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A novel repressor, par-4, modulates transcription and growth suppression functions of the Wilms' tumor suppressor WT1
R W Johnstone1, R H See, S F Sells
1Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Abstract:
The tumor suppressor WT1 represses and activates transcription. The loss and/or imbalance of the dual transcriptional activity of WT1 may contribute to Wilms' tumor. In this study, we identified par-4 (for prostate apoptosis response) as a WT1-interacting protein that itself functions as a transcriptional repressor. par-4 contains a putative leucine zipper domain and is specifically upregulated during apoptosis of prostate cells (S. F. Sells, D. P. Wood, Jr., S. S. Joshi-Barve, S. Muthukkumar, R. J. Jacob, S. A. Crist, S. Humphreys, and V. M. Rangnekar, Cell Growth Differ. 5:457-466, 1994). The leucine repeat domain of par-4 was shown to interact with the zinc finger DNA binding domain of WT1. Immunoprecipitation-Western blot (immunoblot) analyses demonstrated in vivo WT1-par-4 interactions. par-4 was ubiquitously expressed, and the protein was found in both the nucleus and the cytoplasm. Functionally, par-4 inhibited transcription activated by WT1, but not by the related protein EGR1. Inhibition of WT1-mediated transcription was dependent on the domain of par-4 that mediates its physical association with WT1. In addition, par-4 augmented WT1-mediated repression, possibly by contributing an additional repression domain. Consistent with these results, par-4 functioned as a transcriptional repressor when brought to a promoter via a heterologous DNA binding domain. Significantly, par-4, but not a mutant unable to interact with WT1, rescued growth suppression caused by WT1. Thus, we identified a novel repressor that modulates transcription as well as growth suppression functions of WT1.
Insights
Researchers discovered par-4, a protein that interacts with the tumor suppressor WT1. Par-4 modulates WT1
Area of Science:
- Molecular Biology
- Cancer Research
- Transcriptional Regulation
Background:
- The Wilms tumor suppressor (WT1) protein exhibits dual transcriptional activity (repression and activation).
- Dysregulation of WT1's transcriptional functions is implicated in Wilms tumor development.
- Prostate apoptosis response gene-4 (par-4) is a known transcriptional repressor upregulated during prostate cell apoptosis.
Purpose of the Study:
- To identify novel WT1-interacting proteins.
- To investigate the functional consequences of WT1-par-4 interaction on WT1's transcriptional and growth-suppressive activities.
Main Methods:
- Co-immunoprecipitation and Western blotting to confirm in vivo interaction between WT1 and par-4.
- Yeast two-hybrid assays to map the interaction domains (par-4 leucine repeat and WT1 zinc finger).
- Reporter gene assays to assess the effect of par-4 on WT1-mediated transcription.
- Growth suppression assays using WT1 and par-4 (wild-type and mutant).
Main Results:
- Par-4 directly interacts with WT1 through its leucine repeat domain and WT1's zinc finger DNA-binding domain.
- Par-4 inhibits WT1-activated transcription and enhances WT1-mediated repression.
- Par-4's transcriptional repressor activity is dependent on its interaction domain with WT1.
- Par-4, but not a non-interacting mutant, rescues WT1-induced growth suppression.
Conclusions:
- Par-4 is a novel WT1-interacting protein that modulates both the transcriptional and growth-suppressive functions of WT1.
- The interaction between par-4 and WT1 plays a significant role in regulating cellular processes relevant to cancer, particularly Wilms tumor.
- Par-4 represents a potential therapeutic target for cancers associated with WT1 dysregulation.