CRISPR/Cas9-Mediated Mutagenesis of Two Alleles Encoding a Glutathione S-Transferase Confirms Their Identity as Fhb7 Conferring Fusarium Head Blight Resistance in Wheat.
TL;DR
GST was confirmed as the causal gene for Fhb7-mediated FHB resistance and the power of wheat × maize hybridization coupled with genome editing in validating gene function in wheat was demonstrated.
Abstract
Fhb7 is a gene derived from the homoeologous group-7 chromosomes of tall wheatgrass (Thinopyrum species), which confers resistance to Fusarium head blight (FHB) in wheat. Two resistance alleles at the Fhb7 locus (Fhb7Thp and Fhb7The2) were identified from Th. ponticum and Th. elongatum, respectively, and introgressed into the wheat genome by homoeologous recombination. Fhb7Thp was first cloned and reported to encode a glutathione S-transferase (GST), which detoxifies trichothecenes through de-epoxidation. However, a recent study has questioned the role of GST in Fhb7-mediated FHB resistance. To decipher the function of two GST-encoding alleles, Fhb7Thp and Fhb7The2, in FHB resistance, we generated haploid plants from three spring wheat lines (CS-Fhb7, WGC002, and FW23-009-80) carrying Fhb7Thp or Fhb7The2 through wide hybridizations with transgenic maize expressing Cas9 and guide RNA (gRNA) targeting GST. Of 57, 35, and 74 haploid plants derived from CS-Fhb7, WGC002, and FW23-009-80, respectively, 16, 5, and 9 had an insertion/deletion at the target sites of GST. Greenhouse evaluation of doubled haploid (DH) lines derived from the haploid plants for FHB resistance indicated that the GST-edited DH lines were significantly more susceptible and accumulated more deoxynivalenol (DON) than their corresponding wildtypes and non-edited DH lines, confirming GST as the causal gene for Fhb7-mediated FHB resistance. This study also demonstrated the power of wheat × maize hybridization coupled with genome editing in validating gene function in wheat.
