scholarly journals Reconstitution of recombinant chromatin establishes a requirement for histone-tail modifications during chromatin assembly and transcription

2001 ◽  
Vol 15 (21) ◽  
pp. 2837-2851
Author(s):  
Alejandra Loyola ◽  
Gary LeRoy ◽  
Yuh-Hwa Wang ◽  
Danny Reinberg
Keyword(s):  
Rna Polymerase Ii ◽  
Histone H3 ◽  
Dependent Manner ◽  
Histone Tail ◽  
Histone Tails ◽  
Transcription System ◽  
Nuclear Extracts ◽  
Chromatin Acetylation ◽  
Rna Polymerase Ii Transcription ◽  
Nucleosome Arrays

The human ISWI-containing factor RSF (remodeling andspacing factor) was found to mediate nucleosome deposition and, in the presence of ATP, generate regularly spaced nucleosome arrays. Using this system, recombinant chromatin was reconstituted with bacterially produced histones. Acetylation of the histone tails was found to play an important role in establishing regularly spaced nucleosome arrays. Recombinant chromatin lacking histone acetylation was impaired in directing transcription. Histone-tail modifications were found to regulate transcription from the recombinant chromatin. Acetylation of the histone tails by p300 was found to increase transcription. Methylation of the histone H3 tail by Suv39H1 was found to repress transcription in an HP1-dependent manner. The effects of histone-tail modifications were observed in nuclear extracts. A highly reconstituted RNA polymerase II transcription system was refractory to the effect imposed by acetylation and methylation.

scholarly journals Activation of yeast polymerase II transcription by herpesvirus VP16 and GAL4 derivatives in vitro.

1989 ◽  
Vol 9 (11) ◽  
pp. 4746-4749 ◽  
Author(s):  
D I Chasman ◽  
J Leatherwood ◽  
M Carey ◽  
M Ptashne ◽  
R D Kornberg
Keyword(s):  
Rna Polymerase Ii ◽  
Fusion Proteins ◽  
In Vitro System ◽  
Transcription System ◽  
Natural Mechanism ◽  
Transcription In Vitro ◽  
Rna Polymerase Ii Transcription ◽  
Fold Stimulation

Fusion proteins known to activate transcription in vivo were tested for the ability to stimulate transcription in vitro in a recently developed Saccharomyces cerevisiae RNA polymerase II transcription system. One fusion protein, whose activation domain was derived from the herpesvirus transcriptional activator VP16, gave more than 100-fold stimulation in the in vitro system. The order of effects of the various proteins was the same for transcription in vitro and in vivo, suggesting that the natural mechanism of activation is preserved in vitro.

Pausing and termination of human RNA polymerase II transcription at a procaryotic terminator

1983 ◽  
Vol 3 (10) ◽  
pp. 1687-1693
Author(s):  
G W Hatfield ◽  
J A Sharp ◽  
M Rosenberg
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Sequence Specificity ◽  
Late Promoter ◽  
Transcription System ◽  
A Cell ◽  
Major Late Promoter ◽  
Dyad Symmetry ◽  
Rna Polymerase Ii Transcription ◽  
Terminator Sequence

Kinetic analyses of runoff transcription in a cell-free eucaryotic transcription system revealed that the bacteriophage lambda 4S RNA terminator caused human RNA polymerase II to pause on the template and partially terminate transcription of transcripts initiated by the adenovirus 2 major late promoter. Analogous to the procaryotic RNA polymerase, the eucaryotic enzyme terminated just beyond the guanine-plus-cytosine-rich region of dyad symmetry in the terminator sequence. These results suggest that the eucaryotic RNA polymerase II may respond to transcription termination sequences similar to those used by the procaryotic enzyme. However, similar templates containing lambda tint or lambda tR1 terminators did not elicit pausing or termination, suggesting that other features, such as sequence specificity, may also be involved.

scholarly journals Pausing and termination of human RNA polymerase II transcription at a procaryotic terminator.

1983 ◽  
Vol 3 (10) ◽  
pp. 1687-1693 ◽  
Author(s):  
G W Hatfield ◽  
J A Sharp ◽  
M Rosenberg
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Sequence Specificity ◽  
Late Promoter ◽  
Transcription System ◽  
A Cell ◽  
Major Late Promoter ◽  
Dyad Symmetry ◽  
Rna Polymerase Ii Transcription ◽  
Terminator Sequence

Kinetic analyses of runoff transcription in a cell-free eucaryotic transcription system revealed that the bacteriophage lambda 4S RNA terminator caused human RNA polymerase II to pause on the template and partially terminate transcription of transcripts initiated by the adenovirus 2 major late promoter. Analogous to the procaryotic RNA polymerase, the eucaryotic enzyme terminated just beyond the guanine-plus-cytosine-rich region of dyad symmetry in the terminator sequence. These results suggest that the eucaryotic RNA polymerase II may respond to transcription termination sequences similar to those used by the procaryotic enzyme. However, similar templates containing lambda tint or lambda tR1 terminators did not elicit pausing or termination, suggesting that other features, such as sequence specificity, may also be involved.

