Enrico Coen

Complementary floral homeotic phenotypes result from opposite orientations of a transposon at the plena locus of antirrhinum

Recessive mutations at the plena (ple) locus result in a homeotic conversion of sex organs to sterile perianth organs in flowers of Antirrhinum majus. A complementary phenotype, in which sex organs replace sterile organs, is conferred by semidominant ovulata mutations. The ple locus was identified and isolated using a homologous gene, agamous from Arabidopsis, as a probe. The expression of ple is normally restricted to the inner two whorls of the flower, where sex organs develop. However, in ovulata mutants, ple is expressed ectopically in the outer two whorls of the flower and in vegetative organs. These mutants correspond to gain-of-function alleles of ple, suggesting that ple is sufficient for promoting sex organ development within the context of the flower. The plena and ovulata phenotypes result from opposite orientations of the transposon Tam3 inserted in the large intron of ple.

Transposable elements generate novel spatial patterns of gene expression in antirrhinum majus

The pallida gene of A. majus encodes a product required for the synthesis of red flower pigment. We have shown that the unstable pallida(recurrens) mutation is due to the insertion of the Tam3 transposable element near the promoter of the gene. Imprecise excision of Tam3 alters pallida gene expression and generates new spatial patterns or different intensities of flower pigmentation. Distinct spatial patterns may also result from rearrangements induced by Tam3 that alter the relative position of the pallida gene. Changes in Tam3 structure or position result in new unstable phenotypes. These findings suggest that genes may be rendered genetically hypervariable as a consequence of transposable element insertion and excision.

Comparison of genetic behaviour of the transposable element Tam3 at two unlinked pigment loci in Antirrhinum majus

SummaryIn Antirrhinum majus the transposable element Tam3 has been described at two unlinked loci pallida and nivea, both of which are required for the production of anthocyanin pigment in flowers. In each case the element is inserted in the promoter region and gives a variegated phenotype. We show that the rate of Tam3 excision at both loci is greatly affected by temperature, being approximately 1000-fold higher at 15°C compared with 25°C. Tam3 is also controlled by an unlinked gene Stabiliser, which considerably reduces excision rate. We show that the high degree of sensitivity to temperature and Stabiliser is an intrinsic property of Tam3 which is not shared by an unrelated element, Tam1. The Tam3 insertion at nivea gives rise to a series of alleles which confer reduced pigmentation, novel spatial patterns and changed instability. These are probably a result of imprecise excision and rearrangements of the Tam3 element.

Transposable elements in Antirrhinum majus: generators of genetic diversity

Abstract Several unstable mutations in Antirrhinum majus are caused by the insertion of transposable elements into genes. Excision and rearrangements of these elements can give rise to quantitative genetic variation and altered spatial patterns of gene expression. Element transposition is controlled by genetic and environmental factors and may make an important contribution to natural variation.

De novo activation of the transposable element Tam2 of Antirrhinum majus

SummaryThe nivea locus of Antirrhinum majus encodes the enzyme chalcone synthase required for the synthesis of red anthocyanin pigment. The stable allele niv-44 contains an insertion in the nivea gene (Tam2) which has all the structural features of a transposable element. We have shown that this insertion can excise from the nivea locus when niv-44 is combined with another allele (niv-99) in a heterozygote. Activation of Tam2 excision is caused by a factor tightly linked to the niv-99 allele and may be due to complementation between Tam2 and a related element, Tam1. Factors which repress the excision of Tam2 and Tam1 are also described. Repression is not inherited in a simple mendelian way. Many stable mutations may be due to the insertion of transposable elements. Our data suggest that their stability may be due to the absence in the genome of activating factors and to the presence of repressors.

Phenotypic effects of short-range and aberrant transposition in Antirrhinum majus

We describe two novel ways in which changes in gene expression in Antirrhinum majus may arise as a consequence of the Tam3 transposition mechanism. One involves excision of Tam3 from the nivea gene promoter and insertion of two new Tam3 copies 3.4 kb and 2.1 kb away, on either side of the excision site. One of the new insertions is in the nivea coding region and completely blocks production of an active gene product. This allele probably arose by a symmetrical double transposition, following chromosome replication. The second case involves a small deletion at one end of Tam3 in the pallida gene, flanked by a sequence typical of a Tam3 excision footprint. This suggests that the end of Tam3 was cleaved at an early step in an attempted transposition and re-ligated back to its original flanking sequence. The alteration restores some expression to the pallida gene, suggesting that the ends of the intact Tam3 element contain components which can actively inhibit gene expression. The implications of these findings for the mechanism of Tam3 transposition and for the effects of Tam3 on host gene expression are discussed.

