Plant-specific pecularities of the enzyme machinery of DNA recombination
Final Report Abstract
With the current project we were able to refine our understanding about proteins involved in meiotic recombination in plants. SPO11, a homolog of the subunit A of the archaebacterial topoisomerase VI, is essential for double-strand break (DSB)–induced initiation of meiotic recombination. In contrast with single homologs in animals and yeasts, three homologs are present in Arabidopsis thaliana and other higher plants. Whereas AtSPO11-3 is involved in somatic endoreduplication, AtSPO11-1 and AtSPO11-2 are essential for the initiation of meiotic recombination. Further defining the role of AtSPO11-2, we were able to demonstrate that it is required for proper chromosome segregation, as its loss resulted in aneuploidy in the surviving progeny. The double mutant spo11-1/spo11-2 does not differ phenotypically from the single mutants, indicating that both proteins are required for the same step. Contrary to the observations for the Atrad51-1 single mutant, the combination of spo11-2 and rad51-1 did not lead to chromosome fragmentation, indicating that SPO11-2, like SPO11-1, is required for DSB induction. As the meiotic phenotype of both single SPO11 mutants can be reversed by complementation using the full-length genes but not the same constructs mutated in their respective catalytically active Tyr, both proteins seem to participate directly in the DNA breakage reaction. The active involvement of two SPO11 homologs for DSB formation reveals a striking difference between plants and other eukaryotes in meiosis. Futhermore, we were able to demonstrate that Topoisomerase 3α (TOP3α) functions in the resolution step of homologous recombination intermediates. Using a hypomorphic insertion mutant of Arabidopsis thaliana (top3α-2), which is viable but completely sterile, we were able to define three different functions of the protein in mitosis and meiosis. AtTOP3α, together with AtRECQ4A and AtRMI1, is involved in the suppression of crossover recombination in somatic cells as well as DNA repair in both mammals and Arabidopsis thaliana. Surprisingly, AtTOP3α is also essential for meiosis. The phenotype of chromosome fragmentation, bridges and telophase I arrest can be suppressed by AtSPO11 and AtRAD51 mutations, indicating that the protein is required for the resolution of recombination intermediates. As Atrmi1 mutants have a similar meiotic phenotype as Attop3α mutants, both proteins seem to be involved in a mechanism safeguarding the entangling of homologous chromosomes during meiosis. The requirement of AtTOP3α and AtRMI1 in a late step of meiotic recombination strongly hints at the possibility that the dissolution of double Holliday Junctions via a hemicatenane intermediate is indeed an indispensable step of meiotic recombination, a finding highly relevant not only for plants but all eukaryotes.
Publications
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(2007) The catalytically active tyrosine residues of bothSPO11-1 and SPO11-2 are required for meiotic DSB induction in Arabidopsis. Plant Cell 19, 3090-3099
Hartung F., Wurz-Wildersinn R., Fuchs J., Schubert I., Suer S. and Puchta H.
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(2008) Topoisomerase 3A and RMI1 suppress somatic crossovers and are essential for resolution of meiotic recombination intermediates in Arabidopsis thaliana. PLoS Genetics 4 (12), 1-11
Hartung F., Suer S., Knoll A., Wurz-Wildersinn R. and Puchta H.
