Reducing PSY activity fine tunes threshold levels of a cis-carotene-derived signal that regulates the PIF3/HY5 module and plastid biogenesis

PHYTOENE SYNTHASE (PSY) is a rate-limiting enzyme catalysing the first committed step of carotenoid biosynthesis, and changes in PSY gene expression and/or protein activity alter carotenoid composition and plastid differentiation in plants. Here we identified four genetic variants of PSY (psy−4, psy−90, psy−130 and psy−145) using a forward genetics approach that rescued leaf virescence phenotypes displayed by the Arabidopsis CAROTENOID ISOMERASE (CRTISO) mutant ccr2 (carotenoid and chloroplast regulation 2) when grown under a shorter photoperiod. The four non-lethal mutations affected alternative splicing, enzyme-substrate interactions, and PSY:ORANGE multi-enzyme complex binding, constituting the dynamic posttranscriptional fine-tuning of PSY levels and activity without changing localization to the stroma and protothylakoid membranes. psy genetic variants did not alter overall total xanthophyll or cis-carotene accumulation in ccr2 yet reduced specific acyclic linear cis-carotenes linked to the biosynthesis of a yet-to-be-identified apocarotenoid signal. ccr2 psy variants modulated the ratio of PHYTOCHROME-INTERACTING FACTOR 3/ELONGATED HYPOCOTYL 5 (PIF3/HY5), displayed a normal PLB formation in etioplasts, and chlorophyll accumulation during seedling photomorphogenesis. Thus, suppressing PSY activity and impairing PSY:ORANGE protein interactions reveals how threshold specific cis-carotene levels can be fine-tuned through holoenzyme-metabolon interactions to control plastid development. ### Competing Interest Statement The authors have declared no competing interest.