Transcriptional regulation in apple skin cell layers during fruit development and ripening

Abstract

The fruit skin is a key component in many aspects of fruit development and quality. It is the first line of defence against pathogens, controls water loss, contains pigments and phytochemicals, and controls fruit growth through its ability to grow and stretch. Malus x domestica (apple) skin has two distinct cell layers, the epidermis and hypodermis, which cover the cortex (flesh). These distinct cell types are specialised and are known to regulate the different biomechanical properties of the skin during development. Because of the complex nature of the skin and the difficulty in studying individual cell layers, there is a poor understanding of how the skin changes during development and ripening. With accessibility to laser capture microdissection technology and the increased sensitivity in molecular analysis it is now possible to investigate this in more detail. During fruit ripening the skin becomes more porous to water, which contributes to the softening process. Spatiotemporal gene regulation of cell wall-related enzymes linked to increased porosity seen in fruit during ripening was investigated. A laser capture microdissection methodology was developed for isolating the two specific cell types of the skin (hypodermis and epidermis), along with the underlying cortex cells. This methodology was optimized to retain mRNA quality in each specific cell type. The value of cell-type dissection as a transcript-enrichment strategy was confirmed with the enrichment in epidermal and hypodermal transcripts associated with cuticle biosynthesis and flavour volatiles. Gene expression analysis of these tissues has uncovered many specialized genes and expression patterns within key biochemical pathways in fruit development. Active cell wall metabolism was also deduced exclusively in the epidermis tissues based on the expression of some genes involved in cell wall polysaccharide synthesis and modification. Novel functions of the hypodermis tissue layer were uncovered, with expression patterns that suggest a significant role in cell wall loosening and anthocyanin/flavonoid production. Uncovering a detailed map of tissue-specific gene regulation involved in fruit expansion and quality after harvest can enhance our understanding of the mechanisms that govern fruit quality. It is anticipated that the discrete spatial and temporal patterns of gene expression seen in apple can contribute to future breeding programmes and molecular discovery programmes aimed at improving skin quality for an economically important crop.