Abstract
The promyelocytic leukemia protein (PML) participates in several cellular functions, including transcriptional regulation, apoptosis and maintenance of genomic stability. A key feature of this protein is its ability to induce the assembly of nuclear compartments termed PML-nuclear bodies (PML-NBs). Here we show that these nuclear structures recruit single-stranded DNA (ssDNA) molecules in response to exogenous DNA damage. ssDNA was readily detected in PML-NBs within 1 hour following exposure of cells to UV light. Confocal real-time imaging of cells expressing YFP-tagged PML did not reveal de novo formation of new PML-NBs following UV-irradiation, which shows that ssDNA focus formation occurred within pre-existing PML-NBs. Moreover, siRNA-mediated depletion of PML prevented ssDNA focus formation and sensitized cells to UV-induced apoptosis. PML-dependent ssDNA focus formation was found to be particularly efficient during S-phase of the cell cycle, and PML-depleted cells became retarded in S-phase upon growth in the presence of etoposide. In addition, we found that caffeine and the poly(ADP-ribose) polymerase (PARP) inhibitor NU1027 enhanced UV-induced recruitment of ssDNA to PML-NBs. Together, our results show that PML-NBs have the capacity to accommodate DNA metabolic activities that are associated with processing of damaged DNA.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Apoptosis / drug effects
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Apoptosis / radiation effects
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Caffeine / pharmacology
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Cell Nucleus Structures / genetics
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Cell Nucleus Structures / metabolism*
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Cells, Cultured
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Central Nervous System Stimulants / pharmacology
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DNA Damage / drug effects
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DNA Damage / radiation effects*
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DNA Repair*
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DNA, Neoplasm / genetics
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DNA, Neoplasm / metabolism*
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DNA, Single-Stranded / genetics
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DNA, Single-Stranded / metabolism*
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Endothelium, Vascular / drug effects
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Endothelium, Vascular / metabolism
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Endothelium, Vascular / radiation effects
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Humans
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Neoplasm Proteins / antagonists & inhibitors
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Neoplasm Proteins / genetics
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Neoplasm Proteins / metabolism*
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Nuclear Proteins / antagonists & inhibitors
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism*
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Promyelocytic Leukemia Protein
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RNA, Small Interfering / pharmacology
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S Phase / drug effects
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S Phase / radiation effects
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Transcription Factors / antagonists & inhibitors
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Transcription Factors / genetics
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Transcription Factors / metabolism*
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Tumor Suppressor Proteins / antagonists & inhibitors
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Tumor Suppressor Proteins / genetics
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Tumor Suppressor Proteins / metabolism*
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Ultraviolet Rays
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Umbilical Veins / drug effects
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Umbilical Veins / metabolism
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Umbilical Veins / radiation effects
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Zinc Fingers
Substances
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Central Nervous System Stimulants
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DNA, Neoplasm
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DNA, Single-Stranded
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Neoplasm Proteins
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Nuclear Proteins
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Promyelocytic Leukemia Protein
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RNA, Small Interfering
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Transcription Factors
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Tumor Suppressor Proteins
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PML protein, human
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Caffeine