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| Parental purple (Wp, left) and progeny pink (wp, right) soy-bean flowers. From Zabala and Vodkin 2007. |
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| Highly recommended reading or viewing, |
In 1989 a variegated chimeric* flower mutant spontaneously appeared in a field of soy-beans during the cross-breading of white and purple soy-beans.
Soybeans breeders subsequently obtained a stable pink flower line -- which they call a wp mutant -- from the unstable variegated plant. From purple and white parents they obtained a rare pink offspring
Investigation of the altered structure of the DNA in the gene concerned with pink flower colour revealed that the new mutation is the outcome of insertion of a novel genetic parasite within the gene for flower colour.
The genetic parasite itself turned out to be a remarkably complex object in that it has randomly captured and scrambled the DNA from several other plant genes within its own genetic structure.
It illustrates there can be layers and layers of complexity and chaotic randomness to the process by which novel gene variants can emerge in natural cross-breeding to confer new flower colour patterns. Not only is flower colour changed, but the wp mutant has 22% larger seed size, less oil and more protein, so there are practical implications to this work too.
Two key scientific papers published by Gracia Zabala and Lila Vodkin giving details of this soy-bean flower colour investigation are excerpted below, but most of all the Pundit is grateful for being able to reproduce the image at the top, comparing the purple Wp parental soy flowers and the pink wp mutant flowers whose evolutionary history and emergence as an accidental breeding event have been carefully documented by soybean geneticists. The only genetic engineering involved was done by the plants themselves and their own promiscuous genetic parasites.
The wp Mutation of Glycine max Carries a Gene-Fragment-Rich Transposon of the CACTA Superfamily
Gracia Zabala and Lila O. Vodkin 2005
Summary
We used soybean (Glycine max) cDNA microarrays to identify candidate genes for a stable mutation at the Wp locus in soybean, which changed a purple-flowered phenotype to pink, and found that flavanone 3-hydroxylase cDNAs were overexpressed in purple flower buds relative to the pink. Restriction fragment length polymorphism analysis and RNA gel blots of purple and pink flower isolines, as well as the presence of a 5.7-kb transposon insertion in the wp mutant allele, have unequivocally shown that flavanone 3-hydroxylase gene 1 is the Wp locus. Moreover, the 5.7-kb insertion in wp represents a novel transposable element (termed Tgm-Express1) with inverted repeats closely related to those of other Tgms (transposable-like elements, G. max) but distinct in several characteristics, including the lack of subterminal inverted repeats. More significantly, Tgm-Express1 contains four truncated cellular genes from the soybean genome, resembling the Pack-MULEs (Mutator-like transposable elements) found in maize (Zea mays), rice (Oryza sativa), and Arabidopsis thaliana and the Helitrons of maize. The presence of the Tgm-Express1 element causing the wp mutation, as well as a second Tgm-Express2 element elsewhere in the soybean genome, extends the ability to acquire and transport host DNA segments to the CACTA family of elements, which includes both Tgm and the prototypical maize Spm/En.
Introduction
Soybean (Glycine max) plants display diverse coloration in their flowers, seed coats, hypocotyls, and trichome hairs (pubescence). Genetic studies have identified at least five loci affecting flower pigmentation, W1, W3, W4, Wm, and Wp, and these loci are distinct from those (I, R, and T) determining seed coat and pubescence coloration (Bernard and Weiss, 1973; Palmer and Kilen, 1987; Groose and Palmer, 1991). Cultivars with a purple flower phenotype have a W1W1 genotype, while those with white flowers have w1w1. Pink-flowered plants (W1_wpwp) were first observed in 1989 (Stephens and Nickell, 1991) and were derived from a mutable, chimeric plant having purple and pink flowers on the same plant. Interestingly, the derived pink flower lines averaged 22% higher in seed weight, 4% higher in protein, and 3% lower in oil compared with the purple-flowered lines derived from the chimeric plant (Stephens and Nickell, 1992; Stephens et al., 1993). The inheritance of the mutable phenotype and derived pink and purple lines showed a high rate of instability (Johnson et al., 1998).
The soybean color phenotypes are likely the result of mutations affecting different enzymes of the anthocyanin and proanthocyanidin pathways. Molecular data indicated that the W3 locus encodes a dihydroflavonol reductase (Fasoula et al., 1995). The I locus corresponds to a 27-kb-long chalcone synthase gene cluster that exhibits a unique tissue-specific gene silencing mechanism in the seed coats mediated by short-interfering RNA (Todd and Vodkin, 1996; Senda et al., 2004; Tuteja et al., 2004). Recently, it has been shown that the pleitropic T locus that affects seed coat pigmentation and cell wall integrity encodes a flavonoid 3′ hydroxylase (Toda et al., 2002; Zabala and Vodkin, 2003) (Figure 1).
