scholarly journals PPARβ/δ controls Mediator recruitment and transcription initiation

2019 ◽  
Author(s):  
Nathalie Legrand ◽  
Clemens L. Bretscher ◽  
Svenja Zielke ◽  
Bernhard Wilke ◽  
Michael Daude ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Repression ◽  
Transcription Initiation ◽  
Target Genes ◽  
Hdac Inhibitors ◽  
Inverse Agonist ◽  
Inverse Agonists ◽  
A Subunit ◽  
Basal Transcription Factors

AbstractRepression of transcription by nuclear receptors involves NCOR and SMRT corepressor complexes, which harbour the deacetylase HDAC3 as a subunit. Both deacetylase-dependent and -independent repression mechanisms have been reported for these complexes. In the absence of ligands, the nuclear receptor PPARβ/δ recruits NCOR and SMRT and represses expression of its canonical targets including the ANGPTL4 gene. Agonistic ligands cause corepressor dissociation and enable enhanced induction of transcription by coactivators. Vice versa, recently developed synthetic inverse agonists lead to augmented corepressor recruitment and repression that dominates over activating stimuli. Both basal repression of ANGPTL4 and reinforced repression elicited by inverse agonists are partially insensitive to HDAC inhibition. This raises the question of how PPARβ/δ represses transcription mechanistically.Here, we show that the PPARβ/δ inverse agonist PT-S264 impairs transcription initiation in human cells. Inverse agonist-bound PPARβ/δ interferes with recruitment of Mediator, RNA polymerase II, and TFIIB, but not with recruitment of other basal transcription factors, to the ANGPTL4 promoter. We identify NCOR as the main ligand-dependent interactor of PPARβ/δ in the presence of PT-S264. In PPARβ/δ knockout cells, reconstitution with PPARβ/δ mutants deficient in basal repression recruit less NCOR, SMRT, and HDAC3 to chromatin, concomitant with increased binding of RNA polymerase II. PT-S264 restores binding of NCOR, SMRT, and HDAC3, resulting in diminished polymerase II binding and transcriptional repression. In the presence of HDAC inhibitors, ligand-mediated repression of PPARβ/δ target genes is only partially relieved. Our findings corroborate deacetylase-dependent and -independent repressive functions of HDAC3-containing complexes. Deacetylase-independent repression mediated by binding of inverse agonists to PPARβ/δ involve NCOR/SMRT recruitment and interference with Mediator, TFIIB, and RNA polymerase II binding.

scholarly journals PPARβ/δ recruits NCOR and regulates transcription reinitiation of ANGPTL4

2019 ◽  
Vol 47 (18) ◽  
pp. 9573-9591 ◽  
Author(s):  
Nathalie Legrand ◽  
Clemens L Bretscher ◽  
Svenja Zielke ◽  
Bernhard Wilke ◽  
Michael Daude ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Nuclear Receptor ◽  
Transcription Initiation ◽  
Target Gene ◽  
Target Genes ◽  
Hdac Inhibition ◽  
Inverse Agonists ◽  
Transcription Reinitiation

Abstract In the absence of ligands, the nuclear receptor PPARβ/δ recruits the NCOR and SMRT corepressors, which form complexes with HDAC3, to canonical target genes. Agonistic ligands cause dissociation of corepressors and enable enhanced transcription. Vice versa, synthetic inverse agonists augment corepressor recruitment and repression. Both basal repression of the target gene ANGPTL4 and reinforced repression elicited by inverse agonists are partially insensitive to HDAC inhibition. This raises the question how PPARβ/δ represses transcription mechanistically. We show that the PPARβ/δ inverse agonist PT-S264 impairs transcription initiation by decreasing recruitment of activating Mediator subunits, RNA polymerase II, and TFIIB, but not of TFIIA, to the ANGPTL4 promoter. Mass spectrometry identifies NCOR as the main PT-S264-dependent interactor of PPARβ/δ. Reconstitution of knockout cells with PPARβ/δ mutants deficient in basal repression results in diminished recruitment of NCOR, SMRT, and HDAC3 to PPAR target genes, while occupancy by RNA polymerase II is increased. PT-S264 restores binding of NCOR, SMRT, and HDAC3 to the mutants, resulting in reduced polymerase II occupancy. Our findings corroborate deacetylase-dependent and -independent repressive functions of HDAC3-containing complexes, which act in parallel to downregulate transcription.

