scholarly journals Human hHpr1/p84/Thoc1 Regulates Transcriptional Elongation and Physically Links RNA Polymerase II and RNA Processing Factors

2005 ◽  
Vol 25 (10) ◽  
pp. 4023-4033 ◽  
Author(s):  
Yanping Li ◽  
Xiaoling Wang ◽  
Xiaojing Zhang ◽  
David W. Goodrich
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Processing ◽  
Rna Splicing ◽  
Transcriptional Elongation ◽  
Intact Cells ◽  
Efficient Gene ◽  
Cellular Phenotypes ◽  
Processing Factors

ABSTRACT Cotranscriptional loading of RNA processing factors onto nascent RNA facilitates efficient gene expression. Mechanisms responsible for coupling transcription and RNA processing are not well defined, but the Saccharomyces cerevisiae TREX complex provides an example. TREX is composed of the subcomplex THO that associates with RNA polymerase II and is required for normal transcriptional elongation. THO associates with proteins involved in RNA splicing and export to form the larger TREX complex. Hence, assembly of TREX physically couples transcriptional elongation with RNA processing factors. Whether metazoan species with long, intron-containing genes utilize a similar mechanism has not been established. Here we show that human hHpr1/p84/Thoc1 associates with elongating RNA polymerase II and the RNA splicing and export factor UAP56 in intact cells. Depletion of hHpr1/p84/Thoc1 causes transcriptional elongation defects and associated cellular phenotypes similar to those observed in THO-deficient yeast. We conclude that hHpr1/p84/Thoc1 regulates transcriptional elongation and may participate in a protein complex functionally analogous to yeast TREX, physically linking elongating RNA polymerase II with RNA processing factors.

In vivo interactions of RNA polymerase II with genes of Drosophila melanogaster

1985 ◽  
Vol 5 (8) ◽  
pp. 2009-2018
Author(s):  
D S Gilmour ◽  
J T Lis
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Protein Characterization ◽  
Heat Shock Gene ◽  
Transcriptional Level ◽  
Intact Cells

We describe a method for examining the in vivo distribution of a protein on specific eucaryotic DNA sequences. In this method, proteins are cross-linked to DNA in intact cells, and the protein-DNA adducts are isolated by immunoprecipitation with antiserum against the protein. Characterization of the DNA cross-linked to the precipitated protein identifies the sequences with which the protein is associated in vivo. Here, we applied these methods to detect RNA polymerase II-DNA interactions in heat-shocked and untreated Drosophila melanogaster Schneider line 2 cells. The level of RNA polymerase II associated with several heat shock genes increased dramatically in response to heat shock, whereas the level associated with the copia genes decreased, indicating that both induction of heat shock gene expression and repression of the copia gene expression by heat shock occur at the transcriptional level. Low levels of RNA polymerase II were present on DNA outside of the transcription units, and for at least two genes, hsp83 and hsp26, RNA polymerase II initiated binding near the transcription start site. Moreover, for hsp70, the density of RNA polymerase II on sequences downstream of the polyadenylate addition site was much lower than that observed on the gene internal sequences. Examination of the amount of specific restriction fragments cross-linked to RNA polymerase II provides a means of detecting RNA polymerase II on individual members of multigene families. This analysis shows that RNA polymerase II is associated with only one of the two cytoplasmic actin genes.

scholarly journals Elongin B-Mediated Epigenetic Alteration of Viral Chromatin Correlates with Efficient Human Cytomegalovirus Gene Expression and Replication

mBio ◽  
2011 ◽  
Vol 2 (2) ◽  
Author(s):  
Jiwon Hwang ◽  
Ryan T. Saffert ◽  
Robert F. Kalejta
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Human Cytomegalovirus ◽  
Viral Gene ◽  
Epigenetic Mark ◽  
Protein Accumulation ◽  
Transcriptional Elongation ◽  
Histone H2b ◽  
Cellular Genes

