Titolo della tesi: Balancing defense and cooperation: the role of the apoplastic oxidoreductase CELLOX1 in shaping root–fungus communication
Plant health in natural environments relies on the dynamic interactions established with surrounding microbial communities. At the root interface, fungi engage in associations that range from mutualism to pathogenicity, profoundly influencing plant growth, nutrient status, and stress resilience. Beneficial partners foster development and tolerance to environmental challenges, while pathogenic microbes compromise fitness and productivity. To navigate this microbial diversity, plants must distinguish between compatible and antagonistic organisms, integrating immune and developmental cues to activate either defense or symbiosis. How this negotiation occurs in roots—an organ with a unique immune architecture and central role as a microbial hub—remains a key question in plant–microbe biology.
Among the mechanisms linking growth and defense, redox processes play a central role by modulating perception of biotic cues at the cell wall interface. The apoplast represents an active signaling space where plants recognize microbial or self-derived molecular patterns (MAMPs and DAMPs) and trigger pattern-triggered immunity (PTI), characterized by calcium influx, reactive oxygen species (ROS) production, and broad transcriptional reprogramming. This multilayered system constitutes the core of innate immunity and ensures effective resistance against diverse microorganisms.
In Arabidopsis thaliana, a redox-regulatory mechanism involving the apoplastic oxidase CELLOX1 fine-tunes these immune responses. CELLOX1, a berberine bridge enzyme-like protein, oxidizes specific oligosaccharides derived from cellulose and hemicellulose—cellodextrins and mixed-linked glucans—thereby reducing their elicitor activity and generating hydrogen peroxide as a secondary product. Through this reaction, CELLOX1 contributes to maintaining an appropriate immune threshold and to shaping plant–fungus communication.
Building on previous studies focused on leaves, this thesis investigates CELLOX1 function in roots, where immune and developmental programs intersect under distinct physiological conditions. Two fungal models with contrasting lifestyles were used: the beneficial endophyte Serendipita indica and the necrotrophic pathogen Bipolaris sorokiniana, allowing direct comparison of CELLOX1 function across mutualistic and pathogenic interactions.
Expression and localization analyses revealed that AtCELLOX1 is constitutively active in roots, with strongest expression at the basal and meristematic zones corresponding to fungal entry sites. Transcriptional regulation was dynamic during colonization but unresponsive to purified MAMPs or DAMPs, indicating that activation requires the physiological context of a in vivo interaction. Knockout and overexpressing lines exhibited normal root development, suggesting that CELLOX1 activity may be specifically engaged in biotic signaling rather than morphogenesis.
Functional assays demonstrated that CELLOX1 restricts fungal colonization and controls its spatial patterning. In S. indica interactions, mutants displayed broader and disorganized colonization domains, whereas overexpressing lines showed reduced fungal spread and diminished symbiotic benefits. A similar pattern was observed with B. sorokiniana, where CELLOX1 loss increased fungal proliferation and aggravated root growth inhibition. These findings reveal CELLOX1 as a general regulator of fungal accommodation rather than a determinant that discriminates between beneficial and pathogenic partners.
Analyses of key immune responses—ROS accumulation, callose deposition, and PTI marker expression—highlighted the mechanism underlying this regulation. CELLOX1 modulates the amplitude of immune activation through oxidation of wall-derived oligosaccharides, which both limits elicitor activity and produces hydrogen peroxide that acts as a signal that promotes callose deposition, adding an additional layer of defense modulation.
Overall, this study extends CELLOX1 function to roots, identifying it as a redox-based homeostatic regulator that defines the intensity and spatial boundaries of immune activation without encoding partner specificity. By connecting enzymatic redox control, apoplastic signaling, and callose-mediated defense, CELLOX1 emerges as a key integrator of growth and immunity in the root environment. These insights open new perspectives for engineering redox-regulated immune systems in crops, aiming to enhance resilience while preserving beneficial symbioses under variable environmental conditions.