Improving End-Use Quality Through Introducing Cysteines Into the Central Repeat Domain of High-Molecular-Weight...

In bread wheat (Triticum aestivum L.), cysteine (Cys) residues within the N- and C-termini of high-molecular-weight glutenin subunits (HMW-GSs) critically influence dough quality. However, the functional significance of Cys residue in their central repeat domain (CRD) remains unclear. Using site-directed mutagenesis (SDM), we introduced Cys residues near the N-terminus (m1) and/or C-terminus (m2) of 1Dx2 CRD, generating variants 1Dx2m1, 1Dx2m2 and 1Dx2m1/2. Transgenic lines expressing the variants exhibited superior dough properties, increased loaf volume, and elevated glutenin macropolymers (GMPs) content, attributable to enhanced disulfide bond formation and upregulation of associated genes. Notably, the two Cys residues introduced variant 1Dx2m1/2 demonstrated additive improvements, indicating synergistical effects of Cys residues at both positions. Field trials confirmed these modifications did not compromise key agronomic traits. Our study provides the experimental evidence for the role of CRD-located Cys residues in HMW-GSs on dough quality and offers valuable genetic resources for improving end-use quality without yield penalties in wheat breeding.