MrHex1 is Required for Woronin Body Formation, Fungal Development and Virulence in Metarhizium robertsii

Abstract

The Woronin body (WB) is a peroxisome-derived dense-core vesicle, a self-assembling hexagonal crystal of a single protein Hex1. This organelle is specific to the ascomycete fungi belonging to the Pezizomycotina subphylum by functioning in sealing septal pores in response to mycelium damage and the control of cell heterogeneity. We retrieved all available Hex1-domain containing proteins of different fungi from the GenBank database and found considerable length variations among 460 obtained Hex1 proteins. However, a highly conserved Hex1 domain containing 75 amino acid residues with a specific S/A-R/S-L consensus motif for targeting peroxisome is present at the carboxy-terminus of each protein. A homologous Hex1 gene, named MrHex1, was deleted in the entomopathogenic fungus Metarhizium robertsii. It was found that MrHex1 was responsible for WB formation in M. robertsii and involved in sealing septal pores to maintain cell integrity and heterogeneity. Different assays indicated that, relative to the wild-type (WT) strain, ∆Mrhex1 demonstrated a growth defect on a solid medium and substantial reductions of conidiation, appressorium formation and topical infectivity against insect hosts. However, there was no obvious virulence difference between WT and mutants during injection of insects. We also found that ∆MrHex1 could tolerate different stress conditions like the WT and the gene-rescued mutant of M. robertsii, which is in contrast to the reports of the stress-response defects of the Hex1 null mutants of other fungal species. In addition to revealing the phenotypic/functional alterations of the Hex1 deletion mutants between different pathotype fungi, the results of this study may benefit the understanding of the evolution and WB-control of fungal entomopathogenicity.

Hex1, the Major Component of Woronin Bodies, is Required for Normal Development, Pathogenicity and Stress Response in the Plant Pathogenic FungusVerticillium dahliae

“…In the absence of Hex1, we observed reduced germination of conidia and growth rate upon treatment with either calcofluor white or Congo red, which indicates that changes in cell wall structure and related defects in hyphal integrity possibly result in impaired germination. Contrasting results have been reported as regards to the sensitivity of Δ hex1 mutant strains of other fungi to osmotic and cell wall/plasma membrane stress; deletion of the gene did not significantly affect M. robertsii and A. flavus [25,26], whereas in A. fumigatus and Arthrobotrys oligospora it had a significant impact [13,24].…”

“…The major component of WBs was shown to be the conserved peroxisomal protein Hex1 in N. crassa [15,17]. Its discovery was followed by detailed analyses of its structure and functions, mostly in this and to a lesser extent in other fungi [13,16,18,20,26]. In this study, we identified the single Hex1 homolog of V. dahliae , a plant pathogen with a particularly broad range of hosts and significant economic impact [28].…”

“…The expression of two protein isoforms has also been experimentally demonstrated in other species using Hex1-specific antibodies [17,18,21], which corroborates our suggestion. The inaccuracy of automatic gene annotation in this case might explain the predicted hypervariable N-terminal extensions observed in many sequences from different species [25,26], indicating that this data should be treated with caution.…”

“…Deletion of hex1 in several fungi generally led to excessive hyphal bleeding after wounding [20,27], but also had pleiotropic effects on additional phenotypes associated with vegetative growth, asexual reproduction and stress response against osmotic and cell wall-perturbating agents [24][25][26]. Regarding pathogenicity and virulence, conflicting results have been gathered from studies on plant, insect and human pathogens.…”

Dynamic Regulation of Peroxisomes and Mitochondria during Fungal Development

“…An involvement for Woronin bodies during conidiation has been exposed in Fusarium graminearum (Gibberella zeae) , a devastating pathogen of cereals, where either the deletion or overexpression of HEX1 resulted in a significant reduction in conidiation [ 162 ]. Similarly, loss of Woronin bodies results in reduced conidiation in the nematophagous A. oligospora [ 163 ], in Aspergillus flavus [ 164 ] and the entomopathogenic M. robertsii [ 165 ], whereas in A. nidulans [ 166 ] and N. crassa [ 160 ] it has a minor impact in conidiation. Moreover, no visible conidiation defects are produced by Hex1 elimination in fungi like M. oryzae [ 167 ] and A. fumigatus [ 168 ].…”