Abstract
Trans-lesion synthesis (TLS) is an important DNA-damage tolerance mechanism that permits ongoing DNA synthesis in cells harbouring damaged genomes. The E3 ubiquitin ligase RAD18 activates TLS by promoting recruitment of Y-family DNA polymerases to sites of DNA-damage-induced replication fork stalling. Here we identify the cancer/testes antigen melanoma antigen-A4 (MAGE-A4) as a tumour cell-specific RAD18-binding partner and an activator of TLS. MAGE-A4 depletion from MAGE-A4-expressing cancer cells destabilizes RAD18. Conversely, ectopic expression of MAGE-A4 (in cell lines lacking endogenous MAGE-A4) promotes RAD18 stability. DNA-damage-induced mono-ubiquitination of the RAD18 substrate PCNA is attenuated by MAGE-A4 silencing. MAGE-A4-depleted cells fail to resume DNA synthesis normally following ultraviolet irradiation and accumulate γH2AX, thereby recapitulating major hallmarks of TLS deficiency. Taken together, these results demonstrate a mechanism by which reprogramming of ubiquitin signalling in cancer cells can influence DNA damage tolerance and probably contribute to an altered genomic landscape.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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A549 Cells
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Animals
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Antigens, Neoplasm / genetics*
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Antigens, Neoplasm / metabolism
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Cell Line
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Cell Line, Tumor
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Cloning, Molecular
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DNA Damage
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DNA Repair*
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DNA Replication
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DNA-Binding Proteins / genetics*
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DNA-Binding Proteins / metabolism
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Escherichia coli / genetics
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Escherichia coli / metabolism
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Fibroblasts / cytology
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Fibroblasts / metabolism
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Fibroblasts / radiation effects
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Gene Expression Regulation, Neoplastic*
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Genetic Vectors / chemistry
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Genetic Vectors / metabolism
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HCT116 Cells
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HeLa Cells
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Histones / genetics*
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Histones / metabolism
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Humans
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Mice
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Neoplasm Proteins / genetics*
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Neoplasm Proteins / metabolism
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Signal Transduction
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Ubiquitin-Conjugating Enzymes / genetics
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Ubiquitin-Conjugating Enzymes / metabolism
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Ubiquitin-Protein Ligases / genetics*
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Ubiquitin-Protein Ligases / metabolism
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Ultraviolet Rays
Substances
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Antigens, Neoplasm
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DNA-Binding Proteins
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H2AX protein, human
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Histones
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MAGEA4 protein, human
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Neoplasm Proteins
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RAD18 protein, human
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Recombinant Proteins
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UBE2A protein, human
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Ubiquitin-Conjugating Enzymes
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Ubiquitin-Protein Ligases