scholarly journals The Myc Transactivation Domain Promotes Global Phosphorylation of the RNA Polymerase II Carboxy-Terminal Domain Independently of Direct DNA Binding

2007 ◽  
Vol 27 (6) ◽  
pp. 2059-2073 ◽  
Author(s):  
Victoria H. Cowling ◽  
Michael D. Cole
Keyword(s):  
Dna Binding ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Target Genes ◽  
Dependent Manner ◽  
Transactivation Domain ◽  
Pol Ii ◽  
Terminal Domain ◽  
Ctd Phosphorylation

ABSTRACT Myc is a transcription factor which is dependent on its DNA binding domain for transcriptional regulation of target genes. Here, we report the surprising finding that Myc mutants devoid of direct DNA binding activity and Myc target gene regulation can rescue a substantial fraction of the growth defect in myc −/− fibroblasts. Expression of the Myc transactivation domain alone induces a transcription-independent elevation of the RNA polymerase II (Pol II) C-terminal domain (CTD) kinases cyclin-dependent kinase 7 (CDK7) and CDK9 and a global increase in CTD phosphorylation. The Myc transactivation domain binds to the transcription initiation sites of these promoters and stimulates TFIIH binding in an MBII-dependent manner. Expression of the Myc transactivation domain increases CDK mRNA cap methylation, polysome loading, and the rate of translation. We find that some traditional Myc transcriptional target genes are also regulated by this Myc-driven translation mechanism. We propose that Myc transactivation domain-driven RNA Pol II CTD phosphorylation has broad effects on both transcription and mRNA metabolism.

scholarly journals A spectrum of mechanisms for the assembly of the RNA polymerase II transcription preinitiation complex.

1995 ◽  
Vol 15 (2) ◽  
pp. 1049-1059 ◽  
Author(s):  
C P George ◽  
L M Lira-DeVito ◽  
S L Wampler ◽  
J T Kadonaga
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Selective Inhibition ◽  
Initiation Site ◽  
Tata Box ◽  
Preinitiation Complex ◽  
Basal Transcription ◽  
Tata Box Binding Protein ◽  
Basal Transcription Factors

To explore the diversity in the mechanisms of basal transcription by RNA polymerase II, we have employed a novel biochemical approach that involves perturbation of the transcription reaction with exogenously added TFIIB or TATA box-binding protein (TBP). Under these conditions, we observe promoter-selective inhibition of transcription by excess TFIIB or excess TBP. This inhibition occurs at the level of basal transcription, because it is observed with minimal promoters that comprise only the TATA box and initiation site sequences as well as with preparations of basal transcription factors that have been purified to greater than 90% homogeneity. In addition, the excess basal factors inhibit the assembly of a functional preinitiation complex but do not inhibit transcription initiation from preassembled preinitiation complexes. A study of several promoters revealed a reciprocal trend in the promoter specificity of inhibition by excess TFIIB versus that by excess TBP. At opposite ends of this spectrum, promoters are strongly inhibited by excess TFIIB but not excess TBP and vice versa. These results reveal the existence of a spectrum of mechanisms for preinitiation complex assembly at different promoters. The mechanistic preference appears to be specified by the aggregate of basal promoter elements rather than by an individual component, such as the TATA box or initiation site sequence. This spectrum provides a new parameter by which differences in the function of minimal class II promoters can be analyzed in the context of both basal and regulated transcription.

scholarly journals Structure of TFIIK for phosphorylation of CTD of RNA polymerase II

2021 ◽  
Vol 7 (15) ◽  
pp. eabd4420
Author(s):  
Trevor van Eeuwen ◽  
Tao Li ◽  
Hee Jong Kim ◽  
Jose J. Gorbea Colón ◽  
Mitchell I. Parker ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Activation Loop ◽  
Active Conformation ◽  
General Transcription Factor ◽  
Cryo Electron Microscopy ◽  
Proposed Model ◽  
Carboxyl Terminal ◽  
Catalytic Cleft

During transcription initiation, the general transcription factor TFIIH marks RNA polymerase II by phosphorylating Ser5 of the carboxyl-terminal domain (CTD) of Rpb1, which is followed by extensive modifications coupled to transcription elongation, mRNA processing, and histone dynamics. We have determined a 3.5-Å resolution cryo–electron microscopy (cryo-EM) structure of the TFIIH kinase module (TFIIK in yeast), which is composed of Kin28, Ccl1, and Tfb3, yeast homologs of CDK7, cyclin H, and MAT1, respectively. The carboxyl-terminal region of Tfb3 was lying at the edge of catalytic cleft of Kin28, where a conserved Tfb3 helix served to stabilize the activation loop in its active conformation. By combining the structure of TFIIK with the previous cryo-EM structure of the preinitiation complex, we extend the previously proposed model of the CTD path to the active site of TFIIK.

