A single amino acid change at position 96 (Arg to His) of the sweetpotato Orange protein leads to carotenoid overaccumulation

Kim, So-Eun; Kim, Ho Soo; Ke, Qingbo; Lee, Chan-Ju; Park, Sul-U; Lim, Ye-Hoon; Park, Woo Sung; Ahn, Mi‐Jeong; Kwak, Sang‐Soo

doi:10.1007/s00299-019-02448-4

Cited by 27 publications

(19 citation statements)

References 46 publications

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“…Recently, it was found that the natural variation of the OR gene with a "golden SNP" defines melon fruit flesh color in a broad germplasm collection and governs b-carotene accumulation in melon fruit (Tzuri et al, 2015;Chayut et al, 2017). The "golden SNP" was demonstrated to alter the ability of OR for high levels of carotenoid accumulation (Yuan et al, 2015a;Kim et al, 2019). A recent study reveals that OR with a nonsynonymous mutation is also associated with carotenoid presence in carrot roots (Ellison et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

“…A recent study reveals that OR with a nonsynonymous mutation is also associated with carotenoid presence in carrot roots (Ellison et al, 2018). While ectopic expression of a wild-type OR gene increases carotenoid level (Bai et al, 2016;Park et al, 2016;Berman et al, 2017), likely due to its posttranslational upregulation of PSY protein level and activity, expression of either cauliflower OR mutant allele or an OR variant mimicking the "golden SNP" present in melon greatly promotes carotenoid accumulation in a number of plant species (Lopez et al, 2008;Yuan et al, 2015a;Kim et al, 2019;Yazdani et al, 2019). Interestingly, OR was recently found to regulate chloroplast biogenesis (Sun et al, 2019) and the expression of wild-type OR in sweetpotato and Arabidopsis lines enhances plant resistance to heat and oxidative stress treatments (Park et al, 2016;Kang et al, 2017;Kim et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

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Characterization of Cauliflower OR Mutant Variants

et al. 2020

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Cauliflower Orange (Or) mutant is characterized by high level of b-carotene in its curd. Or mutation affects the OR protein that was shown to be involved in the posttranslational control of phytoene synthase (PSY), a major rate-limiting enzyme of carotenoid biosynthesis, and in maintaining PSY proteostasis with the plastid Clp protease system. A transposon integration into the cauliflower wild-type Or gene (BoOR-wt) results in the formation of three differently spliced transcripts. One of them is characterized by insertion (BoOR-Ins), while the other two have exon-skipping deletions (BoOR-Del and BoOR-LD). We investigated the properties of individual BoOR variants and examined their effects on carotenoid accumulation. Using the yeast split-ubiquitin system, we showed that all variants were able to form OR dimers except BoOR-LD. The deletion in BoOR-LD eliminated the first of two adjacent transmembrane domains and was predicted to result in a misplacement of the C-terminal zinc finger domain to the opposite side of membrane, thus preventing OR dimerization. As interaction with PSY is mediated by the N-terminus of BoOR, which remains unaffected after splicing, all BoOR variants including BoOR-LD maintained interactions with PSY. Expression of individual BoOR mutant variants in Arabidopsis revealed that their protein stability varied greatly. While expression of BoOR-Del and BoOR-Ins resulted in increased BoOR protein levels as BoOR-wt, minimal amounts of BoOR-LD protein accumulated. Carotenoid accumulation showed correlated changes in calli of Arabidopsis expressing these variants. Furthermore, we found that OR also functions in E. coli to increase the proportion of native, enzymatically active PSY from plants upon co-expression, but not of bacterial phytoene synthase CrtB. Taken together, these results suggest that OR dimerization is required for OR stability in planta and that the simultaneous presence of PSY interaction-domains in both OR and PSY proteins is required for the holdase function of OR. The more pronounced effect of simultaneous expression of all BoOR variants in cauliflower Or mutant compared with individual overexpression on carotenoid accumulation suggests an enhanced activity with possible formation of various BoOR heterodimers.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Characterization of Cauliflower OR Mutant Variants

et al. 2020

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“…Both mutations induced the formation of chromoplasts containing carotenoid sequestering structures 7,11,28 . These mutated versions of Or have been shown to boost carotenoid accumulation in different plant species 11,[29][30][31] . However, the Or gene mechanism of action in the regulation of carotenoid accumulation is not clear yet 9,12 .…”

