Cytochrome P450 enzyme CYP79D16 from Prunus sibirica seeds presents a novel molecular regulatory target to bioengineering oil accumulation with less amygdalin

Abstract

The seeds of Siberian apricot (Prunus sibirica L.) have abundant oils, but also contain amygdalin causing toxicity issue. This work focused on determining critical cytochrome P450 (CYP) enzyme and revealing its function in controlling amygdalin biosynthesis in P. sibirica seeds. A combination of whole-genomic identification of amygdalin synthesis-related CYPs and quantitative-comparison of transcription of CYP71/79 family members with amygdalin content in P. sibirica seeds among 18 different accessions or developmental stages was applied to identify CYP79D16 specific for seed amygdalin accumulation. The PsCYP79D16 gene was isolated, and expression and mutation were performed in yeast Saccharomyces cerevisiae, revealing high activity of PsCYP79D16 to catalyze the first step in Phe-derived amygdalin biosynthesis with ideal catalytic activity of V_max (175.44 U/mg) and K_m (0.16 mM), and functional site (Asn⁵⁰⁰). An integration of overexpression, mutation and its recovery was performed in Arabidopsis. PsCYP79D16 overexpression increased the amounts of amygdalin biosynthetic precursors and transcriptional levels of amygdalin metabolism-associated enzymes, but repressed oil accumulation and regulatory enzyme transcription (involving carbon partitioning, FA biosynthesis and triacylglycerol assembly), all of which exhibited an opposite status in cyp79d16 mutant that could be compensated by mutation restoration, unraveling a significance of PsCYP79D16 for governing seed oil and amygdalin synthesis. PsCYP79D16 should be as novel regulatory target to future bioengineering oil accumulation with less-amount amygdalin of oilseed plants.