Crystal structure of Ssu72, an essential eukaryotic phosphatase specific for the C-terminal domain of RNA polymerase II, in complex with a transition state analogue

2011 ◽  
Vol 434 (3) ◽  
pp. 435-444 ◽  
Author(s):  
Yong Zhang ◽  
Mengmeng Zhang ◽  
Yan Zhang
Keyword(s):  
Rna Polymerase ◽  
Transition State ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Complex Structure ◽  
Protein Tyrosine Phosphatases ◽  
Cysteine Residue ◽  
Phosphoryl Transfer ◽  
Terminal Domain ◽  
Deep Groove

Reversible phosphorylation of the CTD (C-terminal domain) of the eukaryotic RNA polymerase II largest subunit represents a critical regulatory mechanism during the transcription cycle and mRNA processing. Ssu72 is an essential phosphatase conserved in eukaryotes that dephosphorylates phosphorylated Ser5 of the CTD heptapeptide. Its function is implicated in transcription initiation, elongation and termination, as well as RNA processing. In the present paper we report the high resolution X-ray crystal structures of Drosophila melanogaster Ssu72 phosphatase in the apo form and in complex with an inhibitor mimicking the transition state of phosphoryl transfer. Ssu72 facilitates dephosphorylation of the substrate through a phosphoryl-enzyme intermediate, as visualized in the complex structure of Ssu72 with the oxo-anion compound inhibitor vanadate at a 2.35 Å (1 Å=0.1 nm) resolution. The structure resembles the transition state of the phosphoryl transfer with vanadate exhibiting a trigonal bi-pyramidal geometry covalently bonded to the nucleophilic cysteine residue. Interestingly, the incorporation of oxo-anion compounds greatly stabilizes a flexible loop containing the general acid, as detected by an increase of melting temperature of Ssu72 detected by differential scanning fluorimetry. The Ssu72 structure exhibits a core fold with a similar topology to that of LMWPTPs [low-molecular-mass PTPs (protein tyrosine phosphatases)], but with an insertion of a unique ‘cap’ domain to shelter the active site from the solvent with a deep groove in between where the CTD substrates bind. Mutagenesis studies in this groove established the functional roles of five residues (Met17, Pro46, Asp51, Tyr77 and Met85) that are essential specifically for substrate recognition.

scholarly journals Nuclear distribution of transcription factors in relation to sites of transcription and RNA polymerase II

1997 ◽  
Vol 110 (15) ◽  
pp. 1781-1791 ◽  
Author(s):  
M.A. Grande ◽  
I. van der Kraan ◽  
L. de Jong ◽  
R. van Driel
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Glucocorticoid Receptors ◽  
Cultured Cells ◽  
Spatial Relationship ◽  
Immunofluorescence Labelling ◽  
Transcription Sites ◽  
Nuclear Domains

We have investigated the spatial relationship between sites containing newly synthesized RNA and domains containing proteins involved in transcription, such as RNA polymerase II and the transcription factors TFIIH, Oct1, BRG1, E2F-1 and glucocorticoid receptors, using dual immunofluorescence labelling followed by confocal microscopy on cultured cells. As expected, a high degree of colocalisation between the RNA polymerase II and sites containing newly synthesised RNA was observed. Like the newly synthesised RNA and the RNA polymerase II, we found that all the transcription factors that we studied are distributed more or less homogeneously throughout the nucleoplasm, occupying numerous small domains. In addition to these small domains, TFIIH was found concentrated in coiled bodies and Oct1 in a single large domain of about 1.5 microm in 30% of the cells in an asynchronous HeLa cell culture. Remarkably, we found little or no relationship between the spatial distribution of the glucocorticoid receptor, Oct1 and E2F-1 on the one hand and RNA polymerase II and transcription sites on the other hand. In contrast, a significant but incomplete overlap was observed between the spatial distributions of transcription sites and BRG1 and TFIIH. These results indicate that many of the transcription factor-rich nuclear domains are not actively involved in transcription. They may represent incomplete transcription initiation complexes, inhibitory complexes, or storage sites.

Sign in / Sign up

Export Citation Format

Share Document