Section: Discussionmentioning

confidence: 99%

Differential interaction of Or proteins with the PSY enzymes in saffron

Ahrazem

López

Argandoña

et al. 2020

Sci Rep

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Colored apocarotenoids accumulate at high concentrations in few plant species, where display a role in attraction of pollinators and seed dispersers. Among these apocarotenoids, crocins accumulate at high concentrations in the stigma of saffron and are responsible for the organoleptic and medicinal properties of this spice. Phytoene synthase and Orange protein are key for carotenoid biosynthesis and accumulation. We previously isolated four phytoene synthase genes from saffron with differential roles in carotenoid and apocarotenoid biosynthesis. However, the implications of Orange genes in the regulation of apocarotenoid accumulation are unknown. Here, we have identified two Orange genes from saffron, with different expression patterns. CsOr-a was mainly expressed in vegetative tissues and was induced by light and repressed by heat stress. Both CsOr-a and CsOr-b were expressed in stigmas but showed a different profile during the development of this tissue. The interactions of CsOr-a and CsOr-b were tested with all the four phytoene synthase proteins from saffron and with CsCCD2. None interactions were detected with CCD2 neither with the phytoene synthase 2, involved in apocarotenoid biosynthesis in saffron. The obtained results provide evidence of different mechanisms regulating the phytoene synthase enzymes in saffron by Orange for carotenoid and apocarotenoid accumulation in saffron.Nucleic acid purification and cDNA isolation. Total RNA was isolated from red stigmas 48 , following the manufacturer's protocols (Qiagen, Hilden, Germany). First-strand cDNAs were synthesized by RT from 2 µg of total RNA using an 18-base pair oligo dT primer and a first-strand cDNA synthesis kit (GE Healthcare Life Sciences, www.gelifesciences.com) according to manufacturer's instructions. These cDNAs were used as templates for PCR using specific primers for CsOr-a and CsOr-b (Suppl . Table 1). Thermal cycling parameters were 2 min at 95 °C, 35 cycles of 20 s at 95 °C, 20 s at 60 °C and 1 min 30 s at 72 °C, followed by a final extension of 5 min at 72 °C. The PCR products were separated in a 1% agarose gel, purified, ligated into pGEM-T (Promega, www. promega.com), and then introduced into E. coli. Scientific RepoRtS |(2020) 10:552 | https://doi.

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“…As the only known molecular switch for chromoplast biogenesis, OR His and some other OR variants have been shown to dramatically enhance carotenoid accumulation in various plants (Lopez et al, 2008;Yuan et al, 2015;Endo et al, 2019;Kim et al, 2019;Yazdani et al, 2019). However, the restricted chromoplast number conferred by OR His could limit the capacity for further carotenoid accumulation.…”

Section: Plastid Division Factors Affect Carotenoid Contentmentioning

confidence: 99%

ORHis, a Natural Variant of OR, Specifically Interacts with Plastid Division Factor ARC3 to Regulate Chromoplast Number and Carotenoid Accumulation

et al. 2020

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Chromoplasts are colored plastids that synthesize and store massive amounts of carotenoids. Chromoplast number and size define the sink strength for carotenoid accumulation in plants. However, nothing is known about the mechanisms controlling chromoplast number. Previously, a natural allele of Orange (OR), OR His , was found to promote carotenoid accumulation by activating chromoplast differentiation and increasing carotenoid biosynthesis, but cells in orange tissues in melon fruit and cauliflower OR mutant have only one or two enlarged chromoplasts. In this study, we investigated an OR His variant of Arabidopsis OR, genetically mimicking the melon OR His allele, and found that it also constrains chromoplast number in Arabidopsis calli. Both in vitro and in vivo experiments demonstrate that OR His specifically interacts with the Membrane Occupation and Recognition Nexus domain of ACCUMULATION AND REPLICATION OF CHLOROPLASTS 3 (ARC3), a crucial regulator of chloroplast division. We further showed that OR His interferes with the interaction between ARC3 and PARALOG OF ARC6 (PARC6), another key regulator of chloroplast division, suggesting a role of OR His in competing with PARC6 for binding to ARC3 to restrict chromoplast number. Overexpression or knockout of ARC3 in Arabidopsis OR His plants significantly alters total carotenoid levels. Moreover, overexpression of the plastid division factor PLASTID DIVISION 1 greatly enhances carotenoid accumulation. These division factors likely alter carotenoid levels via their influence on chromoplast number and/or size. Taken together, our findings provide novel mechanistic insights into the machinery controlling chromoplast number and highlight a potential new strategy for enhancing carotenoid accumulation and nutritional value in food crops.

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A single amino acid change at position 96 (Arg to His) of the sweetpotato Orange protein leads to carotenoid overaccumulation

Cited by 27 publications

References 46 publications

Characterization of Cauliflower OR Mutant Variants

Characterization of Cauliflower OR Mutant Variants

Differential interaction of Or proteins with the PSY enzymes in saffron

ORHis, a Natural Variant of OR, Specifically Interacts with Plastid Division Factor ARC3 to Regulate Chromoplast Number and Carotenoid Accumulation

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