistance to fungus. By employing CRISPR-Cas9 to target the Powdery Mildew Resistance 4 (PMR4) gene ortholog SlPMR4, which is involved in callose deposition (PRR gene), plants were genetically modified to increase resistance to the powdery mildew pathogen Oidium neolycopersici (Imam et al. 2016). The results of the CRISPR/Cas9-induced mutation in one (TaMLO-A1) of the three MLO alleles in bread wheat demonstrated enhanced resistance to Blumeria graminis f. sp. tritici infection, indicating a unique function for the TaMLO genes in powdery mildew (Wang et al. 2014). Of the 16 SlMlo genes, the most important one, SlMlo1 was altered twice, and a 48 bp cutting was obtained in tomatoes as a result. To produce plants devoid of the CRISPR/Cas9 cassette, the mutant plants were permitted to self-pollinate. "Tomelo," the recently created nontransgenic cultivar, was impervious to Oidium neolycopersici. Furthermore, no adverse impacts were observed on other genomic areas as a result of this alteration (Nekrasov et al. 2017). In an effort to genetically modify the plants and increase their features, additional susceptibility genes, such as rice Ethylene Response Factor 922 (ERF922) and enhanced disease resistance 1 (EDR1), which are involved in ethylene signaling and pathogen resistance, respectively, were also targeted (Zhang and Gassmann 2007). CRISPR-Cas9 was used to modify rice OsERF922, OsSEC3A, and the outcome was total resistance against the blast disease M. oryzae without affecting the plant's usual growth pattern (Ma et al. 2018a, b). Citrus canker disease in citrus plants is caused by Xanthomonas citri, a bacterial pathogen that is commercially significant and produces significant crop losses (Peng et al. 2017a, b). The CRISPR-Cas9 method was utilized to modify a target gene implicated in ETI in order to develop resistance against X. citri. The CsLOB1 (lateral organ boundaries 1) gene's promoter, the effectors binding element (EBE) PthA4, was altered, leading to a loss of recognition and response to bacterial effectors and an increase in resistance against infection (Jia et al. 2016). This mutation was implicated in host vulnerability. Similar to this, CRISPR-Cas9 was used to modify the tomato ortholog of Jasmonate ZIM-domain-2 (SlJAZ2), resulting in plants that are resistant to the pathogen that causes bacterial speck disease, P. syringae pv. Tomato (Ortigosa et al. 2019).

Rice has been genetically modified using CRISPR-Cas9 to mutate OsSWEET13 in order to develop tolerance against bacterial blight, which is caused by γ-proteobacterium pv. Oryzae (Zhou et al. 2015). In order to investigate PthXo2-dependent susceptibility to disease, Zhou et al. (2015, 2017) discovered a minor mutation in OsSWEET13. In the end, the mutants exhibited resistance against bacterial blight. In Wanjincheng orange, Peng et al. (2017a, b) found a connection between the promoter activity of CsLOB1 and CBC susceptibility. Trimming the EBEPthA4 sequence from CsLOB1 alleles can increase resistance to CBC.

Since viruses lack the ability translation, they hijack the transcription and translational factors systems of plants and exploit them to replicate. Thus, it is possible to develop active resistance against viruses (DNA or RNA viruses) by changing the host susceptibility genes or by focusing on the genes linked to pathogenicity. Different CRISPR-Cas9-based techniques that use the Cas9 or Cas13a nucleases have been utilized recently to genetically modify plants and provide them resistance to plant DNA viruses (Hirano et al. 2016). Three distinct research groups published the first report of employing CRISPR-Cas9 based technologies to create resistance to DNA viruses in 2015. The ability of the CRISPR tool to cause a specific mutation that weakens the virus's ability to replicate and increases plant resistance was confirmed by the authors. Baltes et al. (2015) used 11 sgRNAs to drive hairpin, Rep-joining sites, Rep designs, and BeYDV nonanucleotide series, and they reported the least amount of viral accumulation in N. benthamiana. Ali et al. (2016) investigated the mobility and replication potential of modified viruses in addition to the directing abilities of CRISPR/Cas9. RNA-directed Turnip mosaic virus (TuMV)'s RNA genome has been modified by (Aman et al. 2018). Aman et al. (2018) using the Cas13a ribonuclease within the coat protein (CP). Through CRISPR RNA editing of the GFP2 and HC-Pro genes, the very effective viral intervention was identified. This resulted in less TuMV spreading into tobacco leaves. More recently, A. thaliana evolved resistance to the Clover yellow vein virus (ClYVV) after the eIF4E1 gene was targeted using CRISPR-Cas9 (Bastet et al. 2019).

