Structural and functional insights into the modulation of the activity of a flax cytokinin oxidase by flax rust effector AvrL567‐A

Abstract

Summary During infection, plant pathogens secrete effector proteins to facilitate colonization. In comparison with our knowledge of bacterial effectors, the current understanding of how fungal effectors function is limited. In this study, we show that the effector AvrL567‐A from the flax rust fungus Melampsora lini interacts with a flax cytosolic cytokinin oxidase, LuCKX1.1, using both yeast two‐hybrid and in planta bimolecular fluorescence assays. Purified LuCKX1.1 protein shows catalytic activity against both N6‐(Δ2‐isopentenyl)‐adenine (2iP) and trans ‐zeatin (tZ) substrates. Incubation of LuCKX1.1 with AvrL567‐A results in increased catalytic activity against both substrates. The crystal structure of LuCKX1.1 and docking studies with AvrL567‐A indicate that the AvrL567 binding site involves a flexible surface‐exposed region that surrounds the cytokinin substrate access site, which may explain its effect in modulating LuCKX1.1 activity. Expression of AvrL567‐A in transgenic flax plants gave rise to an epinastic leaf phenotype consistent with hormonal effects, although no difference in overall cytokinin levels was observed. We propose that, during infection, plant pathogens may differentially modify the levels of extracellular and intracellular cytokinins.

Ethylene-Cytokinin Interaction Determines Early Defense Response of Wheat against Stagonospora nodorum Berk.

“…Some studies suggested that pathogens are able to alter CK levels in host plants through the effect on cytokinin oxidases/dehydrogenases (CKXs) [ 97 , 102 , 103 ] involved in the irreversible oxidation of CKs [ 104 ]. Our results showed that pathogen S. nodorum induced transcription of the CKX1 gene in wheat using NE SnTox3 specifically by gene-for-gene interactions in ethylene-dependent and ethylene-independent manners, which correlated with a low level of active CKs in wheat leaves.…”

“…However, it has not been determined as to whether this resulted from the action of effectors secreted by the pathogen. Wan et al (2019) showed that the effector AvrL567-A from the flax rust fungus Melampsora lini interacted with a flax cytosolic cytokinin oxidase, LuCKX1.1, increased its catalytic activity, and reduced the cytosolic cytokinin levels, thereby promoting the colonization of the plant [103]. Moreover, indirect activation of CKX is possible through interference with the hormonal pathways of the plant.…”

Candidate Effectors From Uromyces appendiculatus, the Causal Agent of Rust on Common Bean, Can Be Discriminated Based on Suppression of Immune Responses

“…These few were either identified as avirulence (Avr) proteins (Ellis et al, 2007; Upadhyaya et al, 2014) or directly localized inside plant cells by immunofluorescence microscopy (Kemen et al, 2005). Some of these effectors have been analyzed down to the crystal structure (Wang et al, 2007; Ve et al, 2013) and functions were inferred (Wan et al, 2019) but even for most of those effectors, little is known about their biochemical functions and their roles in virulence (Petre et al, 2014). In contrast to this, it is hypothesized that there are many effectors and that they have important functions, especially in suppression of host resistance.…”

All Roads Lead to Susceptibility: The Many Modes of Action of Fungal and Oomycete Intracellular Effectors

“…The Phytophthora capsici effector Avr3a12 interacts with FKBP15-2 and inhibits its PPIase activity in vitro ( Fan et al., 2018 ). By contrast, the interaction of the Melampsora lini effector AvrL567-A with the flax cytosolic cytokinin oxidase LuCKX1.1 was demonstrated to increase its catalytic activity against its substrates ( Wan et al., 2019 ). Structural analysis suggests that AvrL567-A binding may alter substrate access to the catalytic site, thereby modifying enzyme activity.…”