Comparative structural and functional characterization of putative protein effectors belonging to the PcF toxin family from Phytophthora spp

Abstract

The PcF Toxin Family (Pfam 09461) includes the characterized phytotoxic protein PcF from Phytophthora cactorum, as well as several predicted protein effectors from other Phytophthora species recently identified by comparative genomics. Here we provide first evidence that such 'putatives', recombinantly expressed in bacteria and purified to homogeneity, similarly to PcF, can trigger defense-related responses on tomato, that is leaf withering and phenylalanine ammonia lyase induction, although with various degrees of effectiveness. In addition, structural prediction by computer-aided homology modeling and subsequent structural/functional comparison after rational engineering of the disulfide-structured protein fold by site-directed mutagenesis, highlighted the surface-exposed conserved amino acid stretch SK(E/C)C as a possible structural determinant responsible for the differential phytotoxicity within this family of cognate protein effectors.

Mechanisms and Evolution of Virulence in Oomycetes

“…The related PcF (Phytophthora cactorum factor) and Scr (secreted cysteine-rich) toxin families are specific to oomycetes and show much greater heterogeneity among species (39,59,69,116). The numbers of PcF/Scr family genes in Phytophthora and Pythium genomes are highly heterogeneous (39,55,69,116).…”

“…The numbers of PcF/Scr family genes in Phytophthora and Pythium genomes are highly heterogeneous (39,55,69,116). The PcF subfamily is expanded in P. sojae (16 genes), and the Scr family is expanded in P. infestans (11 genes), whereas P. ramorum and Py.…”

“…Several large families of proteins that trigger cell death in host plants have been found in oomycete genomes, including the NLPs (necrosis and ethylene inducing peptide-like proteins) (76), the PcF family (69,70), and the Scr family (59). NLPs trigger cell death in a wide range of dicotyledonous hosts, producing a response with many similarities to PTI (33, 78).…”

SCR96, a small cysteine‐rich secretory protein of Phytophthora cactorum, can trigger cell death in the Solanaceae and is important for pathogenicity and oxidative stress tolerance

“…PCD induced by phytotoxins is thought to play a central role in providing a source of nutrition for necrotrophic and hemibiotrophic pathogens (Dou and Zhou, ; Mott et al ., ). However, such a response caused by different PcF/SCR effectors shows various degrees of effectiveness (Orsomando et al ., ). In our case, we observed no changes visible to the naked eye when some of the PcF/SCRs, e.g.…”

“…It has been suggested that PcF may mimic the effects of a plant signalling protein because it seemingly has structural homology to the pollen protein Ole-e6 with allergenic activity (Nicastro et al, 2009). Oomycetic SCR proteins can be related to PcF mainly on the basis of their conserved sequences, cysteine patterns and N-terminal secretory signals (Bos et al, 2003;Orsomando et al, 2011). The expansion of PcF/SCR gene families in oomycetes has been noted in whole-genome analyses and the size of the family in several species, such as P. infestans (n = 16), P. sojae (n = 19), P. ramorum (n = 4), Pythium ultimum (n = 3) and Hyaloperonospora arabidopsidis (n = 2), has been estimated through annotation (Baxter et al, 2010;Haas et al, 2009;Lévesque et al, 2010;Tyler et al, 2006).…”