The Primula S locus: gene function and the maintenance and breakdown of heterostyly

Lead Research Organisation: University of East Anglia
Department Name: Biological Sciences

Abstract

Pollination is important, ~70% of our food results directly from pollination as seeds, grains, fruits and berries. It is important to plants too for reproduction, which not only produces the next generation but creates variation upon which natural selection can act. This variation enables plants to adapt to changing environments and colonize new habitats. Most plants produce hermaphrodite flowers, but plants cannot move or actively choose a partner, instead they have evolved intriguing strategies to prevent self-pollination and promote cross-pollination. One of the most remarkable of these strategies is heterostyly, which uses insect pollinators (hetero=different; style=female structure). Charles Darwin observed that Primroses have two forms of flower, pin with a long style and short stamens, and thrum with a short style and longs stamens. These reciprocal positions facilitate pollen transfer by insect visitors between each flower type. A group of genes, known collectively as the S locus, controls development of the two forms of flower. A rich history of scientific research on Primroses by early botanists and geneticist made heterostyly an important textbook example of a plant breeding system. Our study will add to this work by using the latest research tools to explore how heterostyly arose, how it is controlled and why it occasionally goes wrong.

We sequenced the Primrose (Primula vulgaris) genome and identified a group of five genes that control development of its two flower types. These genes are found only in thrum and are absent in pin plants. Only those individuals that inherit this gene cluster can produce thrum flowers. Our studies show the gene which we believe is responsible for elongating stamens in thrum flowers arose ~51.7million years ago in an ancestor of all modern Primula species. We have also identified the gene responsible for controlling style length. The gene cluster contains a further three genes and in this study we will characterize their function to determine how they work together to control the floral architectures of pin and thrum flowers that facilitate insect-mediated pollination in the Primrose. Over the past 51.7 million years, 30 different species of Primula have lost the ability to produce two types of flower, they all have long styles and long stamens and never produce short styled flowers. These homostyles (Homo = same) do not use insect pollinators, but self-fertilize. Our studies suggest the gene that controls style length has been lost or damaged in these species.

These homostyle species provide an exciting opportunity to explore how one gene has, over time, on multiple separate occasions, been lost or damaged during the evolution of the different Primula species. This is important because understanding how development has gone wrong can help explain how it normally works. We will use our Primrose genome sequence to identify and characterize the corresponding S locus gene clusters in these different Primula species to discover the gene mutation that has resulted in the loss of heterostyly. Most plant and animal genes are present in two copies, one set from the mother, one from the father; this is useful, if one copy is damaged, the second copy acts as backup so function is retained, it also enables repair of the damaged copy using the other as a template. This arrangement provides genetic stability from generation to generation. One very surprising finding from our studies of genes controlling heterostyly in Primula is that they are present only as a single copy, there is no back up copy. It is therefore surprising that heterostyly is stable as any genetic damage to the key genes cannot be repaired. We do not yet have an explanation for this conundrum but our project will initiate studies to seek an explanation.

Technical Summary

Heterostyly facilitates insect-mediated pollination, it is found in ~430 Primula species which have two forms of flower, pin (long style, low anthers) and thrum (short style, high anthers); each form is self-incompatible (SI). We sequenced the Primula genome, identified the S locus as a cluster of 5 genes, CCMT, GLOT, CYPT, PUMT and KFBT, found only in thrums and absent in pins, and dated its origin to ~51.7 MYA before speciation in the Primulaceae. We characterised 2 independent long homostyle (LH) mutants (long style, high anthers) and showed that CYPT controls style length, and a short homostyle (SH) mutant (short style, low anthers) that suggests GLOT controls anther elevation. We will analyse 2 new SH mutants to confirm this role for GLOT. There are no mutants for the other 3 genes; we will use gain and loss of function transgenics to define their role in heterostyly and SI. Analysis of S locus gene expression in LH and SH mutants suggests CYPT and GLOT cross-regulate other S locus genes. We will define this regulation using transgenics and transcriptomics to reveal how di-morphic flower development is controlled by the S locus. Since its single origin, Primula heterostyly has broken down ~30 times leading to species with self-fertile homostyle flowers (high anthers, long styles). Our data show the S locus is hemizygous, not heterozygous; heterostyles must therefore arise by mutation of S locus genes. Hemizygosity raises a conundrum: how is the S locus stable if there is no opportunity for recombination repair of such mutants? We will use oligo-select capture of the S locus to define sequence variation in different heterostyle populations of P. vulgaris, and to analyse different homostyle species to determine whether independent loss of heterostyly is due to mutation in CYPT. We will also identify ancestral S locus architecture in out-groups. These studies will explore the origin, maintenance and breakdown of this pollination syndrome.

