Regulatory Genes Controlling Anthocyanin Pigmentation Are Functionally Conserved among Plant Species and Have Distinct Sets of Target Genes (original) (raw)
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Role of the regulatory gene pl in the photocontrol of maize anthocyanin pigmentation
The Plant Cell, 1993
The pl gene encodes a regulatory protein that controls the transcription of a number of structural genes of the anthocyanin biosynthetic pathway i n maize. pl alleles have been classified phenotypically into two categories: dominant (Pr) alleles lead to intense, light-independent pigmentation in vegetative and floral organs of the plant; recessive "sun-red" alleles (pr) lead to light-dependent red pigmentation in which only tissues exposed to light become pigmented. Based on these observations, two alternate pathways leading to anthocyanin synthesis in the plant have been proposed: one requiring light and the other bypassing the light requirement through the action of Pl. To evaluate this hypothesis, we have analyzed light-independent and light-dependent alleles of pl. Sequence analysis revealed that the two types of alleles have very distinct promoters but have the capacity to encode very similar proteins. The protein encoded by one recessive allele was shown to be functional in transient assays. Measurements of husk mRNA levels by quantitative polymerase chain reaction showed that sun-red pl alleles are expressed at much lower levels than a Pl allele, but their expression is increased approximately sixfold by exposure to light. These results lead to the conclusion that the sun-red pl alleles are not null; instead, they synthesize functional mRNA and protein. We propose that the light-dependent pigmentation observed in pl plants is the result of a threshold effect in which light exposure boosts pl mRNA expression past a crucial level necessary to generate sufficient PL protein molecules to activate transcription of the anthocyanin structural genes.
Genes & Development, 1992
The B gene encodes a transcription factor of the basic helix-loop-helix class, which controls the synthesis of the anthocyanin pigments in maize. This gene, as well as the highly homologous R gene family, displays extensive allelic variation in that different alleles cause distinct distributions of anthocyanin pigments in different tissues and at different developmental times. The analysis of the expression of two B alleles, with distinct tissue-specific patterns of anthocyanin synthesis in plant and seed tissues, demonstrates that the amount of B transcripts correlates with the accumulation of anthocyanins in the various tissues. The comparison of the genomic clones for the two alleles reveals high sequence identity in the coding and 3'-flanking regions (98% and approximately 90%, respectively). In contrast, the most 5' region of their mRNAs and the 5'-flanking sequences share no significant sequence identity. This result suggests that the alleles diverged from each oth...
The Plant Cell …, 1989
Genetic studies in maize have identified several regulatory genes that control the tissue-specific synthesis of the purple anthocyanin pigments during development. Two such genes, R and 8, exhibit extensive allelic diversity with respect to the tissue specificity and developmental timing of anthocyanin synthesis. Previous genetic studies demonstrated that certain B alleles can substitute for R function, and in these cases only one functional allele at either locus is required for pigment synthesis in the aleurone. In addition, biochemical studies have shown that both genes act on the same biosynthetic pathway, suggesting that the genes are functionally duplicate. In this report we describe DNA hybridization experiments that demonstrate that the functionally duplicate nature of B and R is reflected in DNA sequence similarity between the two genes. We took advantage of this homology and used the R genomic sequences to clone B. Two different strategies were pursued and two genomic clones isolated, a 2.5kilobase Bglll fragment linked to the b allele in W23 inbred stocks and a 1.0-kilobase Hindlll fragment linked to the B allele in CM37 stocks. Examination of several independent transposable element insertion mutations in B and revertant derivatives demonstrated that our clones recognize the functional B gene. Genomic clones representing the entire B-Peru allele were isolated, and a detailed restriction map was prepared. Using these clones we have identified a 2.2-kilobase mRNA in husks from plants containing either B-I or B-Peru alleles, but no B mRNA was detected in plants containing a b allele. The transcript is at least 100 times more abundant in strongly pigmented B-I husks than in weakly pigmented B-Peru husk tissue. Expression of functional 6 alleles in husk tissue correlates with the coordinate increase in mRNA levels of two structural genes of the pathway, A7 and 627, consistent with the postulated role of B as a regulatory gene.
