The carotenoid content of plants can be increased by overexpression of the regulatory protein ORANGE (OR) or a mutant variant known as the 'golden SNP'. In the present study, transgenic lines of the microalgae Chlamydomonas reinhardtii were generated to overexpress either wild type CrOR (CrOR WT ) or a mutated CrOR (CrOR His ) containing a single histidine substitution for a conserved arginine. Overexpression of both CrOR WT and CrOR His dramatically enhanced the accumulation of several different carotenoids, including β -cartotene, α -carotene, lutein and violaxanthin, in C. reinhardtii and, in contrast to higher plants, upregulated the transcript abundance of several relevant carotenoid biosynthetic genes. In addition, microscopic analysis revealed that the OR transgenic cells were larger than control cells and exhibited larger chloroplasts with a disrupted morphology. Moreover, both CrOR WT and CrOR His cell lines showed increased tolerance to salt and paraquat stress. The levels of endogenous phytohormone abscisic acid (ABA) were also increased in CrOR WT and CrOR His lines, not only in normal growth conditions but also in growth medium supplemented with paraquat. Together these results offer new insights regarding the role of the OR protein in regulating carotenoid biosynthesis and accumulation in microalgae, and establish a new functional role for OR to modulate oxidative stress tolerance mediated by ABA. 1 0 2 8 Berman J, Zorrilla-López U, Medina V, Farré G, Sandmann G, Capell T, Christou P, Zhu C (2017) The Arabidopsis ORANGE (AtOR) gene promotes carotenoid accumulation in transgenic corn hybrids derived from parental lines with limited carotenoid pools. Plant Cell Reports 36: 933-945. Bowler C, Slooten L, Vandenbranden S, De Rycke R, Botterman J, Sybesma C, Van Montagu M, Inzé D (1991) Manganese superoxide dismutase can reduce cellular damage mediated by oxygen radicals in transgenic plants. The EMBO Journal