Planned Impact

This project focuses on one of the most successful pollination syndromes involving insect-mediated pollination. Heterostyly evolved in the Primulaceae ~45 million years ago before speciation and widespread global distribution across five continents. Understanding the control of plant pollinator interactions is of direct relevance to food security and biodiversity in a changing world. ~70% of our food is the direct product of pollination and as climates change, the balance of plant pollinator interactions in crops and native species will also change; this imbalance, coupled to global pollinator decline, means that understanding the mechanisms controlling insect-mediated pollination will identify opportunities to adapt and modify genetic systems from model species into crop species.

Importantly, this highly adapted pollination system has, over the past 45 million years, broken down at 30 independent times, leading to self fertility and loss of reliance on insect pollinators. Studying how a system breaks is valuable in understanding how it works and therefore how it can be exploited. Following Darwin's discovery of this adaptation, historical studies in Primula mark landmark events in the history of genetics and the neoDarwinian synthesis. In combination, this project will have academic, technological and societal impact and will provide tools and resources that could be developed by others to have economic impact and support the knowledge economy.

Who will benefit and how?

Academic community: Academic beneficiaries include plant scientists working on reproduction and pollination, evolutionary biologists focused on gene and genome evolution, speciation and co-adaptation, developmental geneticists exploring plant architecture and cell-cell signalling, and ecologists studying plant-insect interactions, habitat loss and colonization, plus science historians interested in the contribution of this breeding system to modern scientific methods. Knowledge and insight from studies in one species are not limited to that species and can facilitate step change discoveries in unrelated areas.

Informed amateurs: Amateur gardening in the UK has an estimated economic value of at least £89 billion with the average adult spending 2-3 hours per week gardening; as a nation we spend £5bn a year on gardening. Public access to science and the understanding of important scientific advances need to be presented in a stimulating way that people can relate to. The huge audience of informed amateurs and enthusiastic gardeners will benefit from the accessibility of complex scientific knowledge through the vehicle of the familiar common primrose.

Future scientists: Engaging young audiences in the value of science and scientific discovery needs simple accessible stories. The sex life of the common primrose provides an example through which a spectrum of biological concepts can be presented in simple and appealing ways. Understanding the relevance and significance of such concepts through accessible and intriguing stores is an ideal way to engage, educate and enthuse future young scientists.

Agbiotech and horticulture industries: Primula is a key northern-clime horticultural crop, estimated EU annual market value ~400M Euro. Development of molecular genetic tools such as plant regeneration and transformation and genome resources in Primula has value and applications. Understanding the mechanisms that control floral reproductive architecture, and its impact on plant pollinator interactions, has the potential to identify genes and pathways that may be of value in engineering floral traits to increase reproductive productivity, develop responses to pollinator decline, or even prevent self pollination to facilitate hybrid seed production. This project has the potential for economic and food security impact by providing tools and resources that could be exploited and applied for commercial benefit.

Publications

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Related Projects

Project Reference Relationship Related To Start End Award Value
BB/P022081/1 01/10/2017 31/03/2019 £735,335
BB/P022081/2 Transfer BB/P022081/1 01/04/2019 31/03/2022 £418,476
 
Description The genome sequence of Primula vulgaris has been published, and a transformation system has been developed and published. Gene function studies are underway as outlined in the proposal and are providing valuable insights, including gene silencing and promoter reporter construct analysis in Primula a manuscript is in preparation from the studies of one of the genes. The oligo-select sequencing work outlined in the proposal has provided new insights into the conservation of the S locus gene cluster in different Primula species. Comparative genome data has also revealed insight into the evolutionary divergence of three different primula Species, and a manuscript is under review in Nature Plants.
Exploitation Route On going work from this project will be published in open access journals when the studies are complete and these data will be of benefit to others working in the field of plant reproductive development and pollination. There are potential longer term applications for the manipulation of plant reproduction for agroicultural purposes.
Sectors Agriculture, Food and Drink,Education,Leisure Activities, including Sports, Recreation and Tourism,Culture, Heritage, Museums and Collections

 
Description Public engagement through press releases, print media stories, radio interview, and social media.
First Year Of Impact 2018
Sector Education,Other
Impact Types Cultural,Societal

 
Title A plant transformation system for Primula 
Description Publication of a plant transformation system for Primula 
Type Of Material Biological samples 
Year Produced 2018 
Provided To Others? Yes  
Impact The first report of a transformation system or Primula, will support us, and others in addressing the function of genes controlling heterostyly. 
 
