Screening of Secreted Proteins of Sporisorium reilianum f. sp. zeae for Cell Death Suppression in Nicotiana benthamiana

Abstract

Sporisorium reilianum f. sp. zeae (SRZ) is a biotrophic fungus causing head smut in maize. Maize infection with SRZ leads to very little cell death suggesting the presence of celldeath suppressinpg effectors. Several hundred effector proteins have been predicted based on genome annotation, genome comparison, and bioinformatic analysis. For only very few of these effectors, an involvement in virulence has been shown. In this work, we started to test a considerable subset of these predicted effector proteins for a possible function in suppressing cell death. We generated an expression library of 62 proteins of SRZ under the control of a strong constitutive plant promoter for delivery into plant cells via Agrobacterium tumefaciens-mediated transient transformation. Potential apoplastic effectors with high cysteine content were cloned with signal peptide while potential intracellular effectors were also cloned without signal peptide to ensure proper localization after expression in plant cells. After infiltration of Nicotiana benthamiana leaves, infiltration sites were evaluated for apparent signs of hypersensitive cell death in absence or presence of the elicitin INF1 of Phytophthora infestans. None of the tested candidates was able to induce cell death, and most were unable to suppress INF1induced cell death. However, the screen revealed one predicted cytoplasmic effector (sr16441) of SRZ that was able to reliably suppress INF1-induced cell death when transiently expressed in N. benthamiana lacking its predicted secretion signal peptide. This way, we discovered a putative function for one new effector of SRZ.

UhAVR1, an HR-Triggering Avirulence Effector of Ustilago hordei, Is Secreted via the ER–Golgi Pathway, Localizes to the Cytosol of Barley Cells during in Planta-Expression, and Contributes to Virulence Early in Infection

“…Apoplastic effectors target surface receptors or extracellular components of resistance [ 12 ]. They have also been implicated in protecting fungi from degradation or recognition by the plant immune system [ 9 ], and because they operate in the harsh apoplastic environment loaded with proteases, many have an overrepresentation of cysteine residues which allow for the formation of disulfide bridges to stabilize and protect the effector protein [ 11 , 12 , 78 ]. In contrast, cytosolic effectors are translocated inside the host cell where they interact with components of cytoplasmic immunity some of which are coded by R genes [ 9 , 11 , 12 ].…”

Estimation of the Resistance of Maize Lines to Sphacelotheca Reiliana in the Northern Steppe of Ukraine

“…S. reiliana уражує і інші види рослин, окрім кукурудзи. Проте відмічається певна специфічність та пристосованість штамів до організму-хазяїна (Poloni & Schirawski, 2016;Dutra et al, 2020).…”

Transcriptome Analysis of Fusarium–Tomato Interaction Based on an Updated Genome Annotation of Fusarium oxysporum f. sp. lycopersici Identifies Novel Effector Candidates That Suppress or Induce Cell Death in Nicotiana benthamiana

“…Cell death triggered by the pro-apoptotic mouse protein Bax in plants represents a similar defense-related cell-death response [ 19 ]. The suppression of Bax-induced cell death by co-expression of effectors in N. benthamiana with Bax has been widely used as an efficient screening tool to identify many putative fungal effectors [ 11 , 15 , 20 , 21 , 22 , 23 , 24 , 25 ]. A growing number of fungal effectors have also been found to induce cell death by transient expression in planta, and they can be initially considered “elicitors” or “toxins” or potential avirulence determinants recognized by plant resistance proteins [ 26 , 27 ].…”