Transposable Elements in Antirrhinum Majus

SUMMARY The transposable element, Tam 3, gives rise to large-scale chromosomal rearrangements at a low frequency, when it is inserted at the nivea locus of Antirrhinum majus. Some deletions that result from imprecise excision of Tam 3 provide an allelic series for functional analysis of the nivea gene promoter. Rearrangements involving deletion, dispersion and inverted duplication of flanking sequences, where Tam 3 remains in situ, have also been identified. These rearrangements have been mapped at the molecular level, and the behaviour of Tam 3 following rearrangement has been observed. It is clear that Tam 3 has enormous potential to restructure chromosomes through successive rounds of large-scale rearrangements.

Genetic and Molecular Analysis of Transposable Elements in Antirrhinum Majus

Transposable element activity in Antirrhinum majus has been studied genetically for many years. More recently the genetic analysis has been combined with molecular techniques, leading to a much greater understanding of various properties of these transposons. We have shown that the frequency of transposition of specific transposable elements can be controlled by a number of different factors including the environmental conditions under which the plants are grown and the genetic background. Transposable elements are also able to alter gene expression by imprecise excision, deletions, inversions, and chromosomal rearrangements, thus giving rise to allelic series. The knowledge gained from these studies has enabled transposable elements to be used for gene isolation. In this paper we describe the main features of the behavior of transposable elements in Antirrhinum and how they may be used to study gene action.

Self-incompatibility in Antirrhinum

Self-incompatibility (SI) systems have been reported in almost half of the families of all flowering plants (de Nettancourt, 1977). Most commonly, SI is regulated by a single multiallelic locus, with the compatibility of the pollen with respect to the stigma being controlled by the haploid nucleus (East, 1940). The genus Antirrhinum possesses such a gametophytically-regulated monofactorial SI system and, when pollen and pistil share SI (S) alleles, tube growth is arrested in the style. The significance of SI systems to plant breeders, combined with the fact that they represent some of the best defined systems of intercellular interaction in plants has resulted in their being intensively studied (Anderson et al., 1986; Nasrallah et al., 1985). For example, molecular and biochemical research has identified the female S-allele products of members of the Solanaceae as being highly charged glycoproteins with molecular weights of between 20–35 kDa (for review, see Ebert et al., 1989). Many putative S-alleles have been cloned and sequenced, and it has been recently discovered that all of the S-glycoproteins in the Solanaceae have ribonuclease activity (McClure et al., 1990), leading to the proposal that this type of SI may involve a step in which pollen tube RNA is selectively degraded. Although progress has been made in our understanding of the S-allele products of the pistil in gametophytically-controlled SI (GSI), and also in the sporophytically-controlled systems where pollen compatibility is regulated by two alleles (Bateman, 1952; Nasrallah et al., 1970), little is known of the organisation of the S-locus and of S-allele expression in the pollen.

Floral Homoeotic and Pigment Mutations Produced by Transposon-Mutagenesis in Antirrhinum majus

Many homoeotic genes affecting flower morphogenesis have been described in diverse species, although there have been few attempts to relate these in a systematic way to the mechanism of floral development (Meyerowitz et al., 1989). In 1742 it was the peloric form of Linaria vulgaris that made Linnaeus change his mind about the fixity of species (Linnaeus, 1744) and over a century later Darwin described, and was intrigued by, the behaviour of the peloric Antirrhinum (Darwin, 1868). We have used transposon-mutagenesis to generate floral homoeotic mutations in Antirrhinum with a view to studying and isolating the genes involved. One advantage of this approach is that transposon integration can be used to identify genes and subsequent excision can be used to prove that the correct gene has been isolated (Martin et al., 1985). In addition, imprecise excision can generate alleles with altered gene expression (Almeida et al., 1989).