Interplay of jasmonic acid and salicylic acid signalling pathways

Ecological costs are associated with activation of plant defences (Pieterse et al. 2012). Defense mechanisms for JA and SA typically oppose one another. In addition to giving plant regulatory ability to control its immune response, this interference between phytohormone pathways causes indirect interactions between various invader types on the same plant, enabling them to make use of their limited resources for growth and survival (Van der Does et al. 2013). Increased resistance against necrotrophs is often correlated with enhanced susceptibility to biotrophs, and vice versa. Necrotrophic pathogens actively kill host tissue as they colonize and thrive on the contents of dead cells, while, as this lifestyle contrasts with that of biotrophic pathogens which derive nutrients from living cells and therefore must maintain host viability. Gymnosperms and angiosperms, have mechanism of SA–JA antagonism established. This suggests that interactions between SA and JA are conserved in angiosperms or may have existed prior to split of gymnosperms and angiosperms (Thaler et al. 2012). JA-response was shown to be inhibited by SA according to first evidence of SA–JA crosstalk in tomatoes (Pena cortes et al. 1993). When applied externally, SA has potential to suppress genes involved in JA biosynthesis (Spoel 2003). SA application had a deleterious impact on JA-dependent gene expression in lima beans (Zhang et al. 2009). In tobacco, herbivore-derived elicitors can initiate JA and ET bursts while inhibiting SA bursts. This results in coordinated interaction of SA–JA signals and, consequently, defense against herbivory (Diezel et al. 2009) (Fig. 3).

Fig. 3.

