Plant cell wall dynamics and wall-related susceptibility in plant–pathogen interactions

1Phytopathogenic fungi, bacteria, and nematodes infect, grow and reproduce themselves on the plant tissues and, at least at the early stages of infection, require breaking the integrity of the host cell wall. Beyond the cuticle layer, the interaction with the plant cell wall and the extent of the wall degradation are determined by the lifestyle of the pathogen. Plants perceive a diverse set of microbial molecules referred to as microbial/pathogen associated molecular patterns (MAMPs/PAMPs; ) through high-affinity cell surface pattern recognition receptors (PRRs) leading to intracellular signaling, transcriptional reprogramming, and biosynthesis of defense metabolites that limit the microbial infection. Emerging evidences indicate that plant cells also exploit sophisticated mechanisms of sensing the alteration of cell wall integrity (CWI) during biotic stress. For instance, they perceive endogenous molecules produced in damaged tissues (the so-called damage-associated molecular patterns, or DAMPs) through membrane receptors. The loss of CWI induced by pathogens activates a variety of defense responses including a cell wall remodeling required to prevent the disease. To escape recognition, pathogens produce effector proteins that counteract the plant defenses and, sometimes, exploit the host cell wall metabolism to favor the infection process.

2Infection by fungal necrotrophs is a complex process that includes conidial attachment, germination, host penetration, lesion formation and expansion, and tissue maceration followed by sporulation. Penetration may be achieved by degrading the external cuticle through the action of cutinases and lipases. The role of the cuticle in plant defense is discussed elsewhere. Once penetrated the cuticle, necrotrophs have a spatial and temporal strategy of attacking the plant cell wall by producing several cell wall degrading enzymes (CWDEs) belonging to multiple families (Figure 1A). The diversity of these enzymes mirrors the structural complexity and the dynamics of the cell wall as well as the lifestyle and host adaptation of the pathogen. The extensive degradation of cell wall polysaccharides by necrotrophs is sensed by plants. The leucine-rich repeat receptor-like kinase (LRR-RLK) ERECTA (ER) and the heterotrimeric G-protein are involved in cell wall remodeling during Arabidopsis defense response against Plectosphaerella cucumerina and probably control CWI. The impairment of cellulose synthases involved in secondary cell wall deposition is also a mechanism of sensing CWI and enhances disease resistance of Arabidopsis to P. cucumerina and Ralstonia solanacearum.

3One of the strategies used by plants to limit the degradation of the cell wall polysaccharides by microbial CWDEs is the production of proteinaceous inhibitors (Figures 1A, B). Polygalacturonases (PGs) are pathogenicity factors produced at the earlier stages of a microbial infection that depolymerize the homogalacturonan (HG), i.e., the main component of pectin in dicots but also present in monocots. Against microbial and insect PGs, plants produce cell wall-associated polygalacturonase-inhibiting proteins (PGIPs; ). The over expression of PGIPs improves the resistance to fungal and bacterial necrotrophs in different plants. The PG-PGIP interaction results in the accumulation of elicitor-active oligogalacturonides (OGs) that are perceived in Arabidopsis by the receptor Wall Associated Kinase 1 (WAK1; ) to activate the plant immune responses. Accumulation and sensing of OGs in response to pathogens is critical for monitoring the pectin integrity and, in general, a tissue injury. Alteration of pectin integrity caused by the expression of PGII from Aspergillus niger in tobacco and Arabidopsis causes a constitutive activation of defense genes and resistance against Botrytis cinerea. Recently, B. cinerea and A. niger PGs have been proposed to function themselves as PAMPs recognized by the Arabidopsis Receptor-Like Responsiveness to Botrytis PolyGalacturonase 1 (RBPG1) belonging to a super clade of LRR receptor-like proteins (RLPs; ).

4Xylan is the major hemicellulose polymer in cereals. To counteract xylan degradation by microbial endoxylanases, graminaceous monocots produce the Triticum aestivum xylanase inhibitor (TAXI), the xylanase inhibitor protein (XIP) and the thaumatin-like xylanase inhibitor (TL-XI;; ). The constitutive expression of TAXI-III in wheat reduces susceptibility to Fusarium graminearum. On the other hand, fungal xylanases function as PAMPs by eliciting defense responses and promoting necrosis. Ethylene inducing xylanases (EIXs) produced by Trichoderma species are perceived in tomato, by two specific LRR-RLPs receptors, LeEix1 and LeEix2. Both receptors bind Eixs, while only LeEix2 mediates defense responses. LeEix1 heterodimerizes with LeEix2 upon application of the Eixs and attenuates Eix-induced internalization and signaling of the LeEix2 receptor. Xyloglucan, i.e., the main hemicellulosic polysaccharide in the primary walls of dicots and non-graminaceous monocots, is degraded by microbial xyloglucan-specific endoglucanases (XEGs). Fungal XEGs are inhibited by xyloglucan endoglucanase inhibiting proteins (XEGIPs), which so far have been characterized in tomato, carrot and tobacco.