scholarly journals Histone H3 Variant Regulates RNA Polymerase II Transcription Termination and Dual Strand Transcription of siRNA Loci in Trypanosoma brucei

PLoS Genetics ◽  
2016 ◽  
Vol 12 (1) ◽  
pp. e1005758 ◽  
Author(s):  
David Reynolds ◽  
Brigitte T. Hofmeister ◽  
Laura Cliffe ◽  
Magdy Alabady ◽  
T. Nicolai Siegel ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Trypanosoma Brucei ◽  
Transcription Termination ◽  
Histone H3 ◽  
Histone H3 Variant ◽  
Rna Polymerase Ii Transcription

scholarly journals Histone tails cooperate to control the breathing of genomic nucleosomes

2021 ◽  
Vol 17 (6) ◽  
pp. e1009013
Author(s):  
Jan Huertas ◽  
Hans Robert Schöler ◽  
Vlad Cojocaru
Keyword(s):  
Epigenetic Regulation ◽  
Genomic Dna ◽  
Histone H3 ◽  
Molecular Simulations ◽  
Dna Interaction ◽  
Histone Tail ◽  
Chemical Modifications ◽  
Histone Tails ◽  
Chromatin Dynamics ◽  
Nucleosome Dynamics

Genomic DNA is packaged in chromatin, a dynamic fiber variable in size and compaction. In chromatin, repeating nucleosome units wrap 145–147 DNA basepairs around histone proteins. Genetic and epigenetic regulation of genes relies on structural transitions in chromatin which are driven by intra- and inter-nucleosome dynamics and modulated by chemical modifications of the unstructured terminal tails of histones. Here we demonstrate how the interplay between histone H3 and H2A tails control ample nucleosome breathing motions. We monitored large openings of two genomic nucleosomes, and only moderate breathing of an engineered nucleosome in atomistic molecular simulations amounting to 24 μs. Transitions between open and closed nucleosome conformations were mediated by the displacement and changes in compaction of the two histone tails. These motions involved changes in the DNA interaction profiles of clusters of epigenetic regulatory aminoacids in the tails. Removing the histone tails resulted in a large increase of the amplitude of nucleosome breathing but did not change the sequence dependent pattern of the motions. Histone tail modulated nucleosome breathing is a key mechanism of chromatin dynamics with important implications for epigenetic regulation.

scholarly journals P54nrb Forms a Heterodimer with PSP1 That Localizes to Paraspeckles in an RNA-dependent Manner

2005 ◽  
Vol 16 (11) ◽  
pp. 5304-5315 ◽  
Author(s):  
Archa H. Fox ◽  
Charles S. Bond ◽  
Angus I. Lamond
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Sequence Similarity ◽  
Dependent Manner ◽  
Rna Pol Ii ◽  
Nuclear Processes ◽  
Rna Polymerase Ii Transcription

P54nrb is a protein implicated in multiple nuclear processes whose specific functions may correlate with its presence at different nuclear locations. Here we characterize paraspeckles, a subnuclear domain containing p54nrb and other RNA-binding proteins including PSP1, a protein with sequence similarity to p54nrb that acts as a marker for paraspeckles. We show that PSP1 interacts in vivo with a subset of the total cellular pool of p54nrb. We map the domain within PSP1 that is mediating this interaction and show it is required for the correct localization of PSP1 to paraspeckles. This interaction is necessary but not sufficient for paraspeckle targeting by PSP1, which also requires an RRM capable of RNA binding. Blocking the reinitiation of RNA Pol II transcription at the end of mitosis with DRB prevents paraspeckle formation, which recommences after removal of DRB, indicating that paraspeckle formation is dependent on RNA Polymerase II transcription. Thus paraspeckles are the sites where a subset of the total cellular pool of p54nrb is targeted in a RNA Polymerase II-dependent manner.

scholarly journals FACT is recruited to the +1 nucleosome of transcribed genes and spreads in a Chd1-dependent manner

2020 ◽  
Author(s):  
Célia Jeronimo ◽  
Andrew Angel ◽  
Christian Poitras ◽  
Pierre Collin ◽  
Jane Mellor ◽  
...  
Keyword(s):  
Mathematical Modeling ◽  
Rna Polymerase Ii ◽  
Molecular Mechanisms ◽  
List Type ◽  
Dependent Manner ◽  
Active Chromatin ◽  
Histone Tails ◽  
High Resolution Mapping ◽  
Resolution Mapping ◽  
Chromatin Integrity