Title Release of genome sequence data and assembly for Primula vulgaris 
Description Public release of the genome assembly, and underpinning sequence data for Primula vulgaris. 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
Impact Publicaton and open availability of the complete geome squnce for Primula vulgaris 
 
Description Japanese collabrorative partnership 
Organisation Kobe University
Country Japan 
Sector Academic/University 
PI Contribution I initiated a collaborative partnership with Prof. Ryo Ohsawa (Tsukuba University), Prof Yasuo Yasui (Kyoto University) and Dr Yasuko Yoshida (Kobe University). Dr Yoshoda was awarded University sabbatical leave to join my laboratory in Norwich for a year, Dr Jonathan Cocker, a UEA-funded PhD student who worked on this project was awarded JSPS funding to visit our collaborating partners in Japan for 3 months. These exchange visits led to the application for UKRI-JSPS joint funding for a collaborative project aimed at comparing heterostyly in two Primula species and two Buckwheat species Unfortunately this application was not successful. This joint funding application was an extension of a previous collaboration set up before the grant started, but has specifically been underpinned by research on this current BBSRC grant.
Collaborator Contribution Our partners have provided plant material, DNA and expertise to enable development of a collaborative research program, We jointly conceived and wrote a collaborative research project anf sought UKRI-SJPS funding. Unfortunately the application was not successful, but the collaboration is still active and we will seek to undertake the work though other routes.
Impact None yet
Start Year 2018
 
Description Japanese collabrorative partnership 
Organisation University of Kyoto
Country Japan 
Sector Academic/University 
PI Contribution I initiated a collaborative partnership with Prof. Ryo Ohsawa (Tsukuba University), Prof Yasuo Yasui (Kyoto University) and Dr Yasuko Yoshida (Kobe University). Dr Yoshoda was awarded University sabbatical leave to join my laboratory in Norwich for a year, Dr Jonathan Cocker, a UEA-funded PhD student who worked on this project was awarded JSPS funding to visit our collaborating partners in Japan for 3 months. These exchange visits led to the application for UKRI-JSPS joint funding for a collaborative project aimed at comparing heterostyly in two Primula species and two Buckwheat species Unfortunately this application was not successful. This joint funding application was an extension of a previous collaboration set up before the grant started, but has specifically been underpinned by research on this current BBSRC grant.
Collaborator Contribution Our partners have provided plant material, DNA and expertise to enable development of a collaborative research program, We jointly conceived and wrote a collaborative research project anf sought UKRI-SJPS funding. Unfortunately the application was not successful, but the collaboration is still active and we will seek to undertake the work though other routes.
Impact None yet
Start Year 2018
 
Description Japanese collabrorative partnership 
Organisation University of Tsukuba
Country Japan 
Sector Academic/University 
PI Contribution I initiated a collaborative partnership with Prof. Ryo Ohsawa (Tsukuba University), Prof Yasuo Yasui (Kyoto University) and Dr Yasuko Yoshida (Kobe University). Dr Yoshoda was awarded University sabbatical leave to join my laboratory in Norwich for a year, Dr Jonathan Cocker, a UEA-funded PhD student who worked on this project was awarded JSPS funding to visit our collaborating partners in Japan for 3 months. These exchange visits led to the application for UKRI-JSPS joint funding for a collaborative project aimed at comparing heterostyly in two Primula species and two Buckwheat species Unfortunately this application was not successful. This joint funding application was an extension of a previous collaboration set up before the grant started, but has specifically been underpinned by research on this current BBSRC grant.
Collaborator Contribution Our partners have provided plant material, DNA and expertise to enable development of a collaborative research program, We jointly conceived and wrote a collaborative research project anf sought UKRI-SJPS funding. Unfortunately the application was not successful, but the collaboration is still active and we will seek to undertake the work though other routes.
Impact None yet
Start Year 2018
 
Description Japanese partnership (still active 2020) 
Organisation Japan Society for the Promotion of Science (JSPS)
Country Japan 
Sector Public 
PI Contribution I initiated a collaborative partnership with Prof. Ryo Ohsawa (Tsukuba University), Prof Yasuo Yasui (Kyoto University) and Dr Yasuko Yoshida (Kobe University). Dr Yoshida applied for University sabbatical leave to join my laboratory in Norwich for a year, Dr Jonathan Cocker, a UEA-funded PhD student who worked on this project applied for JSPS funding to visit our collaborating partners in Japan for 3 months to jointly work on heterostyly in Primula sieboldii (Jjapan) and Primula vulgaris (UK).
Collaborator Contribution Dr Yoshida applied for and secured funding to visit our lab and Prof. Ohsawa and Professor Yasui agreed to host Dr Cocker , who was subsequently funded on the BBSRC grant . on a JSPS Fellowship to gain experience of genomics approaches as well as progress the comparative genomics aspects of the grant. Our Japanese partners have contributed plant material , expertise and unpublished data that have been valuable in developing a collaborative venture. . Work arising from this collaboration is the de novo assembly of the Primula sieboldii genome. This work is ongoing as a collaboration between our lab and our Japanese collaborators ans will enable integration of the genome assembly with molecular genetic mapping data. A publication is anticipated from this work
Impact Research will lead to a join manuscript
Start Year 2016
 
