A ‘golden’ SNP in CmOr governs the fruit flesh color of melon (Cucumis melo)

Abstract

Equal contribution. Accession numbers The sequences of melon CmOr (orange flesh haplotype and green or white flesh haplotype) were deposited in GenBank. Accession numbers are KM505046 and KM505047, respectively. SUMMARYThe flesh color of Cucumis melo (melon) is genetically determined, and can be white, light green or orange, with b-carotene being the predominant pigment. We associated carotenoid accumulation in melon fruit flesh with polymorphism within CmOr, a homolog of the cauliflower BoOr gene, and identified CmOr as the previously described gf locus in melon. CmOr was found to co-segregate with fruit flesh color, and presented two haplotypes (alleles) in a broad germplasm collection, one being associated with orange flesh and the second being associated with either white or green flesh. Allelic variation of CmOr does not affect its transcription or protein level. The variation also does not affect its plastid subcellular localization. Among the identified single nucleotide polymorphisms (SNPs) between CmOr alleles in orange versus green/whiteflesh fruit, a single SNP causes a change of an evolutionarily highly conserved arginine to histidine in the CmOr protein. Functional analysis of CmOr haplotypes in an Arabidopsis callus system confirmed the ability of the CmOr orange haplotype to induce b-carotene accumulation. Site-directed mutagenesis of the CmOr green/white haplotype to change the CmOR arginine to histidine triggered b-carotene accumulation. The identification of the 'golden' SNP in CmOr, which is responsible for the non-orange and orange melon fruit phenotypes, provides new tools for studying the Or mechanism of action, and suggests genome editing of the Or gene for nutritional biofortification of crops.

Distinct Mechanisms of the ORANGE Protein in Controlling Carotenoid Flux

“…The pleiotropic effects of BoOr and the multiple alternatively sliced transcripts of the naturally occurring BoOr active allele make it difficult to resolve the Or mechanism of action. We recently reported that b-carotene accumulation in melon fruit flesh is governed by a single SNP in CmOr, providing a precise biological tool with which to study OR function (Tzuri et al, 2015) and to utilize it in biotechnology (Yuan et al, 2015a). The identification of the novel low-b mutant provides an invaluable tool for further deciphering the Or gene mechanism of action in the regulation of carotenoid accumulation.…”

“…However, the golden SNP, which governs fruit flesh b-carotene levels across a wide spectrum of melon germplasm (Tzuri et al, 2015), was not associated with a reduction in PSY protein level. Green and orange 30-DAA melon fruits showed similar levels of PSY protein independently of the golden SNP genotype and accumulated a similar amount of phytoene 24 h after NF inhibition (Fig.…”

“…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).…”

“…Based on the phenotypic similarity with the recessive homozygous Cmor-Arg melon fruits (Tzuri et al, 2015), we hypothesized that low-b was mutated either in the CmOr gene or alternatively in a structural gene of the carotenoid metabolic pathway. To differentiate between these two options, we conducted a genetic complementation test.…”

“…Fruit flesh (mesocarp) color is an important commercial trait and can be categorized into three distinctive phenotypes: white, green, and orange . The orange phenotype is a result of massive b-carotene accumulation in the fruit mesocarp governed by the golden single-nucleotide polymorphism (SNP) of the Orange (CmOr) gene (Tzuri et al, 2015). Melon plants carrying a dominant CmOr allele bear orange-fleshed fruits, while homozygous recessive Cmor plants bear white-or green-fleshed fruits.…”