The HD-ZIP II Gene PaHAT14 Increases Cuticle Deposition by Downregulating ERF Gene PaERF105 in Phalaenopsis.

To analyze the genes involved in orchid floral development, a homeodomain-leucine zipper II gene PaHAT14, which is specifically and highly expressed in perianth during early flower development, was identified from Phalaenopsis. Transgenic Arabidopsis plants expressing 35S::PaHAT14 and 35S::PaHAT14 + SRDX (fused with the repressor motif SRDX) exhibited similar altered phenotypes, including small leaves, early flowering and bending petals with increased cuticle production. This suggests that PaHAT14 acts as a repressor. In contrast, transgenic Arabidopsis plants expressing 35S::PaHAT14 + VP16 (fused with the activation domain VP16) exhibited curled leaves, late flowering and folded petals with decreased cuticle production within hardly opened flowers. Additionally, the expression of the ERF gene DEWAX2, which negatively regulates cuticular wax biosynthesis, was downregulated in 35S::PaHAT14 and 35S::PaHAT14 + SRDX transgenic Arabidopsis, while it was upregulated in 35S::PaHAT14 + VP16 transgenic Arabidopsis. Furthermore, transient overexpression of PaHAT14 in Phalaenopsis petal/sepal increased cuticle deposition due to the downregulation of PaERF105, a Phalaenopsis DEWAX2 ortholog. On the other hand, transient overexpression of PaERF105 decreased cuticle deposition, whereas cuticle deposition increased and the rate of epidermal water loss was reduced in PaERF105 virus-induced gene silencing Phalaenopsis flowers. Moreover, ectopic expression of PaERF105 not only produced phenotypes similar to those in 35S::PaHAT14 + VP16 Arabidopsis but also compensated for the altered phenotypes observed in 35S::PaHAT14 and 35S::PaHAT14 + SRDX Arabidopsis. These results suggest that PaHAT14 promotes cuticle deposition by negatively regulating downstream gene PaERF105 in orchid flowers.