Paul Bailey

Transcriptional repression by AtMYB4 controls production of UV‐protecting sunscreens in Arabidopsis

An Arabidopsis thaliana line that is mutant for the R2R3 MYB gene, AtMYB4, shows enhanced levels of sinapate esters in its leaves. The mutant line is more tolerant of UV‐B irradiation than wild type. The increase in sinapate ester accumulation in the mutant is associated with an enhanced expression of the gene encoding cinnamate 4‐hydroxylase, which appears to be the principal target of AtMYB4 and an effective rate limiting step in the synthesis of sinapate ester sunscreens. AtMYB4 expression is downregulated by exposure to UV‐B light, indicating that derepression is an important mechanism for acclimation to UV‐B in A.thaliana. The response of target genes to AtMYB4 repression is dose dependent, a feature that operates under physiological conditions to reinforce the silencing effect of AtMYB4 at high activity. AtMYB4 works as a repressor of target gene expression and includes a repression domain. It belongs to a novel group of plant R2R3 MYB proteins involved in transcriptional silencing. The balance between MYB activators and repressors on common target promoters may provide extra flexibility in transcriptional control.

A small family of MYB-regulatory genes controls floral pigmentation intensity and patterning in the genus Antirrhinum

The Rosea1, Rosea2, and Venosa genes encode MYB-related transcription factors active in the flowers of Antirrhinum majus. Analysis of mutant phenotypes shows that these genes control the intensity and pattern of magenta anthocyanin pigmentation in flowers. Despite the structural similarity of these regulatory proteins, they influence the expression of target genes encoding the enzymes of anthocyanin biosynthesis with different specificities. Consequently, they are not equivalent biochemically in their activities. Different species of the genus Antirrhinum, native to Spain and Portugal, show striking differences in their patterns and intensities of floral pigmentation. Differences in anthocyanin pigmentation between at least six species are attributable to variations in the activity of the Rosea and Venosa loci. Set in the context of our understanding of the regulation of anthocyanin production in other genera, the activity of MYB-related genes is probably a primary cause of natural variation in anthocyanin pigmentation in plants.

Retrotransposons Control Fruit-Specific, Cold-Dependent Accumulation of Anthocyanins in Blood Oranges

The cold dependency of pigment formation in blood orange constitutes a major limitation on production worldwide and is due to the cold induction of retrotransposons controlling this gain-of-function trait. Traditionally, Sicilian blood oranges (Citrus sinensis) have been associated with cardiovascular health, and consumption has been shown to prevent obesity in mice fed a high-fat diet. Despite increasing consumer interest in these health-promoting attributes, production of blood oranges remains unreliable due largely to a dependency on cold for full color formation. We show that Sicilian blood orange arose by insertion of a Copia-like retrotransposon adjacent to a gene encoding Ruby, a MYB transcriptional activator of anthocyanin production. The retrotransposon controls Ruby expression, and cold dependency reflects the induction of the retroelement by stress. A blood orange of Chinese origin results from an independent insertion of a similar retrotransposon, and color formation in its fruit is also cold dependent. Our results suggest that transposition and recombination of retroelements are likely important sources of variation in Citrus.

Comparative genomics of flowering time pathways using Brachypodium distachyon as a model for the temperate grasses.