The histone chaperone FACT occupies transcribed regions where it plays prominent roles in maintaining chromatin integrity and preserving epigenetic information. How it is targeted to transcribed regions, however, remains unclear. Proposed models for how FACT finds its way to transcriptionally active chromatin include docking on the RNA polymerase II (RNAPII) C-terminal domain (CTD), recruitment by elongation factors, recognition of modified histone tails and binding partially disassembled nucleosomes. Here, we systematically tested these and other scenarios in Saccharomyces cerevisiae and found that FACT binds transcribed chromatin, not RNAPII. Through a combination of experimental and mathematical modeling evidence, we propose that FACT recognizes the +1 nucleosome, as it is partially unwrapped by the engaging RNAPII, and spreads to downstream nucleosomes aided by the chromatin remodeler Chd1. Our work clarifies how FACT interacts with genes, suggests a processive mechanism for FACT function, and provides a framework to further dissect the molecular mechanisms of transcription-coupled histone chaperoning.HighlightsHigh-resolution mapping of FACT localization in yeastFACT binds partially unwrapped nucleosomes in transcribed genes, not RNAPIIFACT distribution along genes requires Chd1Processive mechanism for FACT function

scholarly journals p53 is phosphorylated by CDK7-cyclin H in a p36MAT1-dependent manner.

1997 ◽  
Vol 17 (12) ◽  
pp. 7220-7229 ◽  
Author(s):  
L J Ko ◽  
S Y Shieh ◽  
X Chen ◽  
L Jayaraman ◽  
K Tamai ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Rna Polymerase Ii ◽  
Excision Repair ◽  
Dependent Manner ◽  
Cycle Arrest ◽  
Basal Transcriptional Machinery ◽  
Nucleotide Excision ◽  
Rna Polymerase Ii Transcription

The tumor suppressor protein p53 acts as a transcriptional activator that can mediate cellular responses to DNA damage by inducing apoptosis and cell cycle arrest. p53 is a nuclear phosphoprotein, and phosphorylation has been proposed to be a means by which the activity of p53 is regulated. The cyclin-dependent kinase (CDK)-activating kinase (CAK) was originally identified as a cellular kinase required for the activation of a CDK-cyclin complex, and CAK is comprised of three subunits: CDK7, cyclin H, and p36MAT1. CAK is part of the transcription factor IIH multiprotein complex, which is required for RNA polymerase II transcription and nucleotide excision repair. Because of the similarities between p53 and CAK in their involvement in the cell cycle, transcription, and repair, we investigated whether p53 could act as a substrate for phosphorylation by CAK. While CDK7-cyclin H is sufficient for phosphorylation of CDK2, we show that p36MAT1 is required for efficient phosphorylation of p53 by CDK7-cyclin H, suggesting that p36MAT1 can act as a substrate specificity-determining factor for CDK7-cyclin H. We have mapped a major site of phosphorylation by CAK to Ser-33 of p53 and have demonstrated as well that p53 is phosphorylated at this site in vivo. Both wild-type and tumor-derived mutant p53 proteins are efficiently phosphorylated by CAK. Furthermore, we show that p36 and p53 can interact both in vitro and in vivo. These studies reveal a potential mechanism for coupling the regulation of p53 with DNA repair and the basal transcriptional machinery.

scholarly journals Histone tails cooperate to control the breathing of genomic nucleosomes

2020 ◽  
Author(s):  
Jan Huertas ◽  
Hans R Schöler ◽  
Vlad Cojocaru
Keyword(s):  
Epigenetic Regulation ◽  
Genomic Dna ◽  
Histone H3 ◽  
Molecular Simulations ◽  
Dna Interaction ◽  
Structural Transitions ◽  
Histone Tail ◽  
Chemical Modifications ◽  
Histone Tails ◽  
Chromatin Dynamics

Genomic DNA is packaged in chromatin, a dynamic fiber variable in size and compaction. In chromatin, repeating nucleosome units wrap 146 DNA basepairs around histone proteins. Genetic and epigenetic regulation of genes relies on structural transitions in chromatin which are driven by intra- and internucleosome dynamics and modulated by chemical modifications of the unstructured terminal tails of histones. Here we demonstrate how the interplay between histone H3 and H2A tails control ample nucleosome breathing motions. We monitored large openings of two genomic nucleosomes, and only moderate breathing of an engineered nucleosome in atomistic molecular simulations amounting to 18μs. Transitions between open and closed nucleosome conformations were driven by the displacement and changes in compaction of the two histone tails. These motions involved changes in the DNA interaction profiles of clusters of epigenetic regulatory aminoacids in the tails. Histone tail modulated nucleosome breathing is a key mechanism of chromatin dynamics.

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