5Reinforcement of the cell wall is initiated at the pathogen penetration sites in response to cell wall damage (Figure 1B). Deposition of callose by the callose synthase PMR4 occurs upon infection of Arabidopsis with P. cucumerina and Alternaria brassicicola. Callose deposition is triggered by PAMPs and DAMPs, is affected by environmental conditions and requires the apoplastic accumulation of the hydrolysis products of glucosinolates or benzoxazinoid metabolites.

6Deposition of lignin has been associated to resistance of cotton to Verticillium dahliae and of Camelina sativa to Sclerotinia sclerotiorum. Lignin makes the cell wall more resistant to CWDEs and also prevents the diffusion of pathogen-produced toxins. The cell wall may also be reinforced by cross-links and insolubilization of structural proteins like the hydroxyproline-rich glycoproteins (HRGPs) by peroxidase-mediated isodityrosine linkages formed in response to pathogen attach. Plant peroxidases catalyze cross-links between phenolic compounds in the secondary walls and between polysaccharides and ferulic acid (FA) upon attack by necrotrophs. Crosslinks between FA and polysaccharides enhance the recalcitrance of the cell wall to digestion by microbial CWDEs and the overall resistance to fungi. On the other hand, fungal FA esterases may shear FA from the cell wall polysaccharides.

7The activities of pectin methyl esterases (PMEs) from both plants and pathogens and the degree and pattern of pectin methyl esterification are critical for the outcome of plant–pathogen infections. The cell walls containing highly methylesterified pectin are somewhat protected against the action of microbial PGs and pectate lyases (PLs; ). PMEs, which remove methyl esters from pectin, are controlled by PME inhibitor proteins (PMEIs) either during growth and development and during plant–pathogen interactions. The biochemical and structural bases of the enzyme/inhibitor interaction have been elucidated. Arabidopsis over expressing PMEIs have a lower level of PME activity, a higher degree of pectin esterification and a concomitant reduced susceptibility to B. cinerea and Pectobacterium carotovorum. The ectopic expression in wheat of a PME inhibitor from kiwi reduces the susceptibility to F. graminearum and Bipolaris sorokiniana. The transcription factor MYB46 which affect the secondary cell wall biosynthesis, regulates the expression of genes encoding several cell wall proteins including PMEI and mediates disease susceptibility of Arabidopsis to B. cinerea. Recently, jasmonic acid has been proposed to modulate the degree of methylesterification in potato to protect pectin degradation by PLs produced by Dickeya dadantii. Acetylation of the cell wall polysaccharides is also a determinant of plant–pathogen interaction. An Arabidopsis mutant with reduced acetylation displays increased tolerance to B. cinerea. Arabidopsis and Brachypodium distachyon plants expressing xylan or pectin acetylesterases from A. nidulans activate specific defense responses and are more resistant to B. cinerea and B. sorokiniana.

8Biotrophic and hemi-biotrophic fungi acquire nutrients from the host cells without causing their death. They often attack the plant surface and penetrate the external barriers by developing appressoria and exploiting the mechanical pressure. In order to breach the host cuticle they also secrete oxidases, esterases, cutinases, and lipases. Small amounts of CWDEs associated with local softening and loosening of plant cell walls are produced by biotrophic microorganisms (Figure 2A). Plants contrast invasion of biotrophs by the apposition of “papillae,” cell wall thickening early produced at the site of pathogen penetration (Figure 2B). Papillae contain callose, as most abundant constituent, cellulose, hemicelluloses, pectins, lignin, and structural proteins such as arabinogalactan proteins and HRGPs. Transgenic Arabidopsis plants overexpressing the callose synthase PMR4 show an early and elevated deposition of callose at the sites of penetration which prevents the haustoria formation and further penetration by Golovinomyces cichoracearum and Blumeria graminis. Papillae are also the sites where antimicrobial peptides, toxic secondary metabolites and reactive oxygen species (ROS) accumulate and contribute to plant resistance. Lignification and cross-links of proteins in the papillar cell wall may entrap the penetration peg of biotrophic fungi and render the cell wall more resistant to the mechanical pressure exerted by fungal appressoria. Lignin downregulation may also activate defense responses and increases the resistance to the hemibiotroph Colletotrichum trifolii in alfalfa (Medicago sativa L.).

9A reduced cellulose content by mutation of cellulose synthase CESA3, involved in primary cell wall formation, leads to production of lignin, and makes Arabidopsis more resistant to different powdery mildew pathogens. RLKs belonging to the Catharanthus roseus (CrRLK)-like protein family are implicated in CWI mechanisms. Among these, THESEUS1 (THE1) is required for lignification in response to inhibition of cellulose biosynthesis.

10Alteration of pectin integrity can trigger plant immunity also against hemibiotrophs. Arabidopsis PMEs, triggered by a Jasmonic acid dependent pathway, contribute to plant immunity against Pseudomonas syringae.