Schematic depicting the intricate crosstalk among SA, JA and ET

The 2b protein primes stimulation of SA biosynthesis by another CMV (Cucumber mosaic virus) gene product or by infection process itself, but it suppresses JA biosynthesis (Lewsey et al. 2010). JA signaling pathway in Arabidopsis is extremely antagonistic to SA, which leads to down-regulation of JA-responsive genes. JA biosynthetic pathway is the target of SA-mediated inhibition of JA-responsive gene expression, whereas ET has been shown to modify NPR1-dependency of crosstalk between SA and JA signaling (Leon-Reyes et al. 2010). SA suppresses induction of RSOsPR10 transcripts and proteins in rice roots, which is caused by JA and ET precursor ACC (Takeuchi et al. 2011). Several findings have also suggested that SA- and JA-dependent signaling interact synergistically (De Wit et al. 2013). Mur et al. (2006) found that, when SA and JA were administered together at low concentrations, there was brief synergistic enhancement in expression of genes linked to SA or JA signaling, but when one hormone was administered at a high dosage, there was an antagonistic interaction. According to a recent study, systemic resistance in roots of tomato against root knot nematode (RKN) exposed to red light at night, was partially generated by the synergistic strengthening of JA and SA defense pathways (Yang et al. 2014). The lack of inhibition of Mzmediated resistance by JA therapy suggests that there are no signaling conflicts between the two defensive mechanisms (Zinovieva et al. 2013). On vulnerable hosts, foliar administration of both SA and JA can likewise dramatically inhibit RKN development and reproduction (Dos Santos et al. 2013). Similar to this, treating Plantago lanceolate with JA and SA alone significantly lowers survival percentage of chewing-biting and piercing-sucking herbivores. However, treating Heliothis virescens with both treatments attenuates the negative effects, suggesting that these hormone pathways are linked by nodes of positive and negative crosstalk as well as convergence and divergence points (Schweiger et al. 2014). In Nicotiana benthamiana, SA and JA are also necessary for systemic resistance against TMV. The best systemic resistance against TMV was produced by the application of JA followed by SA; however, in plants lacking SA or JA biosynthesis and signaling genes, the disease was significantly more severe. It was established that SA was accumulated after TMV infection, whereas JA and MeJA were first concentrated in leaf phloem exudates (Zhu et al. 2014). The plant-pathogen relationship involves both the SA and JA signaling pathways. When SA is applied to Arabidopsis, it triggers SAR against Fusarium graminearum, which is typified by the inoculated plants' simultaneous activation of JA and SA signaling (Makandar et al. 2010). In rice, more than half of the genes induced by SA are also activated by JA, according to microarray research, demonstrating a beneficial interaction between the SA and JA pathways (Tamaoki et al. 2013). However, SA- and JA-dependent pathogen defenses were weakened when Arabidopsis was exposed to low red:far redlight ratios (De Wit et al. 2013). The description above makes it evident that there can be antagonistic or synergistic interactions between the SA and JA pathways; nevertheless, antagonistic interactions appear to be more prevalent (Dong 2004). Further discussion of NPR1, one of the most well-known proteins that is essential for controlling SA/JA cross talk, is warranted. NPR1 is a crucial transcriptional coactivator of SA-responsive PR genes as well as a transmitter of SA-induced redox state (Dong 2004). By inhibiting the JA-signaling pathway, which was partially reliant on the crosstalk modulator NPR1, SA-mediated defences induced by infection of the biotrophic virulent pathogen Pseudomonas syringae made Arabidopsis plants more vulnerable to infection by the necrotrophic pathogen Alternaria brassicicola (Pieterse et al. 2012). SA-mediated inhibition of JA signaling does not depend on NPR1's nuclear localization, suggesting that cytosolic NPR1 has a new role in regulating crosstalk between SA and JA (Spoel 2003). Furthermore, significant regulators of convergence between SA/JA crosstalk are the WRKY transcription factors. It was discovered that SA-JA crosstalk involved WRKY70, WRKY11, WRKY17, WRKY50, WRKY51, and WRKY62 among them (Spoel et al. 2007). According to Li et al. (2004) overexpression of WRKY70 increased the expression of PR genes that are responsive to SA but simultaneously suppressed the expression of PDF1.2, a marker gene that is responsive to JA, in an NPR1-dependent manner. This suggests that WRKY70 functions as a repressor of JA-responsive genes and a positive regulator of SA-induced genes. Through direct transcriptional regulation of WRKY70, an R2R3 MYB transcription factor of Arabidopsis, AtMYB44, controls the antagonistic interplay between SA and JA signaling (Shims et al. 2013). Furthermore, it was established that WRKY50 and WRKY51 proteins mediated the regulation of JA signaling by SA; in wild-type plants, exogenous SA administration suppressed the expression of JA-inducible PDF1.2, but not in plants with WRKY50 or WRKY51 mutants (Gao et al. 2011). It has been found that basal resistance is positively regulated by WRKY53 and HSPRO2, which operate downstream of SA and are adversely regulated by JA and ET signaling (Murray et al. 2007). Nuclear TGA, redox regulators, and mitogen-activated protein kinases (MAPK) are significant regulators influencing the antagonistic relationship between SA and JA-mediated signaling. MAPKs are crucial for plant innate immune signaling because they convert environmental and developmental inputs from sensors to cellular responses (Rodriguez et al. 2010). MAP kinase 4 was found to be a positive regulator of JA signaling and a negative regulator of SA signaling in Arabidopsis (Brodersen et al. 2006). Through EDS1 (increased disease susceptibility 1) and PAD4 (phytoalexin deficient 4), which operate early in the SA pathway and boost SA production, MAP kinase 4 regulates SA and JA/ET-dependent responses (Brodersen et al. 2006). GRX480 and many other ROXY classes are examples of glutaredoxins (GRXs), which are another potential regulator in SA/JA crosstalk. Because these proteins can catalyze disulfide transitions, they are crucial for mediating redox control of protein function (Spoel and Loake 2011). The defense hormones have a major impact on glutathione homeostasis. While JA lowers the glutathione pool, SA raises the amount of glutathione in cells as well as the ratio of reduced to oxidized glutathione (Zander et al. 2012). The GRXs decrease the activation of JA-responsive genes, including PDF1.2 and ORA59, and interact with TGAs which are transcription factors implicated in conferring SA- activated, NPR1-mediated defense gene expression (Zhang et al. 2003). TGAs are crucial for SAR and function as key regulators of SA-induced PR gene expression (Kesarwani et al. 2007). Numerous findings have demonstrated that TGAs interact with NPR1 and GRX480 in Arabidopsis (Zander et al. 2010). Furthermore, it is evident that TGAs are involved in the SA–JA crosstalk, as both NPR1 and GRX480 are involved in SA-mediated repression of the JA pathway. In the absence of a SA stimulus, TGAs function as positive regulators of the expression of defense genes that are dependent on JA and ET (De Lorenzo et al. 2018). The JA-responsive transcript factors EIN3 (Ethylene insensitive 3) and EIL1 (Ethylene-insensitive 3-like 1), which inhibit SA synthesis, are repressed by the JASMONATE-ZIM-DOMAIN (JAZ) proteins. Van der Does et al. (2013). found that via inhibiting the transcriptional activator ORA59, the SA pathway targeted GCC-box motifs in JA responsive promoters, hence inhibiting JA signalling downstream of the SCFCOI1-JAZ complex (Fig. 4).

