Barley BODYGUARD controls cuticular specialisations regulated by SHINE transcription factors

Abstract

The outer epidermis of land plants secretes a cuticular layer, a hydrophobic diffusion barrier which minimises water loss into the atmosphere and protects from pests, ultraviolet light and organ fusion. Cuticles typically comprise a polyester cutin matrix embedded and overlaid with cuticular waxes, but their exact chemical make-up, structure and functions can vary widely depending on the tissue and species. Barley shows two such cuticular specialisations: (1) deposition of a thick {beta}-diketone-rich wax bloom on multiple organs at reproductive stage, common in other Poeceae species and linked to yield; and, (2) secretion of a sticky layer on the grain fruit (caryopsis) pericarp cuticle which adheres to inner floral hulls, leading to barleys distinctive covered grain used in animal feed and malting. Two SHINE/WAX-INDUCER transcription factors in barley, HvWIN1 and NUD, promote the wax bloom and hull to caryopsis adhesion, respectively, yet little is understood about other genes involved. Leveraging near-isogenic lines of wax-deficient mutants, we identify the barley BODYGUARD1 (HvBDG1) gene encoding an /{beta}-hydrolase essential for leaf cuticular integrity and wax bloom deposition. Modelling of functional and defective alleles suggests that HvBDG1 N-terminal region control of protein flexibility is important for HvBDG1 function. In addition to their role in controlling barley epicuticular wax deposition, we show that both HvBDG1 and HvWIN1 are essential for strong hull to caryopsis adhesion. Along with NUD, these gene products differentially contribute to ultrastructural changes on the pericarp associated with a cuticular building programme driven by NUD and HvWIN1 regulation of cuticle metabolism and transport and cell wall-related genes, and correlate with shifts in pericarp surface chemistry. We also show that the previously asserted grain-specific role of NUD should be revised, as our findings reveal that it is essential for maintaining leaf cuticle integrity. Our analyses in barley suggest that NUD and HvWIN1 control cuticular specialisations and cuticle integrity in part via promotion of HvBDG1 expression, while HvWIN1 and NUD likely act independently from each other. Lastly, mining tetraploid wheat mutant populations followed by crossing to combine mutated homoeologues demonstrated that BDG1 and WIN1 orthologues also control wax bloom in wheat. Taken together, our work greatly expands the genetic networks and molecular activities important for cuticle development in cereals and the underlying mechanisms for both shared and species-specific cuticular specialisations.