CRISPR/Cas9 genome-editing applied to MdPGT1 in apple results in reduced foliar phloridzin without impacting plant growth.

Phloridzin is the most abundant polyphenolic compound in apple (Malus x domestica Borkh.), which results from the action of a key phloretin-specific UDP-2'-O-glucosyltransferase (MdPGT1). Here, we simultaneously assessed the effects of targeting MdPGT1 by conventional transgenesis and CRISPR/Cas9-mediated genome editing. To this end, we conducted transcriptomic and metabolic analyses of MdPGT1 RNAi knockdown and genome-edited lines. Knockdown lines exhibited characteristic impairment of plant growth and leaf morphology, whilst genome-edited lines exhibited normal growth despite reduced foliar phloridzin. RNA-Seq analysis identified a common core of regulated genes, involved in phenylpropanoid and flavonoid pathways. However, we identified genes and processes differentially modulated in stunted and genome-edited lines, including key transcription factors and genes involved in phytohormone signalling. Therefore, we conducted a phytohormone profiling to get insight into their role in the phenotypes observed. We found that salicylic and jasmonic acid were increased in dwarf lines, whilst auxin and abscisic acid showed no correlation with the growth phenotype. Furthermore, bioactive brassinosteroids (BRs) were commonly upregulated, whereas gibberellin GA4 was distinctively altered, showing a sharp decrease in RNAi knockdown lines. qRT-PCR expression analysis further confirmed transcriptional regulation of key factors involved in BR and GA interaction. These findings suggest that a differential modulation of phytohormones may be involved in the contrasting effects on growth following phloridzin reduction. This study also illustrates how CRISPR/Cas9 genome editing can be applied to dissect the contribution of genes involved in phloridzin biosynthesis in apple.