BACKGROUND: DNA-protein cross-links (DPCs) are one of the most deleterious DNA lesions, originating from various sources, including enzymatic activity. For instance, topoisomerases, which play a fundamental role in DNA metabolic processes such as replication and transcription, can be trapped and remain covalently bound to DNA in the presence of poisons or nearby DNA damage. Given the complexity of individual DPCs, numerous repair pathways have been described. The protein tyrosyl-DNA phosphodiesterase 1 (Tdp1) has been demonstrated to be responsible for removing topoisomerase 1 (Top1). Nevertheless, studies in budding yeast have indicated that alternative pathways involving Mus81, a structure-specific DNA endonuclease, could also remove Top1 and other DPCs. RESULTS: This study shows that MUS81 can efficiently cleave various DNA substrates modified by fluorescein, streptavidin or proteolytically processed topoisomerase. Furthermore, the inability of MUS81 to cleave substrates bearing native TOP1 suggests that TOP1 must be either dislodged or partially degraded prior to MUS81 cleavage. We demonstrated that MUS81 could cleave a model DPC in nuclear extracts and that depletion of TDP1 in MUS81-KO cells induces sensitivity to the TOP1 poison camptothecin (CPT) and affects cell proliferation. This sensitivity is only partially suppressed by TOP1 depletion, indicating that other DPCs might require the MUS81 activity for cell proliferation. CONCLUSIONS: Our data indicate that MUS81 and TDP1 play independent roles in the repair of CPT-induced lesions, thus representing new therapeutic targets for cancer cell sensitisation in combination with TOP1 inhibitors.

• Keywords
• DNA-protein cross-links repair, MUS81, TDP1, Topoisomerase 1,
• MeSH
• DNA-Binding Proteins * genetics   metabolism   MeSH
• DNA Topoisomerases, Type I genetics   metabolism   MeSH
• Endonucleases * genetics   metabolism   MeSH
• Phosphoric Diester Hydrolases * genetics   metabolism   MeSH
• DNA Repair MeSH
• DNA Damage MeSH
• Saccharomyces cerevisiae Proteins * genetics   metabolism   MeSH
• Saccharomyces cerevisiae MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA-Binding Proteins * MeSH
• DNA Topoisomerases, Type I MeSH
• Endonucleases * MeSH
• Phosphoric Diester Hydrolases * MeSH
• MUS81 protein, S cerevisiae MeSH Browser
• Saccharomyces cerevisiae Proteins * MeSH
• Tdp1 protein, S cerevisiae MeSH Browser
• TOP1 protein, S cerevisiae MeSH Browser

Replication across damaged DNA templates is accompanied by transient formation of sister chromatid junctions (SCJs). Cells lacking Esc2, an adaptor protein containing no known enzymatic domains, are defective in the metabolism of these SCJs. However, how Esc2 is involved in the metabolism of SCJs remains elusive. Here we show interaction between Esc2 and a structure-specific endonuclease Mus81-Mms4 (the Mus81 complex), their involvement in the metabolism of SCJs, and the effects Esc2 has on the enzymatic activity of the Mus81 complex. We found that Esc2 specifically interacts with the Mus81 complex via its SUMO-like domains, stimulates enzymatic activity of the Mus81 complex in vitro, and is involved in the Mus81 complex-dependent resolution of SCJs in vivo Collectively, our data point to the possibility that the involvement of Esc2 in the metabolism of SCJs is, in part, via modulation of the activity of the Mus81 complex.

• MeSH
• Chromatids chemistry   metabolism   MeSH
• DNA, Fungal genetics   metabolism   MeSH
• DNA-Binding Proteins chemistry   genetics   metabolism   MeSH
• Endonucleases chemistry   genetics   metabolism   MeSH
• Escherichia coli genetics   metabolism   MeSH
• Nuclear Proteins chemistry   genetics   metabolism   MeSH
• Cloning, Molecular MeSH
• DNA, Cruciform chemistry   metabolism   MeSH
• Small Ubiquitin-Related Modifier Proteins chemistry   genetics   metabolism   MeSH
• Genomic Instability MeSH
• DNA Damage MeSH
• Protein Domains MeSH
• Cell Cycle Proteins MeSH
• Gene Expression Regulation, Fungal * MeSH
• Recombinant Proteins chemistry   genetics   metabolism   MeSH
• DNA Replication MeSH
• Saccharomyces cerevisiae Proteins chemistry   genetics   metabolism   MeSH
• Saccharomyces cerevisiae genetics   metabolism   MeSH
• Protein Binding MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• DNA, Fungal MeSH
• DNA-Binding Proteins MeSH
• Endonucleases MeSH
• Esc2 protein, S cerevisiae MeSH Browser
• Nuclear Proteins MeSH
• DNA, Cruciform MeSH
• Small Ubiquitin-Related Modifier Proteins MeSH
• MUS81 protein, S cerevisiae MeSH Browser
• Cell Cycle Proteins MeSH
• Recombinant Proteins MeSH
• Saccharomyces cerevisiae Proteins MeSH