PARP-1 Determines Specificity in a Retinoid Signaling Pathway via Direct Modulation of Mediator

Department of Biochemistry, Howard Hughes Medical Institute, University of Medicine and Dentistry of New Jersey, 683 Hoes Lane, Piscataway, New Jersey 08854, USA.
Molecular Cell (Impact Factor: 14.46). 05/2005; 18(1):83-96. DOI: 10.1016/j.molcel.2005.02.034
Source: PubMed

ABSTRACT We show that PARP-1 is indispensable to retinoic acid receptor (RAR)-mediated transcription from the RARbeta2 promoter in a highly purified, reconstituted transcription system and that RA-inducible expression of all RARbeta isoforms is abrogated in PARP-1(-/-) cells in vivo. Importantly, PARP-1 activity was independent of its catalytic domain. PARP-1 directly interacts with RAR and Mediator. Chromatin immunoprecipitation experiments confirmed the presence of PARP-1 and Mediator on RAR-responsive promoters in vivo. Importantly, Mediator was inactive (Cdk8+) under basal conditions but was activated (Cdk8-) upon induction. However, in PARP-1(-/-) cells, Mediator was retained in its inactive state (Cdk8+) upon induction consistent with the absence of gene expression. PARP-1 became dispensable for ligand-dependent transcription in a chromatin reconstituted transcription assay when Mediator was devoid of the Cdk8 module (CRSP). PARP-1 appears to function as a specificity factor regulating the RA-induced switch of Mediator from the inactive (Cdk8+) to the active (Cdk8-) state in RAR-dependent transcription.

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    • "Thus, a mutual allosteric block appears to contribute to pol II–CDK8 module antagonism. Although definitive confirmation in cells is practically and technically difficult, correlations have emerged that suggest mutually exclusive CDK8 module versus pol II occupancy at certain welltested , inducible genes (Kim et al., 2006b; Mo et al., 2004; Pavri et al., 2005). As described later, this CDK8 module–pol II antagonism for binding Mediator may represent a key regulatory checkpoint. "
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    ABSTRACT: Abstract The Mediator complex is a multi-subunit assembly that appears to be required for regulating expression of most RNA polymerase II (pol II) transcripts, which include protein-coding and most non-coding RNA genes. Mediator and pol II function within the pre-initiation complex (PIC), which consists of Mediator, pol II, TFIIA, TFIIB, TFIID, TFIIE, TFIIF and TFIIH and is approximately 4.0 MDa in size. Mediator serves as a central scaffold within the PIC and helps regulate pol II activity in ways that remain poorly understood. Mediator is also generally targeted by sequence-specific, DNA-binding transcription factors (TFs) that work to control gene expression programs in response to developmental or environmental cues. At a basic level, Mediator functions by relaying signals from TFs directly to the pol II enzyme, thereby facilitating TF-dependent regulation of gene expression. Thus, Mediator is essential for converting biological inputs (communicated by TFs) to physiological responses (via changes in gene expression). In this review, we summarize an expansive body of research on the Mediator complex, with an emphasis on yeast and mammalian complexes. We focus on the basics that underlie Mediator function, such as its structure and subunit composition, and describe its broad regulatory influence on gene expression, ranging from chromatin architecture to transcription initiation and elongation, to mRNA processing. We also describe factors that influence Mediator structure and activity, including TFs, non-coding RNAs and the CDK8 module.
    Critical Reviews in Biochemistry and Molecular Biology 10/2013; 48(6). DOI:10.3109/10409238.2013.840259 · 5.81 Impact Factor
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    • "Transcriptional regulation often appears to involve a modulation of chromatin structure via poly(ADP-ribose) synthesis as well [25]. Yet, the enzyme can also act more directly as a co-regulator, which would involve an association with intact DNA, but not necessarily its catalytic activity [31]. Particularly this latter aspect of PARP-1 function might be subject to modulation by SUMO, as suggested by the finding that the modifier interferes with the enzyme's function as a co-activator of HIF1-␣ [32]. "
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    ABSTRACT: Poly(ADP-ribose) polymerase 1 (PARP-1) plays an important role in DNA repair, but also contributes to other aspects of nucleic acid metabolism, such as transcriptional regulation. Modification of PARP-1 with the small ubiquitin-related modifier (SUMO) affects its function as a transcriptional co-activator of hypoxia-responsive genes and promotes induction of the heat shock-induced HSP70.1 promoter. We now report that PARP-1 sumoylation is strongly influenced by DNA. Consistent with a function in transcription, we show that sumoylation in vitro is enhanced by binding to intact, but not to damaged DNA, in a manner clearly distinct from the mechanism by which DNA damage stimulates PARP-1's catalytic activity. An enhanced affinity of PARP-1 for the SUMO-conjugating enzyme Ubc9 upon binding to DNA is likely responsible for this effect. Sumoylation does not interfere with the catalytic or DNA-binding properties of PARP-1, and structural analysis reveals no significant impact of SUMO on the conformation of PARP-1's DNA-binding domain. In vivo, sumoylated PARP-1 is associated with chromatin, but the modification is not responsive to DNA damage and is not affected by PARP-1 catalytic activity. Our results suggest that PARP-1's alternative modes of DNA recognition serve as a means to differentiate between distinct aspects of the enzyme's function.
    DNA repair 07/2013; 12(9). DOI:10.1016/j.dnarep.2013.07.001 · 3.36 Impact Factor
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    • "Of note, the CDK8 module contains four subunits (CDK8, CCNC, MED12, and MED13) (Taatjes 2010) and associates with Mediator through contacts between MED13 and the Mediator core (Knuesel et al. 2009). Our present understanding of the mechanisms that regulate CDK8 module–Mediator interaction is limited to a study demonstrating that PARP-1 is required in a catalytically independent manner for CDK8 module–Mediator dissociation in the specific context of retinoic acid-regulated gene expression (Pavri et al. 2005). However, the general mechanisms that determine the amount of CDK8 module bound to Mediator are largely undefined. "
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    ABSTRACT: The Mediator complex is an essential transcription regulator that bridges transcription factors with RNA polymerase II. This interaction is controlled by dynamic interactions between Mediator and the CDK8 module, but the mechanisms governing CDK8 module-Mediator association remain poorly understood. We show that Fbw7, a tumor suppressor and ubiquitin ligase, binds to CDK8-Mediator and targets MED13/13L for degradation. MED13/13L physically link the CDK8 module to Mediator, and Fbw7 loss increases CDK8 module-Mediator association. Our work reveals a novel mechanism regulating CDK8 module-Mediator association and suggests an expanded role for Fbw7 in transcriptional control and an unanticipated relationship with the CDK8 oncogene.
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