Copy number variation of TdDof controls solid-stemmed architecture in wheat

Stem solidness is an important agronomic trait of durum (Triticum turgidumL. var.durum) and bread (Triticum aestivumL.) wheat that provides resistance to the wheat stem sawfly. This dominant trait is conferred by theSSt1locus on chromosome 3B. However, the molecular identity and mechanisms underpinning stem solidness have not been identified. Here, we demonstrate that copy number variation ofTdDof, a gene encoding a putative DNA binding with one finger protein, controls the stem solidness trait in wheat. Using map-based cloning, we localizedTdDofto within a physical interval of 2.1 Mb inside theSSt1locus. Molecular analysis revealed that hollow-stemmed wheat cultivars such as Kronos carry a single copy ofTdDof, whereas solid-stemmed cultivars such as CDC Fortitude carry multiple identical copies of the gene. Deletion of allTdDofcopies from CDC Fortitude resulted in the loss of stem solidness, whereas the transgenic overexpression ofTdDofrestored stem solidness in theTdDofdeletion mutantpithless1and conferred stem solidness in Kronos. In solid-stemmed cultivars, increasedTdDofexpression was correlated with the down-regulation of genes whose orthologs have been implicated in programmed cell death (PCD) in other species. Anatomical and histochemical analyses revealed that hollow-stemmed lines had stronger PCD-associated signals in the pith cells compared to solid-stemmed lines, which suggests copy number-dependent expression ofTdDofcould be directly or indirectly involved in the negative regulation of PCD. These findings provide opportunities to manipulate stem development in wheat and other monocots for agricultural or industrial purposes.