Related Experiment Video
Updated: Apr 9, 2026
Two- and Three-Dimensional Live Cell Imaging of DNA Damage Response Proteins
Published on: September 28, 2012
DNA Damage Regulates Translation through β-TRCP Targeting of CReP
Theresa B Loveless1, Benjamin R Topacio1, Ajay A Vashisht2
1Department of Biochemistry and Biophysics, Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, San Francisco, California, United States of America.
Abstract:
The Skp1-Cul1-F box complex (SCF) associates with any one of a number of F box proteins, which serve as substrate binding adaptors. The human F box protein βTRCP directs the conjugation of ubiquitin to a variety of substrate proteins, leading to the destruction of the substrate by the proteasome. To identify βTRCP substrates, we employed a recently-developed technique, called Ligase Trapping, wherein a ubiquitin ligase is fused to a ubiquitin-binding domain to "trap" ubiquitinated substrates. 88% of the candidate substrates that we examined were bona fide substrates, comprising twelve previously validated substrates, eleven new substrates and three false positives. One βTRCP substrate, CReP, is a Protein Phosphatase 1 (PP1) specificity subunit that targets the translation initiation factor eIF2α to promote the removal of a stress-induced inhibitory phosphorylation and increase cap-dependent translation. We found that CReP is targeted by βTRCP for degradation upon DNA damage. Using a stable CReP allele, we show that depletion of CReP is required for the full induction of eIF2α phosphorylation upon DNA damage, and contributes to keeping the levels of translation low as cells recover from DNA damage.
Insights
The Skp1-Cul1-F box (SCF) complex targets proteins for destruction. Researchers used Ligase Trapping to identify substrates, finding that CReP degradation is crucial for DNA damage response and translation regulation.
Area of Science:
- Cellular Biology
- Molecular Biology
- Biochemistry
Background:
- The Skp1-Cul1-F box (SCF) complex, a ubiquitin ligase, utilizes F-box proteins as adaptors to target substrates for proteasomal degradation.
- The human F-box protein β-transducin repeat-containing protein 1 (βTRCP) plays a critical role in degrading various substrate proteins, impacting cellular processes.
- Understanding βTRCP substrates is essential for elucidating its role in cellular signaling and stress responses.
Purpose of the Study:
- To identify novel substrates of the βTRCP ubiquitin ligase using an innovative technique.
- To investigate the role of the protein CReP (a PP1 specificity subunit) as a βTRCP substrate.
- To determine the functional significance of CReP degradation in the cellular response to DNA damage.
Main Methods:
- Utilized Ligase Trapping, a method combining a ubiquitin ligase with a ubiquitin-binding domain to capture ubiquitinated substrates.
- Screened candidate substrates for βTRCP-mediated ubiquitination and degradation.
- Employed a stable CReP allele to assess the impact of CReP depletion on DNA damage response pathways.
Main Results:
- Ligase Trapping identified 88% bona fide βTRCP substrates, including 11 new ones.
- Confirmed CReP as a βTRCP substrate, targeted for degradation upon DNA damage.
- Demonstrated that CReP depletion is necessary for full eIF2α phosphorylation induction and contributes to suppressed translation during DNA damage recovery.
Conclusions:
- βTRCP plays a significant role in regulating protein degradation, particularly in response to cellular stress.
- CReP is a key substrate of βTRCP, linking DNA damage to translational control.
- The degradation of CReP by βTRCP is essential for the proper induction of the integrated stress response and efficient recovery from DNA damage.
Related Concept Videos
02:36DNA Damage can Stall the Cell Cycle
02:36DNA Damage Can Stall the Cell Cycle
02:10Translesion DNA Polymerases
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
01:23Negative Regulator Molecules
01:38Nucleotide Excision Repair
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
01:08Nucleotide Excision Repair