A Poplar Rust Effector Protein Associates with Protein Disulfide Isomerase and Enhances Plant Susceptibility.

Mst Hur Madina,Yang Zhang,Hugo Germain,Md Saifur Rahman,Xiaoqiang Huang,Huanquan Zheng

doi:10.3390/biology9090294

Abstract

Melampsora larici-populina (Mlp), the causal agent of Populus leaf rust, secretes an array of effectors into the host through the haustorium to gain nutrients and suppress immunity. The precise mechanisms by which these effectors promote virulence remain unclear. To address this question, we developed a transgenic Arabidopsis line expressing a candidate effector, Mlp124357. Constitutive expression of the effector increased plant susceptibility to pathogens. A GxxxG motif present in Mlp124357 is required for its subcellular localization at the vacuolar membrane of the plant cell, as replacement of the glycine residues with alanines led to the delocalization of Mlp124357 to the nucleus and cytoplasm. We used immunoprecipitation and mass spectrometry (MS) to identify Mlp124357 interaction partners. Only one of the putative interaction partners knock-out line caused delocalization of the effector, indicating that Arabidopsis protein disulfide isomerase-11 (AtPDI-11) is required for the effector localization. This interaction was further confirmed by a complementation test, a yeast-two hybrid assay and a molecular modeling experiment. Moreover, localization results and infection assays suggest that AtPDI-11 act as a helper for Mlp124357. In summary, our findings established that one of Mlp effectors resides at the vacuole surface and modulates plant susceptibility.

Highlights

During infection, plant pathogenic microbes deliver virulence proteins, known as effectors, into host cells to overcome plant immunity and promote parasitic colonization through the manipulation of cellular processes [1]
We provide evidence through mass spectrometry, a genetic complementation test, a yeast-two hybrid assay (Y2H), and in silico modeling that Mlp124357 associates with a protein disulfide isomerase (PDI) which acts as a helper protein for this effector but not for other Melampsora larici-populina (Mlp) effectors having similar numbers of disulfide bridges
To select candidate secreted effector proteins for functional investigation, we followed various criteria that were previously described [5,11]: these included that the sequences must be of small size, possess a signal peptide and conserved cysteines, and must not have conserved sequences outside the order Pucciniales; they must be detected in infection structures

Summary

Introduction

Plant pathogenic microbes deliver virulence proteins, known as effectors, into host cells to overcome plant immunity and promote parasitic colonization through the manipulation of cellular processes [1]. Effectors traffic to various cellular compartments where they interact with host proteins or nucleic acids and exert their virulence function [2,3,4,5,6]. To target these destinations, effectors possess domains or motifs in their sequence; for example, nucleus localized effectors can contain nuclear-localized signals (NLS) and some chloroplast localized effectors may carry a transit peptide [4,7,8]. Biology 2020, 9, 294 in Arabidopsis thaliana and Nicotiana benthamina are extensively used in the functional investigation of effector biology [3,5,10,11,12].

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Results

Conclusion