Fig. 4.

Differential gene expression in CR BJN3-2

Plant epigenetics

The term 'epigenetics' has evolved since it was developed by Waddington in 1942. Initially, Waddington defined epigenetics as the mechanisms by which genes and their products influence phenotype. Epigenetics is now defined as the study of heritable changes in gene function that occur independently of changes in DNA sequence, both mitotically and/or meiotically (Armstrong 2014). These mechanisms involve intricate molecular processes that regulate gene expression without altering the underlying DNA sequence. By modulating the accessibility of genes to transcription factors and other regulatory proteins, epigenetic mechanisms enable plants to fine-tune their immune responses to combat pathogens effectively. These heritable alterations are regulated by cellular processes known as epigenetic mechanisms (Yamaguchi 2022). Among these processes, DNA methylation is one of the most thoroughly studied in plants. DNA methylation occurs when a methyl group forms a covalent link with the 5-carbon of cytosine's pyrimidine ring (C). This mechanism is critical in suppressing the activity of transposable elements (TEs), invading viral DNA, and other potentially harmful genetic elements (Erdmann and Picard 2020). Lopez et al. (2011) demonstrated that Arabidopsis RdDM mutants exhibit increased baseline resistance to the bacterial pathogen Pseudomonas syringae pv tomato DC3000 (Pst), which is associated with the priming of SA-dependent defense genes. In alignment, Yu et al. (2013) discovered greater vulnerability to Pst in the ros1-4 mutant, which corresponded to transcriptional downregulation of genes that facilitate RdDM. Dhawan et al. (2009) performed genome-wide studies of DNA methylation and gene transcription in SA- and Pst-treated Arabidopsis, demonstrating immune-related alterations in DNA methylation, especially at transposable elements (TEs), which were connected with the generation of 21-nt siRNA. These findings are consistent with those of Jaskiewicz et al. (2011), who found that systemic acquired resistance (SAR) in Arabidopsis is mediated by permissive histone post-translational modifications (PTMs) and priming of SA-dependent WRKY genes. Alternatively, heterochromatization and CHG methylation within the first intron of the R-gene RPP7 have been found to promote full-length transcription and efficient immune responses to avirulent downy mildew. This demonstrates the dual involvement of heterochromatin and DNA methylation as positive regulators of plant immunity (Li et al. 2014). The influence of stress-induced epigenetic modifications on the plant immune system's short- and long-term adaptation is revealed in a two-stage process.