11Necrotrophs can force plants to cooperate in disease by altering the host cell wall and favoring the cell wall accessibility to CWDEs (; Figure 1C). The Arabidopsis AtPME3 is induced upon infection with B. cinerea and P. carotovorum and functions as susceptibility factor required for the initial colonization of the host tissue. A PG (LePG) and expansin (LeExp1) cooperatively contribute to cell wall loosening during tomato ripening; their expression is induced by necrotrophic pathogens to successfully infect fruits. Transgenic tomato fruits with suppressed expression of LePG and LeExp1 exhibit a reduced susceptibility to B. cinerea. Silencing of two putative endo β-1,4-endoglucanases, involved in the hydrolysis of cellulose or hemicellulose during ripening, cause a reduced susceptibility of tomato fruits to B. cinerea.

12Biotrophic fungi, at a later stage of infection, produce a limited and localized degradation of the cell wall in the epidermal or mesophyll cells. On the other hand, they form intracellular structures, like the “haustoria,” i.e., feeding organs invaginated into the host membranes to acquire nutrients. Biotrophs need to avoid the host defense responses and carefully regulate the cell-wall degradation at the border of their feeding structures to allow fungal accommodation and haustorium function (Figure 2C). A screening for Arabidopsis powdery mildew-resistant mutants allowed isolating two pectin-related genes, PMR5 and PMR6 which are pathogen-induced and required for susceptibility to G. cichoracearum and G. orontii. PMR5 encodes a protein with unknown function that shares sequence similarity with genes encoding polysaccharide O-acetyltransferase. Therefore, acetylation may be a host susceptibility mechanism that is reprogrammed by biotrophs during infection. PMR6 encodes a putative PL that is, possibly, recruited by the fungi as a susceptible factor to reduce Ca++-pectate domains at the level of haustoria-plasma membrane and facilitate cell wall porosity and accessibility of host nutrients. Callose deposition may also work in favor of the pathogen by contributing to the stability and function of the haustoria and acting as a barrier that renders haustoria less susceptible to toxic metabolites that are produced by the host and accumulate in the site of infection. On the other hand, callose may limit the diffusion of pathogen-derived elicitors, thus reducing the activation of defense responses.

13Many bacterial pathogens utilize a type III secretion system to inject effector proteins directly into the host cytoplasm and manipulate the host cellular activities to their own advantage. The effector AvrPto of P. syringae suppresses a set of Arabidopsis genes that encode cell wall-related defense proteins such as HRGPs.

14Changes in the cell wall metabolism occur during plant infection by nematodes. Like biotrophic pathogens, root-knot and cyst nematodes need to establish feeding structures inside the plant tissue to allow the uptake of nutrients. This process is assisted by the secretion of CWDEs such as pectinases and cellulases produced by the nematodes, and by the local expression of host proteins like expansins and cellulases. The sugar beet cyst nematode Heterodera schactii infects Arabidopsis roots and exploits the host-encoded AtPME3. Transgenic plants overexpressing AtPME3 exhibit an increased susceptibility to the nematode. It has been proposed that AtPME3 locally reduces the pectin esterification and improves the cell wall loosening of pre-syncytial cells during the early stages of syncytium formation.

15Callose deposition at the level of plasmodesmata (PD) limits the cell-to-cell spreading of plant viruses. Due to the small diameter of the PD pore, some viruses utilize the viral movement proteins (MPs) to modify the PD size exclusion limit. Specific interactions of viral MPs with PME are often required. In addition to MP-PME interaction, the PME-dependent formation of methanol has also been reported to be important for viral cell-to-cell movement. The overexpression of PME inhibitor proteins in tobacco and Arabidopsis contrasts the cell-to-cell and systemic movement of tobamoviruses.

16The cell wall is the battleground where plants and pathogens attempt to prevail by implementing contrasting wall-reinforcing and wall-weakening strategies. When pathogens start degrading the plant cell wall components, plants are capable of perceiving the loss of wall integrity and subsequently activate the defense signaling pathways. Pathogens try to escape the plant defenses and sometimes take advantage of the host cell wall metabolism to facilitate their entry into the tissue. These dynamic processes vary according to the lifestyle of the pathogen and the type of plant pathogen interaction. While necrotrophy involves a strong and diffused molecular warfare that may provoke extended lesions of the tissue, during biotrophy the battle involves a weaker cell wall degradation mainly localized at the point of penetration and at the level of the feeding apparatus. Perception of cell wall damage as well as the pathogen- and host-induced cell wall remodeling occurs in both cases. The damage of specific cell wall polysaccharides during infection may be perceived by receptors as THE1, ER and WAK1. Plants may also rely on the recognition of CWDEs by LRR-RLPs receptors, as RBPG1 and LeEIX1-2. Cell wall fragments may be released during infection and sensed as damage signals. Analysis of cell wall mutants has shed light on the relationship between cell wall remodeling and plant response to pathogens. The expression of endogenous and microbial CWDEs and their inhibitors is also a valuable approach for studying the dynamics of the cell wall during plant–pathogen interactions as well as a strategy to improve plant protection.

17The Review Editor, Dr Ferrari, declares that, despite being affiliated to the same institution as authors Dr Daniela Bellincampi and Dr Felice Cervone, the review process was handled objectively. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.