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Sequestration and inhibition of Daxx-mediated transcriptional repression by PML
1Departments of Pharmacology and Molecular Toxicology and Cell Biology, Cancer Center, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA. don.chen@umassmed.edu
Abstract:
PML fuses with retinoic acid receptor alpha (RARalpha) in the t(15;17) translocation that causes acute promyelocytic leukemia (APL). In addition to localizing diffusely throughout the nucleoplasm, PML mainly resides in discrete nuclear structures known as PML oncogenic domains (PODs), which are disrupted in APL and spinocellular ataxia cells. We isolated the Fas-binding protein Daxx as a PML-interacting protein in a yeast two-hybrid screen. Biochemical and immunofluorescence analyses reveal that Daxx is a nuclear protein that interacts and colocalizes with PML in the PODs. Reporter gene assay shows that Daxx drastically represses basal transcription, likely by recruiting histone deacetylases. PML, but not its oncogenic fusion PML-RARalpha, inhibits the repressor function of Daxx. In addition, SUMO-1 modification of PML is required for sequestration of Daxx to the PODs and for efficient inhibition of Daxx-mediated transcriptional repression. Consistently, Daxx is found at condensed chromatin in cells that lack PML. These data suggest that Daxx is a novel nuclear protein bearing transcriptional repressor activity that may be regulated by interaction with PML.
Insights
The nuclear protein Daxx acts as a transcriptional repressor. PML protein sequesters Daxx into PML oncogenic domains (PODs), inhibiting its repressor function, a process crucial for regulating gene expression.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Acute promyelocytic leukemia (APL) is linked to the t(15;17) translocation, involving PML and retinoic acid receptor alpha (RARalpha).
- PML protein localizes to nuclear structures called PML oncogenic domains (PODs), which are altered in APL.
Purpose of the Study:
- To identify proteins interacting with PML.
- To investigate the function of Daxx and its interaction with PML in transcriptional regulation.
Main Methods:
- Yeast two-hybrid screening to identify PML-interacting proteins.
- Biochemical assays and immunofluorescence to study Daxx localization and interaction with PML.
- Reporter gene assays to assess transcriptional activity.
Main Results:
- Daxx, a Fas-binding protein, interacts and colocalizes with PML in PODs.
- Daxx functions as a transcriptional repressor, likely by recruiting histone deacetylases.
- PML, but not the PML-RARalpha fusion, inhibits Daxx's repressor activity.
- SUMO-1 modification of PML is essential for Daxx sequestration into PODs and inhibition of Daxx-mediated repression.
Conclusions:
- Daxx is a novel nuclear protein with transcriptional repressor activity.
- PML regulates Daxx activity through interaction and sequestration within PODs.
- This interaction is dependent on PML's SUMO-1 modification and plays a role in gene expression regulation.