Development of an integrated multi-petal pollinator attractant

We are interested in understanding how very complex structures are formed, and how the signals that cause them to develop are integrated across different tissues and organs of a plant.

Daisies are very complex structures. While they might appear to be simple flowers, a closer look reveals that they are neither simple, nor flowers. A daisy is a structure composed of many very small flowers, arranged together in a group that looks like one large flower. Many daisy species produce 2 types of flowers. Around the outside of the daisy head (the "capitulum") are bilaterally symmetrical flowers, in which some of the petals are absent and some are fused to make a long strap shaped structure (these "ligules" are the structures that you would assume are the petals if a daisy were really a simple flower). Inside these are spirals of small radially symmetrical flowers which have 5 tiny petals each. The whole inflorescence works together to mimic a single large flower.

However, some daisies are even more complex. The South African daisy Gorteria diffusa makes spots on some of its outer flowers, and these mimic the fly that is the main pollinator. Male flies are tricked into thinking the spots are female flies, and they visit the daisies to find mates, even attempting to copulate with the spots. For this system to work, the spots have to be very convincing mimics of female flies, and this requires very detailed coordination and integration of developmental signalling in 3 ways, which we will explore in this project.

First, the spots have to be positioned on only a subset of outer flowers, and that positioning has to be convincingly random in appearance. We will use a combination of approaches, including sequencing of all the genes expressed in developing daisies and comparing varieties with different patterns of spots, to explore the signalling that determines which outer flowers produce spots.

Second, the spots have to develop right across the ligule, which is actually formed from 4 fused petals. The outer 2 petals must have matching mirror image spot edges, which join seamlessly to the spot centre across the 2 inner petals. We will use laser microdissection to understand how this developmental integration is controlled across organ boundaries.

Third, the spot is composed of multiple cell types which must each be positioned correctly with respect to one another. We will use molecular biology approaches to identify the master regulators which specify spot development and coordinate the production of these cell types.

Taken together our data will provide exciting insights into a very complex developmental problem, while also shedding light on a highly unusual pollination system.

The aim of this proposal is to understand the development of a complex three-dimensional structure formed across multiple fused organs. We have developed Gorteria diffusa (a South African daisy) as a model system for this work because it has a very complex feature called the petal spot. This petal spot plays an important role in pollination, acting as a sexually deceptive mimic of a female fly that attracts mate-seeking male flies. The structure has evolved recently within Gorteria, which has undergone a very recent radiation within the Cape Flora.

The petal spots develop on the ligules, structures formed of multiple fused petals which are recognisable as the "rays" on the outsides of daisies. These rays are actually small, highly zygomorphic flowers in which one or more petals has aborted and the others have fused to form the ligule. The petal spot is composed of multiple specialised cell types, which must differentiate in the correct positions with respect to one another and with those relative positions integrated across the ligule. The Integration of all its traits is essential to its function, as is its positioning on a subset of ray florets, and both require exquisite coordination. It is these problems of developmental integration and coordination that we propose to address, to understand (i) how the spotted florets are positioned within the capitulum; (ii) how the spot is coordinated across 4 fused petals in a ligule, and (iii) how the different cell types of the spot are integrated into a convincing whole.

We have generated considerable preliminary data about the development of this petal spot, and are now well placed to bring together a full understanding of this complex developmental problem. Our research will provide insight into how development of complex structures is coordinated within and across organs.

IMPACT STATEMENT This project addresses a problem of basic biology - how to pattern development across multiple organs. It is therefore unrealistic to claim it will have translational impact within a short timeframe. However, it may be of interest to several important audiences. Understanding pollinator attraction may be important in maintaining crop productivity, particularly as climate changes and interactions with herbivorous and pollinating animals change. Understanding how plants attract pollinators has important implications for conservation of biodiversity. Our project therefore has potential long term impact in several sectors, as well as providing great opportunities for public engagement.

AGRIFOOD IMPACT We will disseminate data to farmers and breeders, and increase our engagement with stakeholders, through activities at the University and at NIAB. We will target outreach to these stakeholders through BJG's current contacts with Syngenta, PGRO (the processors and Growers Research Organisation) and a number of plant breeders. Results will be demonstrated at NIAB Innovation Farm Open Days and Symposia. Additional opportunities to interact with industry will be through the University's "Enterprise Tuesdays", where research can be presented to a range of interested companies.

CONSERVATION IMPACT We will disseminate data to conservation and biodiversity stakeholders through the Cambridge Conservation Initiative (CCI). The CCI links a number of conservation charities with the University of Cambridge. We will link our project to the CCI webpage, provide updates on the News section of the webpage, and advertise seminars through their diary section.

IMPACT THROUGH PUBLIC ENGAGEMENT is nationally and internationally increasingly important. Public concerns, fuelled by media coverage, are an important factor in dialogue at all policy levels concerning our management of environmental and agricultural systems. Current activities include presentations at public engagement events including National Science Week, the Cambridge Festival of Plants and the Cambridge Festival of Ideas. Enhanced public impact activities will include linking our project to the Cambridge Conservation Initiative webpage, providing News updates and advertising seminars through their diary section, and setting up a project-specific webpage giving project details and accessible introductions to the concepts involved. In addition, we will work with the Education and Interpretation staff at Cambridge University Botanic Garden to develop a display to be housed in the Asteraceae section of the Garden's historic Systematic Beds. This display will explain daisy anatomy and development to a general audience. Our experience of using daisies at previous outreach events is that the public are always fascinated to discover that a daisy is not a single flower, so extending this work with a display including models of the mature capitulum and its developmental stages will allow us to engage a wide audience with the principles of this grant. The Cambridge University Botanic Garden welcomes 330,000 casual visitors per year, plus 10,000 children on school visits.