Proteomic Analysis of Chromosome Axis-Associated Proteins from Brassica oleracea and Arabidopsis thaliana Provides Insights into Meiotic Recombination

Despite recent progress, the role(s) of the chromosome axes and synaptonemal complex (SC) in co-ordinating key meiotic processes, including the formation and distribution of crossovers (COs), are still not well-understood. As part of the MEIOsys consortium (http://www.meiosys.org/), we are using affinity-based proteomics to isolate axis- and SC-associated protein complexes from Arabidopsis thaliana and its close crop relative, Brassica oleracea, for in-solution analysis by mass spectrometry. This is a powerful technique with the potential to identify new candidate meiotic proteins and reveal the protein-protein interactions involved in key meiotic events. We have identified a large number of proteins from Brassica meiotic tissue which co-immunoprecipitate with the axis-associated protein, ASY1. Mutant analysis of the Arabidopsis equivalents of two of these, ASY3 and a PCH2-like protein, has confirmed a role in CO formation. A number of identified novel proteins are being screened for a meiotic role. Additionally, we were able to detect phospho-modified peptides of BoASY1, providing insights into the signalling events which occur during the early stages of recombination. Currently we are incorporating the ASY1 data into Predictive Protein Interaction networks as a first step towards developing a comprehensive model of meiotic recombination to inform the manipulation of COs in crop species, the ultimate aim of MEIOsys. An example network based on STRING (http://string-db.org/) has revealed a protein cluster associated with DNA replication which includes a DNA polymerase sub-unit. Mutant analysis has confirmed a meiotic role for this protein and we are currently investigating its possible involvement in CO formation.