Description Japanese partnership (still active 2020) 
Organisation University of Kyoto
Country Japan 
Sector Academic/University 
PI Contribution I initiated a collaborative partnership with Prof. Ryo Ohsawa (Tsukuba University), Prof Yasuo Yasui (Kyoto University) and Dr Yasuko Yoshida (Kobe University). Dr Yoshida applied for University sabbatical leave to join my laboratory in Norwich for a year, Dr Jonathan Cocker, a UEA-funded PhD student who worked on this project applied for JSPS funding to visit our collaborating partners in Japan for 3 months to jointly work on heterostyly in Primula sieboldii (Jjapan) and Primula vulgaris (UK).
Collaborator Contribution Dr Yoshida applied for and secured funding to visit our lab and Prof. Ohsawa and Professor Yasui agreed to host Dr Cocker , who was subsequently funded on the BBSRC grant . on a JSPS Fellowship to gain experience of genomics approaches as well as progress the comparative genomics aspects of the grant. Our Japanese partners have contributed plant material , expertise and unpublished data that have been valuable in developing a collaborative venture. . Work arising from this collaboration is the de novo assembly of the Primula sieboldii genome. This work is ongoing as a collaboration between our lab and our Japanese collaborators ans will enable integration of the genome assembly with molecular genetic mapping data. A publication is anticipated from this work
Impact Research will lead to a join manuscript
Start Year 2016
 
Description 25th ICSPR conference in Gifu, Japan 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Attendance at the 25th International Congress on Sexual Plant Reproduction in GIGU Japan. Posters presented by Jinhong Li and Jonathan Coker who are working on this grant.
Year(s) Of Engagement Activity 2018
 
Description Conference presentation in China 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited attendance at Cold Spring Harbor Asia conference in Suzhou, China to resent our research.
Year(s) Of Engagement Activity 2017
 
Description Departmental seminar including undergraduate students 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Undergraduate students
Results and Impact Research seminar at University of Plymouth aimed at undergraduate, postgraduate and academic audience
Year(s) Of Engagement Activity 2020
 
Description Invitation to present course at Cambridge Botanic Gardens on Primula heterostyly 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Course/workshop at Cambridge Botanic Gardens, .on Primula Heterostyly. To be delivered on 28 March 2018. The planning falls in this reporting period, the details of the event will be included in next years report.
Year(s) Of Engagement Activity 2018
 
Description Invited Conference presentation on our research 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited conference presentation at Plant Science meeting in Padua Italy
Year(s) Of Engagement Activity 2017
 
Description JIC Schools openday 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Schools
Results and Impact Poster presentation and discussion with students at the JIC Schools open day.
Year(s) Of Engagement Activity 2018
 
Description John Innes Centre School engagement event 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Schools
Results and Impact Annual John Innes Centre School engagement event hands on presentation to participants as well as poster presentation and other discussion and engagement with researchers engaged on this project. Over 100 pupils attend the event and had the opportunity to engage with our research team, The purpose was to enhance awareness of Science, plant science, research and on a detailed level the importance of plant reproduction, insect pollinators and food security.
Year(s) Of Engagement Activity 2018
 
Description Press releaae and media stories 1 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Press release and multiple media stories following publication of our paper on the Primula genome
Year(s) Of Engagement Activity 2018
 
Description Press release and media stories 2 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Press release and multiple media stories following our paper on the transformation of Primula
Year(s) Of Engagement Activity 2018
 
Description Radio Norfolk 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Media (as a channel to the public)
Results and Impact Interview on Radio Norfolk about our publication of the Primula genome sequence
Year(s) Of Engagement Activity 2018
 
Description Resaerch seminar at Chongqing University Vhina 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact Research seminar given at Chongqing university, China.
Year(s) Of Engagement Activity 2018
 
Description Research Presentation to general scientific audience 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Professional Practitioners
Results and Impact Research presentation to wide scientific audience at the Norwich Research Park Darwin Day symposium
Year(s) Of Engagement Activity 2020
 
Description SMBE conference in Yokohama 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invitation to speak at the Society for Molecular Biology and Evolution in Yokohma Japan.
Year(s) Of Engagement Activity 2018
 
Description School visit 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Visit to Fulford School in York to speak to year 11, 12, and 13 students about Primula heterostyly, Darwin, evolution and careers in plant science,
Year(s) Of Engagement Activity 2020
 
Description Twitter 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Frequent tweets on Primula heterostyly and engagement on social media around flower development and plant reproduction. (@PMGilmartin 2600+ followers)
Year(s) Of Engagement Activity 2017,2018,2019,2020
 
Description Wymondham College VIth Form visit 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Wymondham College VIth Form visit to discuss our research
Year(s) Of Engagement Activity 2019