Comparative and Functional Analysis of a Genetic Pathway Controlling Floral Asymmetry

Although much progress has been made in understanding how genes influence the development of characters, such as the organs in a flower or the eye of a fly, we still know little about how such characters evolved in the first place. Ever since Darwin, it has been thought that novel characters do not emerge completely anew but through modification or tinkering with what was already there before. We now know that this tinkering involved changes in genes but precisely how such changes can lead to the production of novel characters or traits still remains to be elucidated. One way of approaching this problem is to take an example of a novel trait, understand how it is genetically controlled in a species that has the trait and then compare this to the situation in a species that lacks the trait. By defining the genetic differences that allow one species but not the other to produce the trait, it should be possible to understand how particular components may have changed so as to allow the novel character to have emerged during evolution. In this proposal we have chosen to analyse the evolution of a key trait in plants - flower asymmetry. Many flowers, such as buttercups, are radially symmetrical, having similar petals all the way round the flower. Other flower species, such as orchids and snapdragons, have asymmetric flowers in which the upper petals are very different from the lower ones. This asymmetric condition is thought to have evolved many times independently in flowering plants from the symmetrical state as a specialized adaptation to pollinators. The best studied case of floral asymmetry is in the snapdragon (Antirrhinum) where four key genes that regulate the process have been isolated and analysed. However, one difficulty with studying these genes further has been the inability to introduce modified versions of the genes into snapdragons. This obstacle has recently been overcome by the development of a new way of conveniently introducing genes into snapdragons. The aim of this proposal is to exploit this development to analyse further how genes controlling floral asymmetry work in snapdragons and to compare this to the situation in Arabidopsis, a plant that has radially symmetrical flowers and therefore represents a more ancestral condition. This should give important insights into how novel characters can be derived through genetic change.

We propose to analyse the function and divergence of a genetic pathway that underlies a key evolutionary innovation, floral asymmetry. Floral asymmetry is thought to have evolved many times independently and has been most extensively studied in Antirrhinum, where a pathway of four regulatory genes, CYC, DICH, RAD and DIV, has been defined. We will exploit recent advances in Antirrhinum transformation technology to analyse the interactions between these genes and how they modify plant growth. Antirrhinum plants carrying CYC regulatory regions driving a marker gene will be characterised to determine the sequences needed for dorsal-specific expression of CYC. The CYC promoter will then be used to drive expression of CYC or its Arabidopsis counterpart, TCP1, in a cyc mutant background to determine the extent of functional conservation. In parallel, tcp mutants of Arabidopsis will be characterised to determine the role of TCP1 in its normal context. Antirrhinum plants carrying inducible CYC constructs will also be generated and analysed to determine the role of CYC at specific times and locations and its effect on the downstream target, RAD. The requirement of CYC-binding sites in RAD for gene activation will also be investigated. The role of RAD will be studied by through ectopic expression and induction studies and by analysing the function of novel proteins that interact with RAD. The results should provide insights into some of the key developmental and evolutionary steps that underlie the emergence of floral asymmetry.