Arabidopsis OR proteins are the major posttranscriptional regulators of phytoene synthase in controlling carotenoid biosynthesis

Abstract

Carotenoids are indispensable natural pigments to plants and humans. Phytoene synthase (PSY), the rate-limiting enzyme in the carotenoid biosynthetic pathway, and ORANGE (OR), a regulator of chromoplast differentiation and enhancer of carotenoid biosynthesis, represent two key proteins that control carotenoid biosynthesis and accumulation in plants. However, little is known about the mechanisms underlying their posttranscriptional regulation. Here we report that PSY and OR family proteins [Arabidopsis thaliana OR (AtOR) and AtOR-like] physically interacted with each other in plastids. We found that alteration of OR expression in Arabidopsis exerted minimal effect on PSY transcript abundance. However, overexpression of AtOR significantly increased the amount of enzymatically active PSY, whereas an ator ator-like double mutant exhibited a dramatically reduced PSY level. The results indicate that the OR proteins serve as the major posttranscriptional regulators of PSY. The ator or ator-like single mutant had little effect on PSY protein levels, which involves a compensatory mechanism and suggests partial functional redundancy. In addition, modification of PSY expression resulted in altered AtOR protein levels, corroborating a mutual regulation of PSY and OR. Carotenoid content showed a correlated change with OR-mediated PSY level, demonstrating the function of OR in controlling carotenoid biosynthesis by regulating PSY. Our findings reveal a novel mechanism by which carotenoid biosynthesis is controlled via posttranscriptional regulation of PSY in plants.arotenoids are a group of C40 isoprenoids synthesized in chloroplasts, chromoplasts, and other plastids in plants. Carotenoids serve as components of photosynthetic machinery, precursors for phytohormones, and important contributors to fruit nutritional quality and flower color (1, 2). The carotenoid biosynthetic pathway in higher plants has been well defined. However, identification of the regulatory mechanisms underlying carotenoid biosynthesis remains a challenge.Phytoene synthase (PSY) catalyzes the first committed step in carotenoid biosynthesis and controls carbon flux into the carotenoid biosynthetic pathway (1-5). Alteration of PSY expression exerts profound effects on carotenoid content (6-11). A number of factors are known to affect PSY gene expression (12)(13)(14)(15)(16)(17)(18). PSY is found to be repressed by phytochrome-interacting factors in etiolated Arabidopsis seedlings (16). PSY1 expression in tomato fruits is reported to be regulated by cis-carotenoids (14) and requires the MADS-Box transcription factor RIPENING INHIBITOR (18). Recently, it was discovered that PSY protein levels in carrot roots are modulated by a negative feedback emerging from carotenoids (19). The crucial role of PSY in carotenogenesis and the multiple factors affecting its expression suggest a complex regulatory system involved in controlling PSY. However, the factors involved in posttranscriptional regulation of PSY within plastids remain a mystery. No proteins have been re...

Elevated vitamin E content improves all- trans β-carotene accumulation and stability in biofortified sorghum

“…If PSY1 gene expression and protein accumulation can be kept consistently at relatively higher levels throughout the later stages of seed development (e.g., by the use of promoters that achieve higher PSY1 expression at all seed developmental stages, monocot codon optimization, or the selection of more stable and efficient versions of PSY), biofortified sorghum grain will accumulate more β-carotene in mature seeds. These methods combined with the recent discovery that carotenoid biosynthesis is controlled via posttranscriptional regulation of PSY by ORANGE could further improve β-carotene biosynthesis in sorghum grain (46).…”

Distinct Mechanisms of the ORANGE Protein in Controlling Carotenoid Flux

“…Recent studies of OR proteins of melon Tzuri et al, 2015), Arabidopsis (Yuan et al, 2015a;Zhou et al, 2015), and sweet potato (Park et al, 2015) and previous works with cauliflower (Li et al, 2006(Li et al, , 2012 and potato (Solanum tuberosum; Lopez et al, 2008;Li et al, 2012) suggest two complementary roles of OR in carotenoid accumulation: posttranscriptional regulation of PSY protein and biogenesis of chromoplasts. Stabilization of PSY protein in controlling carotenoid pathway flux has been shown in Arabidopsis and transgenic potato (Lopez et al, 2008).…”

Carotenogenesis Is Regulated by 5′UTR-Mediated Translation of Phytoene Synthase Splice Variants

“…Bypassing transcriptional and translational processes, the membrane-association of soluble inactive PSY enzyme leads to rapid enzymatic activation (Welsch et al, 2000). Moreover, PSY protein levels were recently found to be posttranslationally regulated by the ORANGE proteins in Arabidopsis (Zhou et al, 2015). Additionally, potential feedbackregulatory mechanisms involve translation, plastid import, protein-protein interaction, and protein turnover to achieve stoichiometrically defined carotenoid biosynthetic complexes (Cazzonelli and Pogson, 2010;Nogueira et al, 2013;Arango et al, 2014;Avendaño-Vázquez et al, 2014;Tian, 2015).…”