Fragment length polymorphisms between the purple and pink mutant isolines supported the discovery of a transposon insertion in the wp mutant allele, and the aberrant size F3H transcripts in the flower buds and seed coats of the pink-flowered line demonstrated that the Wp locus of soybean encodes an F3H gene. The DNA sequence of a 5.7-kb insertion in the mutant wp allele revealed a transposable element member of the CACTA family of transposons (Tgm, Spm, and Tam) (Vodkin et al., 1983; Rhodes and Vodkin, 1988). However, the element in the wp pink flower mutation differed from the other Tgm family members previously characterized in that it lacks the subterminal repeats and was laden with at least four genic fragments picked up from the host genome. The potential of CACTA elements to carry truncated genic fragments resembles that of the Pack-MULEs found in maize (Zea mays), rice (Oryza sativa), and Arabidopsis thaliana (Talbert and Chandler, 1988; Yu et al., 2000; Turcotte et al., 2001; Jiang et al., 2004) and the Helitron discovered more recently in maize (Lal et al., 2003; Gupta et al., 2005). It has been speculated that these types of elements have the potential to create novel genes through the rearrangement and fusion of noncontiguous genomic sequences captured by the transposons.
The discovery of the wp insertion element, named Tgm-Express1, adds a new level of transposable element complexity to the CACTA family of elements that includes the Spm/En (Suppressor-mutator/Enhancer elements), one of the original maize transposable elements first described genetically in the 1940s by Barbara McClintock and Peter Peterson (reviewed in Wessler 1988; Gierl et al., 1989). Another occurrence of the capture of genomic sequences by a CACTA-type element has been shown for the Tpn1 of the Japanese morning glory (Ipomoea tricolor) (Takahashi et al., 1999). The existence of the Tgm-Express and CACTA family elements in other plant species indicates that the ability to acquire, recombine, and replicate host genomic DNA fragments may be widespread. In addition, they represent only a few genetically described movements of cellular genes revealed as insertional inactivations of the target genes rather than by extrapolation from data mining of high-throughput genome sequencing data.
Results
Two stable isolines, one with purple flowers (WpWp) and a second with pink flowers (wpwp), were recovered from the progeny of a mutant variegated plant that arose spontaneously in the field from a cross that was expected to produce purple- or white-flowered plants.
Figure 2 confirms with an RNA gel blot using the full-length Gm-c1012-683 EST clone as probe that F3H cytoplasmic transcripts are not detectable in flower buds with the pink flower (wp/wp) genotype compared with those of the purple-flowered line (Wp/Wp).
Plant Cell. 2005 October; 17(10): 2619–2632. doi: 10.1105/tpc.105.033506 PMCID: PMC1242261
Novel exon combinations generated by alternative splicing of gene fragments mobilized by a CACTA transposon in Glycine max
Gracia Zabala and Lila Vodkin 2007
Background
The recent discoveries of transposable elements carrying host gene fragments such as the Pack-MULEs (Mutator-like transposable elements) of maize (Zea mays), rice (Oryza sativa) and Arabidopsis thaliana, the Helitrons of maize and the Tgm-Express of soybeans, revealed a widespread genetic mechanism with the potential to rearrange genomes and create novel chimeric genes affecting genomic and proteomic diversity. Not much is known with regard to the mechanisms of gene fragment capture by those transposon elements or the expression of the captured host gene fragments. There is some evidence that chimeric transcripts can be assembled and exist in EST collections.
Results
We report results obtained from analysis of RT-PCR derived cDNAs of the Glycine max mutant flower color gene, wp, that contains a 5.7-kb transposon (Tgm-Express1) in Intron 2 of the flavanone 3-hydroxylase gene (F3H) and is composed of five unrelated host gene fragments. The collection of cDNAs derived from the wp allele represents a multiplicity of processed RNAs varying in length and sequence that includes some identical to the correctly processed mature F3H transcript with three properly spliced exons. Surprisingly, the five gene fragments carried by the Tgm-Express1 were processed through complex alternative splicing as additional exons of the wp transcript.
Conclusion
The gene fragments carried by the Tgm inverted repeat ends appear to be retained as functional exons/introns within the element. The spliceosomes then select indiscriminately the canonical intron splice sites from a pre-mRNA to assemble diverse chimeric transcripts from the exons contained in the wp allele. The multiplicity and randomness of these events provide some insights into the origin and mechanism of alternatively spliced genes.
BMC Plant Biology 2007, 7:38 doi:10.1186/1471-2229-7-38
The electronic version of this article is the complete one and can be found online at: http://www.biomedcentral.com/1471-2229/7/38
© 2007 Zabala and Vodkin; licensee BioMed Central Ltd.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
* The original chimera is a hybrid beast from Roman mythology.
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| The chimera of Arezzo (courtesy of Wikipedia/Lucarelli) |
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