MOLECULAR MECHANISMS OF MEIOTIC RECOMBINATION

Our germ cells, sperm and egg, form the basis for the generation of new human life. Upon fertilisation, the DNA contained within sperm and egg is combined to provide the full complement of genetic information that defines a unique human being. Germ cells are produced through meiosis, a process of cell division in which genetic information contained in our DNA is reshuffled, to generate diversity in the offspring. Meiosis is a fascinating and complicated process that can frequently go wrong, resulting in either sperm or egg with the incorrect number of chromosomes, or in no germ cell being produced at all, causing miscarriage and infertility. It may also result in children being born with genetic diseases such as Down's syndrome. It is important that we explain the molecular processes of meiosis, so that we can understand how genetic diversity is generated and we can diagnose and possibly intervene when meiosis goes wrong.

At the heart of meiosis is the process of genetic exchange between pairs of matching parental chromosomes. During genetic exchange, the chromosomes of each matching pair swap segments of DNA, thus generating novel DNA sequences, in a process known as meiotic recombination. In preparation for the exchange, the chromosomes in each pair must first be brought into alignment with each other. To be paired correctly, the long coils of chromosomal DNA must take up a specific shape, needed to execute the recombination reaction. it is also important that the chromosomes of each matching pair check their alignment, by exchanging homologous DNA strands with their partner at multiple sites along their arms.

A large set of proteins are required to carry out the complicated steps of meiotic recombination. Previous research has shown that a large protein structure known as the synaptonemal complex (SC) acts as a molecular scaffold, keeping matching chromosomes together in the correct way for the exchange of DNA to take place. At the same time, dedicated protein enzymes known as recombinases perform the remarkable reactions of homology search and DNA-strand exchange between the aligned chromosomes. The successful outcome of this complex set of reactions leads eventually to the generation of a novel combination of genetic information in each sperm cells, in preparation for fertilisation.

Despite the great importance of the reaction of genetic exchange between homologous chromosomes in meiosis, we still know relatively little about the molecular mechanisms that determine the three-dimensional organisation of meiotic chromosomes and their recombination. We plan to discover how

Our research will help us improve our molecular understanding of meiotic recombination and explain how defects in its mechanisms, either accidental or inherited, can lead to infertility, miscarriage and genetic disease. In addition, our work will improve our knowledge of the different ways in which human cells organise and handle large chromosomal DNA molecules. This information is important as incorrect sorting of our chromosomes during cellular proliferation, caused by defective packaging of the DNA, can lead to unwanted alterations in the genome and to disease such as cancer.

Sexually reproducing organisms rely on the process of meiosis for the generation of genetic diversity across generations. At the functional centre of meiosis is the process of genetic exchange between homologous chromosomes, of which the reaction of meiotic recombination is the physical manifestation. Despite its paramount role in meiosis, important aspects of meiotic recombination remain poorly understood at molecular level, including the specific architecture of the meiotic chromosome that must be achieved to promote genetic exchange, and the mechanism of the reactions of DNA-strand pairing and exchange that underlie recombination.

Our proposed research aims to improve our knowledge of meiotic recombination, by focusing on two important, related questions that remain poorly understood at structural and mechanistic level: what is the role of the synaptonemal complex (SC) proteins in the three-dimensional organisation of the meiotic chromosome, required for recombination? And what are the molecular principles of meiotic recombination that necessitate the intervention of a specific recombinase, DMC1, acting cooperatively with its mitotic counterpart, RAD51?

The current proposal builds on our long-standing interest in the field of homologous recombination and genomic stability, as well as our more recent focus on the structure and function of SC proteins in meiosis. To achieve our aim, we have designed a multi-disciplinary research plan that includes state-of-the-art biophysical and structural experiments, at ensemble and single-particle level. Our work will advance our understanding of the molecular mechanisms underpinning genetic recombination in meiosis and ultimately human fertility. Furthermore, it will illustrate molecular principles of DNA organisation that will be relevant to our understanding of chromosome structure and dynamics and of pathologies resulting from aneuploidy and incorrect chromosome segregation.

The correct execution of meiotic recombination is essential for successful completion of meiosis and hence for ensuring fertility. However, despite remaining the intense focus of research efforts at genetic, cellular and molecular level, our molecular understanding of key mechanisms of meiotic recombination remains incomplete. The successful completion of the experiments in this proposal will move forward in an important way our understanding of meiotic recombination at the molecular level, leading to the following academic, economic and societal impacts:

Academic impacts
The principal academic impacts of this work come from structural models of SC and recombination proteins and enzymes, and the identification of critical mechanisms governing meiotic chromosome organisation and recombination. Molecular models and testable mechanisms generated by our work will be invaluable to to elucidate aspects of meiotic recombination in vivo. In addition, the communities of interested researchers will benefit from the provision of biochemical reagents, including recombinant recombinant proteins and protein complexes that will advance the biochemical analysis of meiotic recombination. Our work will further benefit biotechnology companies that may develop small-molecule inhibitors of LE assembly or meiotic DNA-strand exchange, of commercial value in basic science, medical, agricultural and livestock industries. The large number of basic science and commercial beneficiaries within this country means that our work will have direct impact on basic science and biotechnology in the UK.

Economic and societal impacts - Medical and agricultural
The most immediate medical impact of our research will come from molecular models of the proteins under investigation, which for instance will enable defects in SC LE assembly or in execution of recombination to be predicted from genetic analysis. This will allow diagnosis of the molecular cause of infertility or recurrent miscarriage, and rational assessment of the risk of aneuploidy in pregnancy. Accurate diagnosis will better guide the choice of assisted reproduction method, tailoring it to individual couples and assisting the NHS in allocating resources to cases for which treatments are most likely to succeed. Fertility is an important concern of a large majority of the UK population, so such work is likely to have widespread impact on health and well-being, providing substantial economic benefits and with significant ability to attract R&D investment.
The methods developed for human fertility will likely first impact on the agricultural industry. Novel methods for controlling the fertility of livestock may increase the number of healthy animals born, reducing animal suffering, ensuring our food security and enhancing profitability. The latter is particularly applicable in the case of prize animals and crops, meaning that this would be likely to attract significant R&D investment.

Economic and societal impacts - Diversity and evolution
A precise understanding of genetic inheritance, through molecular understanding of crossover formation, will help us provide an accurate account of how the current wealth of genetic diversity has been attained. The explanation of our genetic history and the origin of diversity within our species is of particular interest to the general public and will raise awareness of the societal benefits stemming from our research.