Molecular analysis of the maize anthocyanin regulatory locus C1
Proceedings of the National Academy of Sciences, 1986
The Cl gene of maize plays a regulatory role in the production of anthocyanin pigments in the aleurone layer of the endosperm. As an initial step toward understanding the molecular details of how Cl controls pigment biosynthesis, we cloned the Cl gene. This was accomplished by first cloning a mutable allele of Cl, cl-mS, which contains the transposable element Spm. A combination of molecular and genetic analysis was used to identify the Spm at the Cl locus. Individual genomic DNAs from a population in which the cl-mutable phenotype was segregating with the recessive cl phenotype were digested with methyl-sensitive restriction enzymes and probed with a small DNA fragment derived from a defective Spm. One Sal I restriction fragment complementary to the Spm probe was shown to be present in the DNA of individuals with the cl-m5 phenotype but absent from DNA of individuals with a recessive cl phenotype. Subsequent cloning and restriction analysis of this fragment revealed sequences flanking the Spm that proved to be Cl-specific. A DNA fragment derived from the flanking sequences was then used as a probe to clone the wild-type Cl gene and several additional alleles of Cl, including one stable recessive, two mutations caused by Ds insertions, one mutation induced by insertion of a defective Spm, and two dominant mutations, Cl-S and C1-. RNA blot hybridization analysis of three Cl alleles indicates that Cl regulation of the Bzl and Al structural genes in the anthocyanin biosynthetic pathway is at the transcriptional level.
Genetic Regulation and Photocontrol of Anthocyanin Accumulation in Maize Seedlings
THE PLANT CELL ONLINE, 1990
The flavonoid pathway leading to anthocyanin biosynthesis in maize is controlled by multiple regulatory genes and induced by various developmental and environmental factors. We have investigated the effect of the regulatory loci R, B, and f / on anthocyanin accumulation and on the expression of four genes ( C2, A l , 621, and 822) in the biosynthetic pathway during an inductive light treatment. The results show that light-mediated anthocyanin biosynthesis is regulated solely by R; the contributions of B and f / are negligible in young seedlings. lnduction of the A1 and Bz2 genes by high fluence-rate white light requires the expression of a dominant R allele, whereas accumulation of C2 and Bzl mRNA occurs with either a dominant or recessive allele at R. A1 and 822 mRNA accumulate only in response to high fluence-rate white light, but Bzl is fully expressed in dim red light. Some C2 mRNA is induced by dim red light, but accumulation is far greater in high fluence-rate white light. Furthe'more, expression from both dominant and recessive alleles of the regulatory gene R is enhanced by high fluence-rate white light. Seedlings with a recessive allele at R produce functional chalcone synthase protein (the C2 gene product) but accumulate no anthocyanins, suggesting that, in contrast to the R-mediated coordinate regulation of C2 and Bzl observed in the aleurone, C2 expression in seedlings is independent of R and appears to be regulated by a different light-sensitive pathway.
Gene regulation networks generate diverse pigmentation patterns in plants
Plant Signaling & Behavior, 2014
T he diversity of pigmentation patterns observed in plants occurs due to the spatial distribution and accumulation of colored compounds, which may also be associated with structural changes to the tissue. Anthocyanins are flavonoids that provide red/purple/blue coloration to plants, often forming complex patterns such as spots, stripes, and veinassociated pigmentation, particularly in flowers. These patterns are determined by the activity of MYB-bHLH-WDR (MBW) transcription factor complexes, which activate the anthocyanin biosynthesis genes, resulting in anthocyanin pigment accumulation. Recently, we established that the MBW complex controlling anthocyanin synthesis acts within a gene regulation network that is conserved within at least the Eudicots. This network involves hierarchy, reinforcement, and feedback mechanisms that allow for stringent and responsive regulation of the anthocyanin biosynthesis genes. The gene network and mobile nature of the WDR and R3-MYB proteins provide exciting new opportunities to explore the basis of pigmentation patterning, and to investigate the evolutionary history of the MBW components in land plants.
