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During replication stress, non-SMC element 5 (NSE5) is required for Smc5/6 protein complex functionality at stalled
Denise E Bustard1, Demis Menolfi, Kristian Jeppsson
1Department of Biochemistry and Molecular Biology, Southern Alberta Cancer Research Institute, University of Calgary, Calgary, Alberta T2N 4N1, Canada.
The Journal of Biological Chemistry
|February 4, 2012
Summary
Nse5 is crucial for the Smc5/6 complex's stability and function during replication stress. Mutations in NSE5 disrupt Smc5 sumoylation and lead to replisome dissociation and DNA repair defects.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- The Smc5/6 complex, a key player in chromosome structure, includes essential non-Smc elements (Nse1-6).
- Nse2/Mms21 acts as an E3 SUMO ligase, critical for Smc5 sumoylation, but the roles of other Nse proteins remain largely unknown.
Purpose of the Study:
- To investigate the function of Nse5 within the Smc5/6 complex, particularly its role in stability and fork function during replication stress.
- To elucidate the relationship between Smc5 sumoylation and the overall functionality of the Smc5/6 complex.
Main Methods:
- Characterization of two temperature-sensitive mutants, nse5-ts1 and nse5-ts2, in Saccharomyces cerevisiae.
- Analysis of Smc5 sumoylation levels in wild-type and mutant strains.
- Phenotypic analysis including assessment of replisome stability, homologous recombination intermediates, and hydroxyurea sensitivity.
Main Results:
- Both nse5 mutant alleles significantly reduced Smc5 sumoylation, even below levels seen in a catalytically inactive Mms21 mutant.
- A phenotypic distinction emerged: nse5-ts1 cells, but not nse5-ts2, showed replisome dissociation, homologous recombination intermediates, and additive hydroxyurea sensitivity with mms21-11.
- These defects in nse5-ts1 are linked to impaired Smc5/6 complex localization to replication forks.
Conclusions:
- Nse5 is essential for Smc5/6 complex stability and localization to replication forks during replication stress.
- While Nse5 influences Smc5 sumoylation, its role extends beyond this modification, impacting critical interactions within the complex and its overall function.
- These findings highlight Nse5's vital contribution to maintaining genome integrity under stress conditions.
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