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Identification of protein complexes with quantitative proteomics in S. cerevisiae
Published on: March 4, 2009
Identification and characterization of SMARCAL1 protein complexes
Rémy Bétous1, Gloria G Glick, Runxiang Zhao
1Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.
Plos One
|May 15, 2013
Summary
SMARCAL1, a DNA repair enzyme, forms complexes with WRN helicase and Replication Protein A (RPA) at stalled replication forks. These proteins independently prevent DNA cleavage and promote fork restart, suggesting distinct roles in replication stress response.
Area of Science:
- Molecular Biology
- DNA Replication and Repair
- Biochemistry
Background:
- SMARCAL1 is an SNF2 family ATPase crucial for replication fork stability and restart.
- It translocates on DNA, catalyzing strand annealing, branch migration, and fork regression.
- Many SNF2 enzymes function within larger protein complexes.
Purpose of the Study:
- To identify proteins interacting with SMARCAL1.
- To elucidate SMARCAL1's role in the replication stress response.
- To understand the functional relationship between SMARCAL1, WRN, and RPA.
Main Methods:
- Proteomics approach to identify SMARCAL1 interacting proteins.
- Co-localization studies at stalled replication forks.
- Biochemical assays for DNA strand annealing and fork regression.
Main Results:
- SMARCAL1 forms complexes with DNA-PKcs and WRN helicase, in addition to RPA.
- SMARCAL1 and WRN co-localize at stalled forks, with RPA mediating the SMARCAL1-WRN interaction.
- Both SMARCAL1 and WRN independently prevent MUS81 cleavage of stalled forks, with SMARCAL1 showing more efficient fork regression activity.
Conclusions:
- RPA scaffolds SMARCAL1 and WRN at stalled replication forks.
- SMARCAL1 and WRN function independently to protect forks from cleavage and promote restart.
- These findings suggest distinct but cooperative pathways for replication fork repair and restart involving SMARCAL1 and WRN.
