Unravelling the meiotic single-cell transcriptomic atlas for the control of recombination.

Most organisms that reproduce sexually use a special type of cell division, called meiosis, that is important for the creation of genetic variation and halving the chromosome numbers in gametes. During meiosis, numerous programmed DNA double strand-breaks (DSBs) are formed and processed by the meiotic recombination pathway to form crossovers, which are the points of reciprocal exchange of genetic information between chromosomes. Crossovers are essential to create novel genetic variation in each generation. There is a major interest to understand meiosis in plants because domestication and intense selective breeding have led to a substantial loss of genetic variation in crops. Continued genetic improvement of elite cultivars to mitigate the challenge of climate change will require the introgression of beneficial alleles from wild varieties through the formation of crossovers. Unfortunately, crossovers are mainly formed at the end of the chromosomes, representing less than 15% of the genome, whereas more centric regions, which contain certain genes of agricultural relevance, like defence response genes, rarely recombine in most major crops. Therefore, it is both timely and imperative to understand the factors influencing crossover patterning and to create strategies to reposition crossovers in crops.
Substantial cellular variation in DSB and crossover numbers is observed between species and within individuals. Arabidopsis and wheat anthers contain a mixture of hypo- and hyper-recombinant meiotic cells varying in DSB and crossover numbers by up to 70%. Our previous studies revealed that the frequency and position of the crossovers are influenced by the transcript levels of ASY1 and HEI10 in Arabidopsis. Therefore, we propose that the recombination outcome of a meiocyte is influenced by a fine balance of expression of several genes. Hence, heterogeneity in the transcriptome could be responsible for the hypo- and hyper-recombination meiocytes observed in anthers. However, all genomic studies carried out on plant meiosis have so far included pools of cells, thus preventing the identification of heterogeneous factors responsible for such variation.
In this project, we propose to generate a single cell transcriptomic atlas of Arabidopsis meiocytes at two key time points of meiotic recombination (T1 during DSB formation, T2 during crossover formation) to understand the transcriptome dynamics from the formation of DSBs to their conversion into crossovers. In addition, we will group cells that are transcriptionally highly correlated and infer the cluster of cells that contains the hyper-recombinant meiocytes using information from known genes (e.g. higher HEI10 transcript level corresponds to higher crossover rate). We will then use this data to identify genes with a putative role in recombination heterogeneity. We will complement this study with the characterisation of a set of Arabidopsis over-expressing lines to find genes influencing recombination based on their transcript level. Lastly, we will perform a proof-of-principle experiment, using the dosage-sensitive gene ASY1 as a reference, to test if increasing meiotic gene expression in wheat could reposition crossovers to favour recombination in regions which are not easily accessible in conventional breeding. These new data will provide impact through the use of innovative approaches to understand the inter-relationship between transcriptome and recombination heterogeneity, decipher the transcriptome dynamics during meiosis and discover genes involved in meiosis. This project also aims to explore a novel route for impact in wheat using gene over-expression to influence the recombination landscape, which could confer lasting benefits for the breading sector. This proposed work supports BBSRC strategic priorities "Frontier bioscience: understanding the rules of life" and "Bioscience for sustainable agriculture and food".

Meiosis is a specialised type of cell division important for the creation of genetic variation and halving the chromosome numbers in gametes. During meiosis, numerous programmed DNA double strand breaks (DSBs) are formed along the genome. However, only 2-5% of the DSBs in plants mature to form crossovers, which are the points of reciprocal exchange of genetic information between chromosomes. The genetic improvement of elite lines depends on the formation of crossovers. Unfortunately, crossovers are mostly formed at chromosomes ends while over 85% of the genome rarely recombines in most crops. We previously showed that the anthers contain a mixture of hypo- and hyper-recombinant meiotic cells varying in their number of recombination events by up to 70%. We also found three genes influencing the recombination frequency based on their expression level. We propose that transcriptome heterogeneity is responsible for recombination heterogeneity and seek to comprehensively understand the inter-relationship between transcriptome dynamics and the progression of meiotic recombination. We propose to generate the first single cell transcriptomic atlas of plant meiocytes from DSB formation (1st time point) to their conversion into crossovers (2nd time point) using Arabidopsis. We aim to uncover the transcriptome heterogeneity between cells and relate this with recombination heterogeneity to gain knowledge on the transcriptome of the hyper-recombinant meiocytes. We will then characterise a set of Arabidopsis over-expressing lines to discover genes influencing recombination based on their transcript level. This objective has potential for gene discovery to be translated to crops. Lastly, we will perform a proof-of principle experiment to test if over-expressing ASY1, a known dosage-sensitive gene, could reposition wheat crossovers. This project aligns with BBSRC remit "Frontier bioscience: understanding the rules of life" with impact in academic and industrial sectors.