RESEARCH ARTICLES

Caught Red-Handed:Rc Encodes a Basic Helix-Loop-Helix Protein Conditioning Red Pericarp in Rice W OA

Megan T.Sweeney,a Michael J.Thomson,a,1Bernard E.Pfeil,b and Susan McCouch a,2

a Department of Plant Breeding and Genetics,Cornell University,Ithaca,New York14953-1901

b Department of Plant Biology,Cornell University,Ithaca,New York14853

Rc is a domestication-related gene required for red pericarp in rice(Oryza sativa).The red grain color is ubiquitous among the wild ancestors of O.sativa,in which it is closely associated with seed shattering and dormancy.Rc encodes a basic helix-loop-helix(bHLH)protein that wasfine-mapped to an18.5-kb region on rice chromosome7using a cross between Oryza rufipogon(red pericarp)and O.sativa cv Jefferson(white pericarp).Sequencing of the alleles from both mapping parents as well as from two independent genetic stocks of Rc revealed that the dominant red allele differed from the recessive white allele by a14-bp deletion within exon6that knocked out the bHLH domain of the protein.A premature stop codon was identified in the second mutant stock that had a light red pericarp.RT-PCR experiments confirmed that the Rc gene was expressed in both red-and white-grained rice but that a shortened transcript was present in white varieties. Phylogenetic analysis,supported by comparative mapping in rice and maize(Zea mays),showed that Rc,a positive regulator of proanthocyanidin,is orthologous with INTENSIFIER1,a negative regulator of anthocyanin production in maize, and is not in the same clade as rice bHLH anthocyanin regulators.

INTRODUCTION

Most rice(Oryza sativa)that is grown and consumed throughout the world has white pericarp,but rice can also produce grains with brown,red,and purple pericarp.The color is visible when the grains are dehulled,but it can be removed by polishing to reveal the white endosperm.Red pericarp is ubiquitous among the wild ancestors of cultivated rice(Oryza rufipogon),and in some regions of the world red cultivars are preferred for their taste,texture,and ceremonial or medicinal value.Consumer interest in red and purple rices represents a growing specialty market in the United States,but at the same time,the constant presence of weedy red rice in farmers’fields is the most eco-nomically important pest and grain-quality problem faced by U.S.rice growers(Gealy et al.,2002).Red rices,which typically show seed shattering and dormancy along with a red pericarp, can belong to either O.sativa or O.rufipogon(Vaughan et al., 2001),neither of which is native to the United States.They interbreed freely with cultivated,white-grained types,making transgenic herbicide-resistant varieties impractical.

The red pigment in rice grains is proanthocyanidin,also called condensed tannins(Oki et al.,2002).Proanthocyanidins are a branch off the anthocyanin pathway and share many of the same biosynthetic genes(Winkel-Shirley,2001).Proanthocyanidins have been shown to have important deterrent effects on path-ogens and predators,so it is not surprising that spontaneous mutations that inhibit pigment production would be selected against in the wild(Shirley,1998).On the other hand,white grain appears to be associated with the domestication syndrome and remains under strong selection in most rice breeding programs today.

Regardless of the problems associated with red rice as a weed,the red pigment is of interest for nutritional reasons.It serves as a powerful antioxidant that has been demonstrated to reduce atherosclerotic plaque formation,a risk factor associated with cardiovascular disease(Ling et al.,2001).On the negative side,proanthocyanidin pigments reduce the bioavailability of iron,protein,and carbohydrates(Eggum et al.,1981;Carmona et al.,1996;Glahn et al.,2002),which has important implications for people with low nutritional status.A better understanding of the genetics and molecular biology of red pericarp and the association of this characteristic with other wild/weedy traits will provide important information for the better management of both the negative and positive features associated with red rice. Two loci have been identified using classical genetic analysis, Rc(brown pericarp and seed coat)and Rd(red pericarp and seed coat).When present together,these loci produce red seed color (Kato and Ishikawa,1921).Rc in the absence of Rd produces

1Current address:Department of Plant Breeding,Genetics,and

Biotechnology,International Rice Research Institute,Los Ban˜os,

Laguna,Philippines.

2To whom correspondence should be addressed.E-mail srm4@

cornell.edu;fax607-255-6683.

The author responsible for distribution of materials integral to the

findings presented in this article in accordance with the policy described

in the Instructions for Authors()is:Susan R.McCouch

(srm4@cornell.edu).

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Article,publication date,and citation information can be found at

/cgi/doi/10.1105/tpc.105.038430.

The Plant Cell,Vol.18,283–294,February2006,ª2006American Society of Plant Biologists