Award details

Coordination of meiotic recombination and prophase I progression in plants: the role of retinoblastoma (RBR)

Reference	BB/K007505/1
Principal Investigator / Supervisor	Professor Chris Franklin
Co-Investigators / Co-Supervisors	Dr James Higgins, Dr Eugenio Sanchez-Moran
Institution	University of Birmingham
Department	Sch of Biosciences
Funding type	Research
Value (£)	476,722
Status	Completed
Type	Research Grant
Start date	30/06/2013
End date	31/08/2016
Duration	38 months

Abstract

During meiosis the formation of genetic crossovers (CO) is highly controlled to ensure that each pair of homologous chromosomes receives at least one CO and that additional COs are prevented from occurring in an adjacent chromosome region (CO interference). The control of CO frequency and distribution requires coordination between the recombination machinery and extensive chromosome remodelling during meiotic prophase I. How this is achieved remains poorly understood, yet it is of great importance since meiosis provides the basis of genetic variation that underpins plant (and animal) breeding. This project is based on our recent work that has identified for the first time that the cell-cycle control protein retinoblastoma (RBR) is essential for the coordinated formation of COs in Arabidopsis. Based on evidence thus far we hypothesize that RBR provides a direct link between the sites of recombination and the cell-cycle machinery. The first part of the experimental programme will investigate the role of RBR in CO formation. Using a combination of molecular cytogenetics, high-resolution microscopy and proteomics we will determine how RBR interfaces with the recombination machinery and/or components of the chromosome axes to ensure that DNA double-strand breaks (DSBs) are processed to form COs. Second, we will address the inter-relationship between RBR and the cell-cycle machinery to ensure recombination and chromosome remodeling are coordinated. The hypotheses that the phosphorylation status of RBR is a key factor and that it may act as an "environmental sensor" will be investigated. We will seek to confirm that the role of RBR is conserved in crop species (brassica and barley) and explore a possible link between RBR and the function of the Ph1 locus in wheat. These studies are closely aligned with the BBSRC's strategic priority area of Food Security and will inform the development of methodology for the manipulation of meiotic recombination in crop species.

Summary

It is predicted that as a result of population increase, industrialization and climate change global demand for food will double by 2050 (Reaping the Benefit: Royal Society Review 2009). To meet this challenge it will be necessary to develop new crop varieties that are improved in various ways, for example, increased nutritional value and yield and tolerance to biotic and abiotic stresses. Although GM has its part to play, the development of new varieties will remain highly dependent on methodologies derived from traditional breeding methods which are reliant on meiotic recombination to generate variation through the formation of genetic crossovers (COs) which results in new combinations of genes. Understanding the factors that control meiotic recombination is of great significance for the improvement of crop-breeding since it is now clear that many species notably cereals, possess large regions on their chromosomes that rarely recombine. This presents a significant barrier for the introgression of new genetic traits. Hence, to overcome this problem we need to know how the frequency and distribution of COs are controlled. In addition an estimated 50% of plants are polyploid, which creates an additional level of meiotic regulation that we need to understand. Studies indicate that the controlled formation of COs is dependent on the interplay between the proteins that catalyse recombination and those that regulate the extensive remodelling of chromosomes during prophase I of meiosis. How these processes are coordinated remains poorly understood. Recently, we obtained the first evidence in any organism, that the retinoblastoma protein Rb (RBR in plants) plays an essential role in the control of meiotic recombination. The function of Rb in mitotic cell-cycle control and as a tumour-suppressor has been extensively studied. However, it is generally difficult to study its role in development in adult organisms as loss of Rb results in lethality during embryogenesis. Using aspecific rbr mutant we have been able to overcome this problem. Our studies reveal an important coordinating role for RBR in meiosis through a direct interaction with the meiotic chromosomes at the sites of recombination. We now propose to investigate how RBR exerts this coordinating role. In particular we aim to establish how RBR links with components of the cell-cycle machinery to ensure that chromosome remodelling occurs in a timely fashion in relation to meiotic recombination and investigate if it functions as "sensor" to link meiosis with changes in temperature. Although studies will primarily be conducted in the model plant Arabidopsis, as this is the most experimentally tractable system, these will be complimented by additional work in crop species. Our experimental strategy will be based around molecular cytogenetics using antibodies that recognize key meiotic proteins combined with high resolution light microscopy to study meiotic prophase I in wild-type plants and a range of meiotic mutants, including a line lacking RBR. Interactions between RBR, meiotic proteins and cell-cycle components will be studied using mass-spectrometry to analysis protein complexes that have been precipitated from meiocytes using an anti-RBR antibody. Yeast two-hybrid analysis will be used as an alternative strategy and to confirm putative interactions. A further aspect of the analysis will be to investigate the role of RBR in coordinating chromosome pairing and recombination in polyploid organisms. Specifically, we will investigate the relationship between RBR and the function of the Ph1 locus in wheat which is important for the control of chromosome pairing in this hexaploid species. We anticipate these studies will provide important new insights into the control of recombination during meiosis that will be of benefit for plant breeding by providing approaches to enable changes in CO frequency and distribution.

Impact Summary

The proposed project fulfils several BBSRC strategic aims: "maintaining world-class UK Bioscience by supporting the best people and best ideas", "providing skilled researchers needed for academic research". It has particular relevance to BBSRC's strategic priorities in regard to crop breeding/food security. Plant breeding is reliant on the creation of genetic variation that arises during meiotic recombination. It has been known for some time that large segments of chromosomes in crop species, notably cereals, rarely recombine. This is a serious barrier to the breeding of lines with new traits. Thus there is a real need to understand the major factors that control the frequency and distribution of meiotic crossovers during recombination. An additional issue, that is particularly relevant to plants including key crops such as wheat and oilseed rape, is that they are polyploids. Although we know something about the genetic loci that are important for meiotic stability in these species, such as Ph1 in wheat and BHP1 in B. napus, our understanding of how accurate homologous chromosome pairing is established remains to be established. This project is designed to increase our fundamental understanding of the mechanistic basis of these complex issues, thus providing a platform for future translational studies. It is worth pointing out that our fundamental work on meiosis in Arabidopsis has directly informed the translational work that is being conducted in barley and Brassica in our BBSRC project BB/F019351/1 and EU FP7 MEIOsys project. We have links with a number of breeding companies and have regularly attended annual meetings of the UK-Brassica Research Community (UK-BRC) and Monogram, where we have presented talks. We propose to continue this as these meetings provide an ideal forum to present our work to wider crop research community and to plant breeders. As part of our EU FP7 MEIOsys project we plan to have a workshop in 2015 that will directly engage with plant breeders. Hence, this will provide an additional opportunity to present the work in this project to the plant breeding community. FCHF and ESM have been actively involved in interacting with the end-users, media, public and schools to make science more accessible for many years. FCHF has contributed to writing press releases for both the University and BBSRC. The work on barley meiosis has featured in a BBSRC road show and was also a feature in BUZZ magazine (2009), a widely distributed University magazine. FCHF has been involved in the BBSRC Schools Liaison scheme in the past. FCHF and ESM co-organized science activities at "ThinkTank" (Science and Technology museum in Birmingham) for the public. This introduced the public to chromosomes and chromosome segregation allowing "hands-on experience". FCHF also co-organized a similar event at the International Plant Sexual Reproduction meeting in Bristol in 2010. We are now developing links with NIAB to join their Innovation Farm ERDF partnership, which will provide an opportunity to showcase the work in this project in a workshop to the general public and plant breeders.

Committee	Research Committee B (Plants, microbes, food & sustainability)
Research Topics	Crop Science, Plant Science
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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