Brachypodium distachyon (Brachypodium) is a model for the temperate grasses which include important cereals such as barley, wheat and oats. Comparison of the Brachypodium genome (accession Bd21) with those of the model dicot Arabidopsis thaliana and the tropical cereal rice (Oryza sativa) provides an opportunity to compare and contrast genetic pathways controlling important traits. We analysed the homologies of genes controlling the induction of flowering using pathways curated in Arabidopsis Reactome as a starting point. Pathways include those detecting and responding to the environmental cues of day length (photoperiod) and extended periods of low temperature (vernalization). Variation in these responses has been selected during cereal domestication, providing an interesting comparison with the wild genome of Brachypodium. Brachypodium Bd21 has well conserved homologues of circadian clock, photoperiod pathway and autonomous pathway genes defined in Arabidopsis and homologues of vernalization pathway genes defined in cereals with the exception of VRN2 which was absent. Bd21 also lacked a member of the CO family (CO3). In both cases flanking genes were conserved showing that these genes are deleted in at least this accession. Segmental duplication explains the presence of two CO-like genes in temperate cereals, of which one (Hd1) is retained in rice, and explains many differences in gene family structure between grasses and Arabidopsis. The conserved fine structure of duplications shows that they largely evolved to their present structure before the divergence of the rice and Brachypodium. Of four flowering-time genes found in rice but absent in Arabidopsis, two were found in Bd21 (Id1, OsMADS51) and two were absent (Ghd7, Ehd1). Overall, results suggest that an ancient core photoperiod pathway promoting flowering via the induction of FT has been modified by the recruitment of additional lineage specific pathways that promote or repress FT expression.

AtMYB12 regulates caffeoyl quinic acid and flavonol synthesis in tomato: expression in fruit results in very high levels of both types of polyphenol

Plant polyphenolics exhibit a broad spectrum of health-promoting effects when consumed as part of the diet, and there is considerable interest in enhancing the levels of these bioactive molecules in plants used as foods. AtMYB12 was originally identified as a flavonol-specific transcriptional activator in Arabidopsis thaliana, and this has been confirmed by ectopic expression in tobacco. AtMYB12 is able to induce the expression of additional target genes in tobacco, leading to the accumulation of very high levels of flavonols. When expressed in a tissue-specific manner in tomato, AtMYB12 activates the caffeoyl quinic acid biosynthetic pathway, in addition to the flavonol biosynthetic pathway, an activity which probably mirrors that of the orthologous MYB12-like protein in tomato. As a result of its broad specificity for transcriptional activation in tomato, AtMYB12 can be used to produce fruit with extremely high levels of multiple polyphenolic anti-oxidants. Our data indicate that transcription factors may have different specificities for target genes in different plants, which is of significance when designing strategies to improve metabolite accumulation and the anti-oxidant capacity of foods.

Update on the basic helix-loop-helix transcription factor gene family in Arabidopsis thaliana

Basic helix-loop-helix (bHLH) transcription factors represent a family of proteins that contain a bHLH domain, a motif involved in binding DNA. Recently, two groups independently analyzed the BHLH gene family of Arabidopsis thaliana ([Heim et al., 2003][1]; [Toledo-Ortiz et al., 2003][2]). These

Control of cell and petal morphogenesis by R2R3 MYB transcription factors

Petals of animal-pollinated angiosperms have adapted to attract pollinators. Factors influencing pollinator attention include colour and overall size of flowers. Colour is determined by the nature of the pigments, their environment and by the morphology of the petal epidermal cells. Most angiosperms have conical epidermal cells, which enhance the colour intensity and brightness of petal surfaces. The MYB-related transcription factor MIXTA controls the development of conical epidermal cells in petals of Antirrhinum majus. Another gene encoding an R2R3 MYB factor very closely related to MIXTA, AmMYBML2, is also expressed in flowers of A. majus. We have analysed the roles of AmMYBML2 and two MIXTA-related genes, PhMYB1 from Petunia hybrida and AtMYB16 from Arabidopsis thaliana, in petal development. The structural similarity between these genes, their comparable expression patterns and the similarity of the phenotypes they induce when ectopically expressed in tobacco, suggest they share homologous functions closely related to, but distinct from, that of MIXTA. Detailed phenotypic analysis of a phmyb1 mutant confirmed the role of PhMYB1 in the control of cell morphogenesis in the petal epidermis. The phmyb1 mutant showed that epidermal cell shape affects petal presentation, a phenotypic trait also observed following re-examination of mixta mutants. This suggests that the activity of MIXTA-like genes also contributes to petal form, another important factor influencing pollinator attraction.

