CMPG1-dependent cell death follows perception of diverse pathogen elicitors at the host plasma membrane and is suppressed by Phytophthora infestans RXLR effector AVR3a.

• Little is known about how effectors from filamentous eukaryotic plant pathogens manipulate host defences. Recently, Phytophthora infestans RXLR effector AVR3a has been shown to target and stabilize host E3 ligase CMPG1, which is required for programmed cell death (PCD) triggered by INF1. We investigated the involvement of CMPG1 in PCD elicited by perception of diverse pathogen proteins, and assessed whether AVR3a could suppress each. • The role of CMPG1 in PCD events was investigated using virus-induced gene silencing, and the ability of AVR3a to suppress each was determined by transient expression of natural forms (AVR3a(KI) and AVR3a(EM)) and a mutated form, AVR3a(KI/Y147del) , which is unable to interact with or stabilize CMPG1. • PCD triggered at the host plasma membrane by Cf-9/Avr9, Cf-4/Avr4, Pto/AvrPto or the oomycete pathogen-associated molecular pattern (PAMP), cellulose-binding elicitor lectin (CBEL), required CMPG1 and was suppressed by AVR3a, but not by the AVR3a(KI/Y147del) mutant. Conversely, PCD triggered by nucleotide-binding site-leucine-rich repeat (NBS-LRR) proteins R3a, R2 and Rx was independent of CMPG1 and unaffected by AVR3a. • CMPG1-dependent PCD follows perception of diverse pathogen elicitors externally or in association with the inner surface of the host plasma membrane. We argue that AVR3a targets CMPG1 to block initial signal transduction/regulatory processes following pathogen perception at the plasma membrane.

Phytophthora infestans effector AVR3a is essential for virulence and manipulates plant immunity by stabilizing host E3 ligase CMPG1.

Fungal and oomycete plant pathogens translocate effector proteins into host cells to establish infection. However, virulence targets and modes of action of their effectors are unknown. Effector AVR3a from potato blight pathogen Phytophthora infestans is translocated into host cells and occurs in two forms: AVR3a(KI), which is detected by potato resistance protein R3a, strongly suppresses infestin 1 (INF1)-triggered cell death (ICD), whereas AVR3a(EM), which evades recognition by R3a, weakly suppresses host ICD. Here we show that AVR3a interacts with and stabilizes host U-box E3 ligase CMPG1, which is required for ICD. In contrast, AVR3a(KI/Y147del), a mutant with a deleted C-terminal tyrosine residue that fails to suppress ICD, cannot interact with or stabilize CMPG1. CMPG1 is stabilized by the inhibitors MG132 and epoxomicin, indicating that it is degraded by the 26S proteasome. CMPG1 is degraded during ICD. However, it is stabilized by mutations in the U-box that prevent its E3 ligase activity. In stabilizing CMPG1, AVR3a thus modifies its normal activity. Remarkably, given the potential for hundreds of effector genes in the P. infestans genome, silencing Avr3a compromises P. infestans pathogenicity, suggesting that AVR3a is essential for virulence. Interestingly, Avr3a silencing can be complemented by in planta expression of Avr3a(KI) or Avr3a(EM) but not the Avr3a(KI/Y147del) mutant. Our data provide genetic evidence that AVR3a is an essential virulence factor that targets and stabilizes the plant E3 ligase CMPG1, potentially to prevent host cell death during the biotrophic phase of infection.

Towards understanding the virulence functions of RXLR effectors of the oomycete plant pathogen Phytophthora infestans.

Plant pathogens establish infection by secretion of effector proteins that may be delivered inside host cells to manipulate innate immunity. It is increasingly apparent that the ubiquitin proteasome system (UPS) contributes significantly to the regulation of plant defences and, as such, is a target for pathogen effectors. Bacterial effectors delivered by the type III and IV secretion systems have been shown to interact with components of the host UPS. Some of these effectors possess functional domains that are conserved in UPS enzymes, whilst others contain novel domains with ubiquitination activities. Relatively little is known about effector activities in eukaryotic microbial plant pathogens. Nevertheless, effectors from oomycetes that contain an RXLR motif for translocation to the inside of plant cells have been shown to suppress host defences. Annotation of the genome of one such oomycete, the potato late blight pathogen Phytophthora infestans, and protein-protein interaction assays to discover host proteins targeted by the RXLR effector AVR3a, have revealed that this eukaryotic plant pathogen also has the potential to manipulate host plant UPS functions.