Abstract

The mycotoxin deoxynivalenol (DON) acts as a disease virulence factor for Fusarium fungi, and tolerance of DON enhances wheat resistance to Fusarium head blight (FHB) disease. Two variants of an ATP-binding cassette (ABC) family C transporter gene were cloned from DON-treated wheat mRNA, namely TaABCC3.1 and TaABCC3.2. These represent two of three putative genes identified on chromosomes 3A, 3B, and 3D of the wheat genome sequence. Variant TaABCC3.1 represents the DON-responsive transcript previously associated with DON resistance in wheat. PCR-based mapping and in silico sequence analyses located TaABCC3.1 to the short arm of wheat chromosome 3B (not within the FHB resistance quantitative trait locus Fhb1). In silico analyses of microarray data indicated that TaABCC3 genes are expressed in reproductive tissue and roots, and in response to the DON producer Fusarium graminearum. Gene expression studies showed that TaABCC3.1 is activated as part of the early host response to DON and in response to the FHB defence hormone jasmonic acid. Virus-induced gene silencing (VIGS) confirmed that TaABCC3 genes contributed to DON tolerance. VIGS was performed using two independent viral construct applications: one specifically targeted TaABCC3.1 for silencing, while the other targeted this gene and the chromosome 3A homeologue. In both instances, VIGS resulted in more toxin-induced discoloration of spikelets, compared with the DON effects in non-silenced spikelets at 14 d after toxin treatment (≥2.2-fold increase, P<0.05). Silencing by both VIGS constructs enhanced head ripening, and especially so in DON-treated heads. VIGS of TaABCC3 genes also reduced the grain number by more than 28% (P<0.05), both with and without DON treatment, and the effects were greater for the construct that targeted the two homeologues. Hence, DON-responsive TaABCC3 genes warrant further study to determine their potential as disease resistance breeding targets and their function in grain formation and ripening.

Highlights

  • GAGTGGAATCGAAGAAAGACATAGGAGCTCTG AGTAGCTGTGCTATCAACACGAATGGTACGAG GAGATCGTGCATGTACCATGAGAAGAGG CTTCCTCAGCGTCCTCACCTTCTCCT/ TACCTGGATCGATGATACCAGTTGGTAGACTG CTCTTCTCATGGTACATGCACGATCTCTG/ CCTGGATAGGCTTAAAGAGATTGTTTCACTGC CTTCCTCAGCGTCCTCACCTTCTCCT/ CTACGCGTGCGTCGCTCTGATCGTGTACT GCTCCACTCACCCAGCCTGC/ ACCGCAACGGCTGTTCTGTTCA GCCAACGAATAGATCCTCCA/ ACCACCTTCAGCCTTTTCCT GCACAGGGAATCGTCAAAGT/ CACACATAGCTTCATTCGTTCA

  • By computational mapping of the TaABCC3.1 or TaABCC3.2 mRNA sequences to wheat genomic homologues, both TaABCC3 genes were shown to consist of 10 exons and 9 introns

  • The closest homologue is AetABCC3 from the wheat progenitor Aegilops tauschii [GenBank ID: gi|475536653; TaABCC3.1: 1229/1251aa (98%) identity, 7/1251aa (0%) gaps; TaABCC3.2: 1240/1252aa (99%) identity, 7/1252aa (0%) gaps]

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Summary

Introduction

GAGTGGAATCGAAGAAAGACATAGGAGCTCTG AGTAGCTGTGCTATCAACACGAATGGTACGAG GAGATCGTGCATGTACCATGAGAAGAGG CTTCCTCAGCGTCCTCACCTTCTCCT/ TACCTGGATCGATGATACCAGTTGGTAGACTG CTCTTCTCATGGTACATGCACGATCTCTG/ CCTGGATAGGCTTAAAGAGATTGTTTCACTGC CTTCCTCAGCGTCCTCACCTTCTCCT/ CTACGCGTGCGTCGCTCTGATCGTGTACT GCTCCACTCACCCAGCCTGC/ ACCGCAACGGCTGTTCTGTTCA GCCAACGAATAGATCCTCCA/ ACCACCTTCAGCCTTTTCCT GCACAGGGAATCGTCAAAGT/ CACACATAGCTTCATTCGTTCA Supplemental Results: Characterisation and phylogeny of the TaABCC3 genes and their encoded proteins. After mapping of all cloned and downloaded TaABCC3.2 transcript sequences (N=91) to the TaABCC3.2 gene the mRNA length was determined to be at least 4992bp.

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