Transcripts of Vp-1 homeologues are misspliced in modern wheat and ancestral species

The maize (Zea mays) Viviparous 1 (Vp1) transcription factor has been shown previously to be a major regulator of seed development, simultaneously activating embryo maturation and repressing germination. Hexaploid bread wheat (Triticum aestivum) caryopses are characterized by relatively weak embryo dormancy and are susceptible to preharvest sprouting (PHS), a phenomenon that is phenotypically similar to the maize vp1 mutation. Analysis of Vp-1 transcript structure in wheat embryos during grain development showed that each homeologue produces cytoplasmic mRNAs of different sizes. The majority of transcripts are spliced incorrectly, contain insertions of intron sequences or deletions of coding region, and do not have the capacity to encode full-length proteins. Several VP-1-related lower molecular weight protein species were present in wheat embryo nuclei. Embryos of a closely related tetraploid species (Triticum turgidum) and ancestral diploids also contained misspliced Vp-1 transcripts that were structurally similar or identical to those found in modern hexaploid wheat, which suggests that compromised structure and expression of Vp-1 transcripts in modern wheat are inherited from ancestral species. Developing embryos from transgenic wheat grains expressing the Avena fatua Vp1 gene showed enhanced responsiveness to applied abscisic acid compared with the control. In addition, ripening ears of transgenic plants were less susceptible to PHS. Our results suggest that missplicing of wheat Vp-1 genes contributes to susceptibility to PHS in modern hexaploid wheat varieties and identifies a possible route to increase resistance to this environmentally triggered disorder.

Genetic map locations for orthologous Vp1 genes in wheat and rice

Abstract Chromosome locations for gene orthologues of the dormancy-related maize transcription factor VIVIPAROUS-1, encoded by the Vp1 locus on maize chromosome 3, were determined in wheat (Triticum aestivum L.) and rice (Oryza sativa L.) via linkage to markers on existing molecular maps using a cDNA of a wheat Vp1 orthologue as a probe in genomic Southern analyses. Vp1-orthologous loci were detected on the long arms of wheat chromosomes 3A, 3B and 3D [Xlars10 (taVp1) loci] and rice chromosome 1 (osVp1), in line with previous evidence of synteny between these regions of the rice and wheat genomes and chromosome 3 of maize. The wheat loci mapped some 30 cM from the centromeres and some 30 cM proximal to the red grain (R) loci that control seed colour and coat-imposed dormancy. This unequivocal, genetic separation of the Vp1 and R loci may offer an opportunity for improving resistance to pre-harvest sprouting in wheat by combining the coat-imposed dormancy associated with red seed colour and true embryo dormancy regulated by Vp1.

Separating homeologs by phasing in the tetraploid wheat transcriptome

BackgroundThe high level of identity among duplicated homoeologous genomes in tetraploid pasta wheat presents substantial challenges for de novo transcriptome assembly. To solve this problem, we develop a specialized bioinformatics workflow that optimizes transcriptome assembly and separation of merged homoeologs. To evaluate our strategy, we sequence and assemble the transcriptome of one of the diploid ancestors of pasta wheat, and compare both assemblies with a benchmark set of 13,472 full-length, non-redundant bread wheat cDNAs.ResultsA total of 489 million 100 bp paired-end reads from tetraploid wheat assemble in 140,118 contigs, including 96% of the benchmark cDNAs. We used a comparative genomics approach to annotate 66,633 open reading frames. The multiple k-mer assembly strategy increases the proportion of cDNAs assembled full-length in a single contig by 22% relative to the best single k-mer size. Homoeologs are separated using a post-assembly pipeline that includes polymorphism identification, phasing of SNPs, read sorting, and re-assembly of phased reads. Using a reference set of genes, we determine that 98.7% of SNPs analyzed are correctly separated by phasing.ConclusionsOur study shows that de novo transcriptome assembly of tetraploid wheat benefit from multiple k-mer assembly strategies more than diploid wheat. Our results also demonstrate that phasing approaches originally designed for heterozygous diploid organisms can be used to separate the close homoeologous genomes of tetraploid wheat. The predicted tetraploid wheat proteome and gene models provide a valuable tool for the wheat research community and for those interested in comparative genomic studies.