A dissection of the molecular mechanisms behind DNA double strand
break repair during meiosis.
Tanja Thybo (1), Michael Lisby (2), Rodney Rothstein (2), Uffe
Mortensen (1)
(1) BioCentrum-DTU, Tech. University of Denmark, Building 223, Lyngby,
DK-2800, Denmark;
(2) Department of Genetics and Development, Columbia University, USA
Rad52 is a key component in DNA double strand break (DSB) repair,
homologous recombination and gene targeting. Thus, deletion of
RAD52 causes severe defects in these processes (1). To analyze
recombination in vivo in S. Cerevisiae the yellow
fluorescent protein (YFP) has been fused to RAD52. Thus, the
subcellular localization of Rad52 can be visualized. Induction of DSB's
in the genome relocalizes the diffuse nuclear distribution of Rad52-YFP
to distinct nuclear foci that reflects active repair of DNA lesions by
recombination (2). During meiosis several DSB's are induced to ensure
proper chromosome pairing and segregation. The repair of these DSB's can
be followed visually by the appearance and disappearance of distinct
foci in the nucleus. Hence, the molecular requirements of DNA DSB repair
in meiosis can be investigated by studying the fate of Rad52-YFP foci in
strains that contain relevant mutations. For this purpose, various genes
involved in homologous recombination have been deleted. Furthermore, a
collection of rad52 mutant strains has been created which can be
used to further dissect the mechanism of recombination (3). The meiotic
consequences of these mutants are under investigation. Interesting
mutant Rad52 species will be tagged with YFP and the impact of the
mutations on the in vivo localization of Rad52 will be assessed.
1. Paques et al., 1999., 2. Lisby, M., et al., 2001., 3. Mortensen,
U.H., et al., 2002.
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