Stage 1 When exposed to biotic stress, the plant's epigenome experiences changes that promote persistent up-regulation and/or priming of defense genes. This epigenetic stress memory may be passed down through generations and involves changes in the silencing of transposable elements (TEs) via DNA methylation, histone modifications, and noncoding RNAs. Stage 2 Prolonged stress increases the mutagenesis activity of functional class I ("copy and paste") and class II ("cut and paste") transposable elements, collectively known as the mobilome. As a result of this process, modest and large changes occur at excision and insertion sites (classes 1 and 2), further changing the plant's genomic response to stress. The integration of transposable elements (TEs) is influenced by the histone variant H2A.Z (Quadrana et al. 2019). TEs prefer to target environmentally sensitive genes (ERGs), which are significantly enriched with H2A.Z (Coleman-Derr and Ziberman 2012). In stage 3, TE-induced mutations enhance ERG genetic diversity, hastening the emergence of new defensive regulatory genes. Because TEs are highly epigenetically regulated, newly developed defense genes and their related pathways are still vulnerable to stress-dependent epigenetic control. This dynamic mechanism increases both the genetic and epigenetic regulatory capacity to combat biotic stress. Mutations in epigenetic regulatory pathways can have a variety of impacts on defense-response genes, causing major changes in cellular physiology and homeostasis. Ambros and his colleagues ran an experiment to see how epigenetic pathways influence the formation and evolution of a viral disease. In Arabidopsis thaliana plants, mutations affecting essential parts of two epigenetic processes led to the evolution of five distinct lineages of a naïve turnip mosaic virus (TuMV). They compared these findings to viral lineages that arose in wild-type plants. All developed lineages demonstrated adaptability to the lack of epigenetically controlled responses, with considerable increases in infectivity, pathogenicity, and viral burden. However, the degree of improvement varied according to the plant genotype. These features emerged quickly in the early passages but plateaued later on. Viral load exhibited a significant link with numerous markers of pathogenicity; however, the degree of these relationships differed across ancestral and evolved viruses. This work proposes that epigenetic regulatory mechanisms control not just host antiviral responses and viral interference with such defences, but also the development of RNA viruses. This impact may be exerted by direct contact with host factors, modification of the expression of additional resistance genes, or the production of a unique cellular milieu in comparison to wild-type plants. These mechanisms involve intricate molecular processes that regulate gene expression without altering the underlying DNA sequence. By modulating the accessibility of genes to transcription factors and other regulatory proteins, epigenetic mechanisms enable plants to fine-tune their immune responses to combat pathogens effectively (Essemine et al. 2024).

Conclusions

Plants employ a dual-layered immune system to safeguard against infections. PTI is initiated by the perception of PAMPs by PRRs, while NLRs mediate ETI in response to the perception of effectors. Both PTI and ETI signalling pathways undergo meticulous regulation to prevent autoimmunity. Contrary to the traditional view of PTI and ETI as distinct routes, emerging evidence suggests their collaborative and interdependent nature in effectively thwarting infections. Investigating the molecular underpinnings of plant defense mechanisms holds promise for enhancing development of pathogen-resistant crops. CRISPR/Cas9 is a powerful tool having gene manipulation abilities assisted in understanding plant immunity and developing crop resistance. We described how stress induced epigenetic change facilitates short- and long-term adaptation of the plant immune system. Remarkably, introducing Arabidopsis EFR into a variety of plant species, such as rice and apples, has conferred broad-spectrum resistance to diverse microorganisms. The identification of novel PRRs responsive to PAMPs or other elicitors stands as a valuable avenue for advancing the development of disease-resistant crops. The intricate interplay between jasmonic acid and salicylic acid signalling pathways in plant defense takes into consideration both synergistic and antagonistic interactions. The outcome of these SA-JA interactions is context-dependent, with instances of both positive and negative regulatory effects on defense responses. Understanding these complexities provides valuable insights for devising strategies to enhance plant immunity, while considering the ecological costs associated with the activation of defense mechanisms.

Declarations

Conflict of interest

The authors have no conflicts of interest to declare.

Ethical approval

Not applicable.