Group Leader

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Dr. Bernd Reiß
Group Leader
Phone:+49 221 5062 220Fax:+49 221 5062 113
Email:reiss@...

Recombination Research (Bernd Reiss)

Recombination Research

Research Program

Figure 1: A. thaliana, a typical flowering plant. Zoom Image
Figure 1: A. thaliana, a typical flowering plant.

Studies of the same process in different organisms reveal important basic features as well as adaptations to special functions. We focused on two model plants greatly differing in phylogenetic position, the flowering plant Arabidopsis thaliana (Fig. 1) and the moss Physcomitrella patens (Fig. 2). While A. thaliana is a typical flowering plant with a complex body plan and elaborated developmental patterns, the moss P. patens is a lower plant with comparably simple patterns. The development of P. patens begins with a meiospore that grows out to a haploid, filamentous structure, the protonema which is characterised by one-dimensional growth. Three-dimensional structures develop later giving rise to the adult gametophyte, the moss plant with leafy shoots. These shoots produce male and female reproductive organs fusion of which gives rise to the diploid generation, the sporophyte that eventually
produces haploid spores by meiosis.

Figure 2: The moss P. patens. Leafy shoots develop on a one-dimensional structure, the haploid protonema. Zoom Image
Figure 2: The moss P. patens. Leafy shoots develop on a one-dimensional structure, the haploid protonema.

We are interested in plant-specific aspects of DNA recombination. A. thaliana and P. patens differ fundamentally in the efficiency of gene targeting, a process generally considered to reflect the use of homologous recombination (HR) over the non-homologous end-joining pathway (NHEJ) in somatic cells and thus a preference for error-free over error-prone DNA damage repair.

Figure 3: Empty and well developed stamens on a young wild-type and atrad51 mutant A. thaliana flower. Zoom Image
Figure 3: Empty and well developed stamens on a young wild-type and atrad51 mutant A. thaliana flower.

DNA damage repair is involved in the maintenance of genome instability and in humans errors in this process are associated with cancer. NHEJ is considered to contribute to genome instability while HR leads to precise DNA damage repair and thus contributes to the prevention of cancer. RAD51, a homologue of the bacterial recA gene, plays a key role in eukaryotic HR and is essential for DNA damage repair and meiosis in yeast. In vertebrates like human, mouse and chicken, RAD51 seems to have acquired an additional function at the interface of DNA damage repair and cell cycle control. To analyse this process in plants with a special attention to the additional function, mutants were produced that lost RAD51function in both A. thaliana and P. patens. In contrast to animals, these mutants are fully viable indicating that the role of RAD51 in plants is restricted to recombination functions. This finding is typical for plants since this feature is conserved across a large phylogenetic distance and independent of the use of DNA repair pathways. In addition, these data imply that plants and animals differ in important aspects in the link of DNA damage repair to cell cycle control.

Figure 4: Flower and silique development in wild-type (left) and atrad51 (right) A. thalianaplants. Zoom Image
Figure 4: Flower and silique development in wild-type (left) and atrad51 (right) A. thalianaplants.

In A. thaliana the main function of RAD51 and by implication the homologous pathway of recombination appears to be in meiosis and homozygous mutants are male and female sterile. This phenotype becomes apparent early after the onset of flowering and is manifested in empty stamens visible in opening flowers (Fig. 3) and aborted siliques and ovule (Fig. 4) development at later stages of development, respectively. A more detailed analysis of meiosis by the group of Prof. Dr. Hong Ma (The Pennsylvania State University, University Park, PA 16802, USA) revealed the function of RAD51 in meiosis. The chromosomes fail to synapse in prophase I in the mutant and become extensively fragmented. Chromosome fragmentation is suppressed by atspo11-1, indicating that AtRAD51 functions downstream of AtSPO11-1. Therefore, AtRAD51 likely plays a crucial role in the repair of DNA double-stranded breaks generated by AtSPO11-1. These results suggest that RAD51 function is essential for chromosome pairing and synapsis at early stages in meiosis in A. thaliana. Furthermore, major aspects of meiotic recombination seem to be conserved between yeast and plants, especially the fact that chromosome pairing and synapsis are dependent on the function of SPO11 and RAD51. The function of RAD51 and its role in development has immediate relevance for the development of cancer in humans and therefore the A. thaliana RAD51 project has a direct link to cancer research.

A major application of recombination research is gene targeting. The naturally high efficiency of gene targeting in Physcomitrella patens allows the direct application of this technology in reverse genetics. To exemplify the application of this technology we have started to analyse the function of the CONSTANS (CO) gene in Physcomitrella patens. The transition from vegetative growth to flowering in A. thaliana is under environmental control. The CO gene plays a central role in the pathway that promotes flowering specifically in response to long days. The CO pathway is highly conserved among the angiosperms and some of these genes complement the mutation suggesting functional conservation. In is part of a gene family with 17 members of CO and CO-like genes. Although some of these CO-like proteins are highly similar to CO, they have no apparent function in flowering time regulation.

Database searches identified only three genes that had all of the hallmarks of CO. These are most similar to CO-like genes AtCOL3/AtCOL4/AtCOL5, a group of A. thaliana genes closely related to, but distinct from CO, suggesting that the CO branch of the AtCOL phylogeny does not exist in the P. patens genome. The data also indicate that CO-like genes must have existed in the common ancestor of bryophytes and flowering plants, and that CO originated in the group of CO-like genes represented by AtCOL3/AtCOL4/AtCOL5. Furthermore, expression of the three closely related Physcomitrella homologues is regulated by light, suggesting that the role of CO in flowering time control was probably derived from an ancestral function in light-signal transduction. A functional analysis of these genes is in progress.

 
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