Distinct Amino Acids of the Phytophthora infestans Effector AVR3a Condition Activation of R3a Hypersensitivity and Suppression of Cell Death

Abstract

The AVR3a protein of Phytophthora infestans is a polymorphic member of the RXLR class of cytoplasmic effectors with dual functions. AVR3a(KI) but not AVR3a(EM) activates innate immunity triggered by the potato resistance protein R3a and is a strong suppressor of the cell-death response induced by INF1 elicitin, a secreted P. infestans protein that has features of pathogen-associated molecular patterns. To gain insights into the molecular basis of AVR3a activities, we performed structure-function analyses of both AVR3a forms. We utilized saturated high-throughput mutant screens to identify amino acids important for R3a activation. Of 6,500 AVR3a(EM) clones tested, we identified 136 AVR3a(EM) mutant clones that gained the ability to induce R3a hypersensitivity. Fifteen amino-acid sites were affected in this set of mutant clones. Most of these mutants did not suppress cell death at a level similar to that of AVR3a(KI). A similar loss-of-function screen of 4,500 AVR3a(KI) clones identified only 13 mutants with altered activity. These results point to models in which AVR3a functions by interacting with one or more host proteins and are not consistent with the recognition of AVR3a through an enzymatic activity. The identification of mutants that gain R3a activation but not cell-death suppression activity suggests that distinct amino acids condition the two AVR3a effector activities.

Evidence for intragenic recombination and selective sweep in an effector gene of Phytophthora infestans

“…In this population, Avr3a EM and virulent phenotype accounted for ~90% and ~70%, respectively. The lower phenotypic than genotypic frequency in Fujian and other three populations likely results from recessive nature of Avr3a EM or gain of function mutants (Armstrong et al., 2005; Bos et al., 2009). This result indicates that increases in air temperature during global warming may slow the appearance of Avr3a and other virulence factors in P. infestans (Wu et al., 2016).…”

“…This isoform can be recognized by potato hosts with R3a resistance; it also promotes pathogen pathogenicity by suppressing the programmed cell death of resistant host plants. Avr3a KI/147del and Avr3a KI/Y147S all demonstrate the ability to activate the hypersensitive response in plants carrying R3a resistance but do not enhance the ability of the pathogen to infect (Bos, Chaparro‐Garcia, Quesada‐Ocampo, Gardener, & Kamoun, 2009). …”

Programmed cell death in the plant immune system

“…58 Oomycete effectors can also inhibit HR in plants. [59][60][61] The mechanisms by which HR is suppressed remain unknown, but systematic characterization of the increasing number of effectors identified will help us understand how they interfere with plant defenses, including the control of HR.…”

In Planta Expression Screens ofPhytophthora infestansRXLR Effectors Reveal Diverse Phenotypes, Including Activation of theSolanum bulbocastanumDisease Resistance Protein Rpi-blb2

“…Elicitins are structurally conserved proteins in oomycetes that trigger defenses in a variety of solanaceous plants and have features of PAMPs (Nurnberger and Brunner, 2002;Vleeshouwers et al, 2006). Previously, we showed that the P. infestans RXLR effector AVR3a suppresses the cell death induced by INF1 elicitin in N. benthamiana (Bos et al, 2006(Bos et al, , 2009. To identify other RXLR effectors that suppress INF1 cell death, we infiltrated A. tumefaciens strains carrying the 62 pGR106-PexRD constructs and the negative control pGR106-DGFP (for green fluorescent protein) in N. benthamiana leaves to express the candidate suppressors.…”

“…One of the best-studied oomycete RXLR effectors is P. infestans AVR3a, which confers avirulence on potato plants carrying the R3a gene . In addition to its avirulence activity, AVR3a suppresses the cell death induced by INF1 elicitin, another secreted protein of P. infestans with features of pathogen-associated molecular patterns (PAMPs) (Bos et al, 2006(Bos et al, , 2009. AVR3a is thought to contribute to virulence through this PAMP suppression activity (Bos et al, 2009).…”