Identification of the Pr1 Gene Product Completes the Anthocyanin Biosynthesis Pathway of Maize
Genetics, 2011
In maize, mutations in the pr1 locus lead to the accumulation of pelargonidin (red) rather than cyanidin (purple) pigments in aleurone cells where the anthocyanin biosynthetic pathway is active. We characterized pr1 mutation and isolated a putative F39H encoding gene (Zmf39h1) and showed by segregation analysis that the red kernel phenotype is linked to this gene. Genetic mapping using SNP markers confirms its position on chromosome 5L. Furthermore, genetic complementation experiments using a CaMV 35S::ZmF39H1 promoter-gene construct established that the encoded protein product was sufficient to perform a 39hydroxylation reaction. The Zmf39h1-specific transcripts were detected in floral and vegetative tissues of Pr1 plants and were absent in pr1. Four pr1 alleles were characterized: two carry a 24 TA dinucleotide repeat insertion in the 59-upstream promoter region, a third has a 17-bp deletion near the TATA box, and a fourth contains a Ds insertion in exon1. Genetic and transcription assays demonstrated that the pr1 gene is under the regulatory control of anthocyanin transcription factors red1 and colorless1. The cloning and characterization of pr1 completes the molecular identification of all genes encoding structural enzymes of the anthocyanin pathway of maize.
THE PLANT CELL ONLINE, 2002
ANTHOCYANIN1 (AN1) of petunia is a transcription factor of the basic helix-loop-helix (bHLH) family that is required for the synthesis of anthocyanin pigments. Here, we show that AN1 controls additional aspects of cell differentiation: the acidification of vacuoles in petal cells, and the size and morphology of cells in the seed coat epidermis. We identified an1 alleles, formerly known as ph6 , that sustain anthocyanin synthesis but not vacuolar acidification and seed coat morphogenesis. These alleles express truncated proteins lacking the C-terminal half of AN1, including the bHLH domain, at an ف 30-fold higher level than wild-type AN1. An allelic series in which one, two, or three amino acids were inserted into the bHLH domain indicated that this domain is required for both anthocyanin synthesis and vacuolar acidification. These findings show that AN1 controls more aspects of epidermal cell differentiation than previously thought through partially separable domains.
Journal of Plant Physiology, 2008
In this work we analysed, at the transcript level, the response of Arabidopsis anthocyanin regulatory genes of the MYB (PAP1 and PAP2), bHLH (TT8, EGL3 and GL3) and WD40 (TTG1) families to white light in seedlings and to different light qualities in rosette leaves. Our experiments showed strong light induction of the MYB genes PAP1 and PAP2. In particular, the kinetics of PAP1 expression preceded those of PAP2 and all of the structural genes (CHS, DFR, F3H, LDOX), consistent with the hypothesis that it has a key role in light induction of anthocyanin biosynthesis. All bHLH genes analysed showed light induction, and in seedlings their expression preceded that of the late structural genes, suggesting their possible role in light regulation of these structural genes. TTG1 expression is essentially constitutive in both systems. Experiments with transgenic lines over-expressing the MYB factors show that PAP1, but not PAP2, strongly stimulates expression of the anthocyanin structural gene encoding dihydroflavonol reductase, but neither factor affected expression of the early flavonoid biosynthesis gene encoding chalcone synthase. Consistent with these findings, PAP1, but not PAP2, stimulated light induction of anthocyanin biosynthesis in seedlings. We conclude that specific members of the MYB and bHLH families play important roles in regulating anthocyanin biosynthesis in response to different light qualities in Arabidopsis.