The life and death of Rad52p.
Erin Asleson, Dennis
Livingston
Biochem.,Mol.Biol.& Biophysics, University of Minnesota,
321 Church St. SE, Minneapolis, MN 55455, USA
We have determined the
half-lives of theSaccharomyces cerevisiaeRad52 and Rad51 proteins
by placing an epitope-tagged copy of each gene behind the repressible
GAL1promoter. By this method, we have found the half-life of
Rad52p to be fifteen minutes, while that of Rad51p is more than two
hours. While many mutant Rad52 proteins are more labile than the wild-type protein, mutant proteins with alterations between residues 210 and
327 have elevated half-lives. Both internal deletions within this region
and a single amino acid change of residue 235 (Arg to Gly) double the
half-life of the protein, demonstrating that this region plays a role in
the turnover of the Rad52 protein. We have searched for factors that may
be involved in the regulated turnover of Rad52p and have found that
several obvious candidates do not affect its half-life. These include
high level expression of RAD51, RFA2,and UBC9,as well as
mutations in the proteasome. Proteasomal mutations specifically affect
the steady state level of RAD52mRNA, but not the half-life of the
mRNA suggesting that the proteasome may act on a positive
RAD52transcription factor. These studies indicate that the
quantity and the longevity of Rad52p are controlled by both
transcriptional and post-translational regulation. This regulation may
have implications for the dynamic rearrangement of recombinational
repair complexes that contain Rad52p.
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