Chalcone Synthase mRNA and Activity Are Reduced in Yellow Soybean Seed Coats with Dominant I Alleles

Abstract

The seed of all wild Clycine accessions have black or brown pigments because of the homozygous recessive i allele in combination with alleles at the R and T loci. In contrast, nearly all commercial soybean (GIycine max) varieties are yellow due to the presence of a dominant allele of the I locus (either I or rl) that inhibits pigmentation in the seed coats. Spontaneous mutations to the recessive i allele occur in these varieties and result in pigmented seed coats. We have isolated a clone for a soybean dihydroflavonol reductase (DFR) gene using polymerase chain reaction. We examined expression of DFR and two other genes of the flavonoid pathway during soybean seed coat development in a series of nearisogenic isolines that vary in pigmentation as specified by combinations of alleles of the I , R, and T loci. The expression of phenylalanine ammonia-lyase and DFR mRNAs was similar in all of the gene combinations at each stage of seed coat development. In contrast, chalcone synthase (CHS) mRNA was barely detectable at all stages of development in seed coats that carry the dominant I allele that results in yellow seed coats. CHS activity in yellow seed coats (I) was also 7-to 10-fold less than in the pigmented seed coats that have the homozygous recessive i allele. It appears that the dominant I allele results in reduction of CHS mRNA, leading to reduction of CHS activity as the basis for inhibition of anthocyanin and proanthocyanin synthesis in soybean seed coats. A further connection between CHS and the I locus is indicated by the occurrence of multiple restriction site polymorphisms in genomic DNA blots of the CHS gene family in near-isogenic lines containing alleles of the I locus.Pigmentation of soybean (Glycine max) seed coats is influenced by at least three genes, I, R, and T. The I gene (for inhibitor) controls synthesis and spatial occurrence of pigments in the epidermal layer of the seed coat, whereas R and T are responsible for the specific color (reviewed in Bemard and Weiss, 1973;Palmer and Kilen, 1987). The dominant I allele inhibits pigment accumulation, resulting in a yellow seed coat color at maturity, whereas the homozygous, recessive i allele specifies full pigmentation across the entire seedWe are grateful for a graduate fellowship award from the. Na- Recently, we showed that polymeric proanthocyanidins are also synthesized in pigmented seed coats with the homozygous recessive i genotype (Todd and Vodkin, 1993). Proanthocyanidins are not precursors of anthocyanins but are synthesized as polymeric condensation products of leucoanthocyanins, the precursors of anthocyanins. Brown (irT) and buff (irt) seed coats do not contain anthocyanins but they do synthesize proanthocyanidins. The mechanism of inhibition of anthocyanin and proanthocyanidin synthesis by the I allele is unknown.Spontaneous mutations of the dominant I alleles ( I or 2%) to the recessive i allele have occurred in many soybean varieties, resulting in isogenic lines with pigmented seed coats. A Prorich cell wall protein, PRP1, and its ...

Duplications That Suppress and Deletions That Restore Expression from a Chalcone Synthase Multigene Family.

“…ther the i' allele (pigmented hilum and yellow seed coat) or the I allele (yellow seed coat) at all stages of seed development (Wang et al, 1994). Moreover, CHS enzyme activity levels are reduced sevenfold to 10-fold in seed coats that are homozygous for the I allele (yellow seed coat) or the ii alleie (pigmented hilum and yellow seed coat) when compared with enzyme activity levels in homozygous recessive (ili) genotypes.…”

“…Moreover, CHS enzyme activity levels are reduced sevenfold to 10-fold in seed coats that are homozygous for the I allele (yellow seed coat) or the ii alleie (pigmented hilum and yellow seed coat) when compared with enzyme activity levels in homozygous recessive (ili) genotypes. Multiple restriction enzyme polymorphisms also have been found in genomic DNA blots with a CHS probe (Wang et al, 1994).…”

“…The I locus affects only seed coat pigmentation and the deposition of proanthocyanidins in the seed coat (Xodd and Vodkin, 1993;Wang et al, 1994). It does not phenotypically affect flower pigmentation, trichome color, or hypocotyl color.…”

“…Total RNA was extracted from seed coats from the designated weight class of seed by using phenol-chloroform and lithium chloride precipitation (McCarty, 1986;Wang et al, 1994). RNAwas then stored at -70°C until use.…”

Tissue-Specific Gene Silencing Mediated by a Naturally Occurring Chalcone Synthase Gene Cluster inGlycine max [W]

“…The Dominant I Allele Inhibits CHS Expression Only in the Seed Coats Previous work had shown that the I locus, corresponding to a region of chalcone synthase gene duplications, in its dominant form prevents the accumulation of anthocyanins in the G. max seed coat (Wang et al, 1994;Todd and Vodkin, 1996). Evidently, chalcone synthase, a key regulatory enzyme in the flavonoid pathway, is not only involved in the production of pigment in the seed coat, but plays a significant role in the synthesis of fundamental secondary metabolites functioning as UV protectants, phytoalexins, insect protectants, and symbiosis initiators in various plant tissues.…”