Transposition of the miniature inverted‐repeat transposable element mimp1 in the wheat pathogen Fusarium culmorum

Abstract

High-throughput methods are needed for functional genomics analysis in Fusarium culmorum, the cause of crown and foot rot on wheat and a type B trichothecene producer. Our aim was to develop and test the efficacy of a double-component system based on the ability of the impala transposase to transactivate the miniature inverted-repeat transposable element mimp1 of Fusarium oxysporum. We report, for the first time, the application of a tagging system based on a heterologous transposon and of splinkerette-polymerase chain reaction to identify mimp1 flanking regions in the filamentous fungus F. culmorum. Similar to previous observations in Fusarium graminearum, mimp1 transposes in F. culmorum by a cut-and-paste mechanism into TA dinucleotides, which are duplicated on insertion. mimp1 was reinserted in open reading frames in 16.4% (i.e. 10 of 61) of the strains analysed, probably spanning throughout the entire genome of F. culmorum. The effectiveness of the mimp1/impala double-component system for gene tagging in F. culmorum was confirmed phenotypically for a putative aurofusarin gene. This system also allowed the identification of two genes putatively involved in oxidative stress-coping capabilities in F. culmorum, as well as a sequence specific to this fungus, thus suggesting the valuable exploratory role of this tool.

FcStuA from Fusarium culmorum Controls Wheat Foot and Root Rot in a Toxin Dispensable Manner

“…The lack of significant amino acid differences between the two sequences indicated an identical structure of the two proteins. When comparing the similarity of FgStuA and FcStuA to a general level of similarity between the two species, at least with partial data obtained from a transposon tagging approach [53], the level of conservation seems to be higher than average: 98.5% vs 96%. It is therefore suggested that FcStuA is under strong selective pressure having a crucial role in cell regulation and therefore being highly conserved between the two closely related species.…”

Fusarium culmorum: causal agent of foot and root rot and head blight on wheat

“…The availability of genomes would facilitate targeted functional genomics studies that, at the moment, are based on the similarities of genes with F. graminearum (Baldwin et al, 2010), but this cannot explore genes that are peculiar to F. culmorum (Spanu et al, 2012).…”

FcRav2, a gene with a ROGDI domain involved in Fusarium head blight and crown rot on durum wheat caused by Fusarium culmorum

“…Identification of the reinsertion site of mimp1 was achieved by Splinkerette‐PCR (Potter and Luo, ), according to the protocol described by Spanu et al . (). Flanking sequences were blast ed to the F. culmorum genome (Urban et al ., ) in order to identify mimp1 locations.…”

“…A mimp1-mediated revertant (see below) strain collection was generated from the F. culmorum FcM7 co-transformant strain, integrating a single copy of the niaD::mimp1 construct and the impalaE transposase gene under the constitutive control of the gpdA promoter (Dufresne et al, 2007;Spanu et al, 2012).…”

“…With the aim to identify new F. culmorum genes playing a role in both FHB and FRR on durum wheat, we performed a transposon-mediated mutagenesis approach based on the heterologous element mimp1 (Hua- Van et al, 2000). mimp1 does not encode a transposase gene, but can be trans-mobilized by the transposase of the impala element (Dufresne et al, 2007(Dufresne et al, , 2008Spanu et al, 2012).…”