WtsE, an AvrE-Family Effector Protein from Pantoea stewartii subsp. stewartii, Causes Disease-Associated Cell Death in Corn and Requires a Chaperone Protein for Stability

Abstract

The pathogenicity of Pantoea stewartii subsp. stewartii to sweet corn and maize requires a Hrp type III secretion system. In this study, we genetically and functionally characterized a disease-specific (Dsp) effector locus, composed of wtsE and wtsF, that is adjacent to the hrp gene cluster. WtsE, a member of the AvrE family of effector proteins, was essential for pathogenesis on corn and was complemented by DspA/E from Erwinia amylovora. An intact C-terminus of WtsE, which contained a putative endoplasmic reticulum membrane retention signal, was important for function of WtsE. Delivery of WtsE into sweet corn leaves by an Escherichia coli strain carrying the hrp cluster of Erwinia chrysanthemi caused water-soaking and necrosis. WtsE-induced cell death was not inhibited by cycloheximide treatment, unlike the hypersensitive response caused by a known Avr protein, AvrRxol. WtsF, the putative chaperone of WtsE, was not required for secretion of WtsE from P. stewartii, and the virulence of wtsF mutants was reduced only at low inoculum concentrations. However, WtsF was required for full accumulation of WtsE within the bacteria at low temperatures. In contrast, WtsF was needed for efficient delivery of WtsE from E. coli via the Erwinia chrysanthemi Hrp system.

Perturbation of Maize Phenylpropanoid Metabolism by an AvrE Family Type III Effector fromPantoea stewartii

“…This system utilized a strain of E. coli MC4100 carrying the entire Dickeya dadantii hrp gene cluster in plasmid pCPP2156 (Ham et al, 1998), which becomes a WtsE delivery system (EcWtsE) when it carries a second plasmid expressing wtsEF or derivatives thereof. Previous work has indicated that WtsE, when delivered by EcWtsE, induces disease-like water-soaking symptoms in leaves of susceptible maize seedlings (Ham et al, 2006(Ham et al, , 2009. While validating the use of the EcDS to study the mechanism of action of WtsE, this study also highlighted the importance of selecting a time point that precedes potential secondary transcriptional changes associated with widespread cell death.…”

“…In nonhost plants, both DspA/E of E. amylovora and WtsE of Pnss have been shown to increase bacterial growth before bacterial population decline begins, presumably due to the onset of host defenses (Oh et al, 2007;Ham et al, 2008). Our previous work indicates that there are at least two main functions for WtsE (Ham et al, 2006(Ham et al, , 2008(Ham et al, , 2009). First, it induces a water-soaking response in maize that may release water and nutrients to support initial and continued bacterial growth.…”

“…For this purpose, we used DspA/E from E. amylovora, which we have shown can complement the virulence defect of a Pnss wtsE mutant (Ham et al, 2006). DspA/E, when delivered by DM5101, as compared with DM5101 with empty vector, induced an increased expression of TD, 4CL, C4H, P/AD, and PAL at 13 and 19 hai; these differences were statistically significant at 13 hai for all but C4H (Supplemental Fig.…”

“…The WxxxE motifs of AvrE family members are hypothesized to form part of the fold that mimics the active site of a guanine-exchange factor (GEF) protein and thus to modulate the activity of host GTPase proteins, as has been demonstrated for Map/IpgB/Sif family members from bacterial pathogens of animals (Huang et al, 2009). Confirmation of this biochemical prediction, along with localization in plant cells, have proven difficult due to the cell toxicity of the AvrE family T3Es (Ham et al, 2006). Intriguingly, HopM1, which is functionally redundant to AvrE for promoting the full virulence of P. syringae pv tomato strain DC3000, targets for degradation an ADP ribosylation factor-GEF called Arabidopsis HopMInteractor7 (AtMIN7; Nomura et al, 2006).…”

Maize Homologs of CCoAOMT and HCT, Two Key Enzymes in Lignin Biosynthesis, Form Complexes with the NLR Rp1 Protein to Modulate the Defense Response

“…WtsE is a functional type III effector secreted from Pnss. Delivering WtsE into the cells of maize seedling leaves by an Escherichia coli system induces disease-like cell death symptoms in leaves of susceptible maize seedlings and elicits numerous plant responses, including the induced expression of genes in the phenylpropanoid pathway and enhanced accumulation of coumaroyl tyramine (Ham et al, 2006(Ham et al, , 2009Asselin et al, 2015). Interestingly, HCT1806, HCT4918, and the two CCoAOMT homologs are also induced in leaves expressing WtsE (Asselin et al, 2015).…”