A mutation in class III homeodomain-leucine zipper (HD-ZIP III) transcription factor results in curly leaf (cul) in cucumber (Cucumis sativus L.).

KEY MESSAGE: We identified two curly-leaf (cul) mutants in cucumber. Map-based cloning revealed that both mutants are due to allelic mutations in the CsPHB gene, a homolog of the Arabidopsis PHABULOSA which encodes a class III homeodomain-leucine zipper (HD-ZIP III) transcription factor. Leaf rolling is an important agronomic trait in crop breeding. Moderate leaf rolling minimizes shadowing between leaves, leading to improved photosynthetic efficiency. Although a number of genes controlling rolled leaf have been identified from rice and other plant species, none have been mapped or cloned in cucurbit crops. In this study, we identified and characterized two curly leaf (cul) mutants, cul-1 and cul-2 in cucumber. With map-based cloning, we show that cul-1 and cul-2 are allelic mutations and CsPHB (Csa6G525430) was the candidate gene for both mutants. The CsPHB gene encoded a class III homeodomain-leucine zipper (HD-ZIP III) transcription factor. A single non-synonymous mutation in the fourth and fifth exons of the CsPHB was responsible for the cul-1 and cul-2 mutant phenotypes, respectively. The single-nucleotide substitutions in cul-1 and cul-2 were both located in cs-miRNA165/166 complementary sites of CsPHB. The expression level of CsPHB gene in multiple organs of cul-1 and cul-2 mutants was higher than that in the wild type, while the expression of cs-miRNA165/166 in the two genotypes showed the opposite trend. We speculate that disruption of the binding between the mutant allele of CsPHB and cs-miRNA165/166 leads to the curly-leaf phenotype. This is the first report to clone and characterize the CsPHB gene in the family Cucurbitaceae. Taken together, these results support CsPHB as an important player in the modulation of leaf shape development in cucumber.