ABSTRACTElongins B and C are members of complexes that increase the efficiency of transcriptional elongation by RNA polymerase II (RNAPII) and enhance the monoubiquitination of histone H2B, an epigenetic mark of actively transcribed genes. Here we show that, in addition to its role in facilitating transcription of the cellular genome, elongin B also enhances gene expression from the double-stranded DNA genome of human cytomegalovirus (HCMV), a pathogenic herpesvirus. Reducing the level of elongin B by small interfering RNA- or short hairpin RNA-mediated knockdown decreased viral mRNA expression, viral protein accumulation, viral DNA replication, and infectious virion production. Chromatin immunoprecipitation analysis indicated viral genome occupancy of the elongating form of RNAPII, and monoubiquitinated histone H2B was reduced in elongin B-deficient cells. These data suggest that, in addition to the previously documented epigenetic regulation of transcriptional initiation, HCMV also subverts cellular elongin B-mediated epigenetic mechanisms for enhancing transcriptional elongation to enhance viral gene expression and virus replication.IMPORTANCEThe genetic and epigenetic control of transcription initiation at both cellular and viral promoters is well documented. Recently, the epigenetic modification of histone H2B monoubiquitination throughout the bodies of cellular genes has been shown to enhance the elongation of RNA polymerase II-initiated transcripts. Mechanisms that might control the elongation of viral transcripts are less well studied. Here we show that, as with cellular genes, elongin B-mediated monoubiquitination of histone H2B also facilitates the transcriptional elongation of human cytomegalovirus genes. This and perhaps other epigenetic markings of actively transcribed regions may help in identifying viral genes expressed duringin vitrolatency or during natural infections of humans. Furthermore, this work identifies a novel, tractable model system to further study the regulation of transcriptional elongation in living cells.

scholarly journals Coordination of transcription, processing, and export of highly expressed RNAs by distinct biomolecular condensates

2020 ◽  
Vol 4 (3) ◽  
pp. 281-291 ◽  
Author(s):  
Alexander M. Ishov ◽  
Aishwarya Gurumurthy ◽  
Jörg Bungert
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Rna Processing ◽  
Transcription Initiation ◽  
Nuclear Speckles ◽  
High Concentration ◽  
Pol Ii ◽  
Processing Factors

Genes under control of super-enhancers are expressed at extremely high levels and are frequently associated with nuclear speckles. Recent data suggest that the high concentration of unphosphorylated RNA polymerase II (Pol II) and Mediator recruited to super-enhancers create phase-separated condensates. Transcription initiates within or at the surface of these phase-separated droplets and the phosphorylation of Pol II, associated with transcription initiation and elongation, dissociates Pol II from these domains leading to engagement with nuclear speckles, which are enriched with RNA processing factors. The transitioning of Pol II from transcription initiation domains to RNA processing domains effectively co-ordinates transcription and processing of highly expressed RNAs which are then rapidly exported into the cytoplasm.

scholarly journals In vivo interactions of RNA polymerase II with genes of Drosophila melanogaster.

1985 ◽  
Vol 5 (8) ◽  
pp. 2009-2018 ◽  
Author(s):  
D S Gilmour ◽  
J T Lis
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Protein Characterization ◽  
Heat Shock Gene ◽  
Transcriptional Level ◽  
Intact Cells

We describe a method for examining the in vivo distribution of a protein on specific eucaryotic DNA sequences. In this method, proteins are cross-linked to DNA in intact cells, and the protein-DNA adducts are isolated by immunoprecipitation with antiserum against the protein. Characterization of the DNA cross-linked to the precipitated protein identifies the sequences with which the protein is associated in vivo. Here, we applied these methods to detect RNA polymerase II-DNA interactions in heat-shocked and untreated Drosophila melanogaster Schneider line 2 cells. The level of RNA polymerase II associated with several heat shock genes increased dramatically in response to heat shock, whereas the level associated with the copia genes decreased, indicating that both induction of heat shock gene expression and repression of the copia gene expression by heat shock occur at the transcriptional level. Low levels of RNA polymerase II were present on DNA outside of the transcription units, and for at least two genes, hsp83 and hsp26, RNA polymerase II initiated binding near the transcription start site. Moreover, for hsp70, the density of RNA polymerase II on sequences downstream of the polyadenylate addition site was much lower than that observed on the gene internal sequences. Examination of the amount of specific restriction fragments cross-linked to RNA polymerase II provides a means of detecting RNA polymerase II on individual members of multigene families. This analysis shows that RNA polymerase II is associated with only one of the two cytoplasmic actin genes.

Nutrient-regulated gene expression in eukaryotes

2006 ◽  
Vol 73 ◽  
pp. 85-96 ◽  
Author(s):  
Richard J. Reece ◽  
Laila Beynon ◽  
Stacey Holden ◽  
Amanda D. Hughes ◽  
Karine Rébora ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Control ◽  
Direct Interaction ◽  
Signalling Pathways ◽  
A Cell ◽  
One Step ◽  
Higher Eukaryotes ◽  
Regulated Gene Expression

The recognition of changes in environmental conditions, and the ability to adapt to these changes, is essential for the viability of cells. There are numerous well characterized systems by which the presence or absence of an individual metabolite may be recognized by a cell. However, the recognition of a metabolite is just one step in a process that often results in changes in the expression of whole sets of genes required to respond to that metabolite. In higher eukaryotes, the signalling pathway between metabolite recognition and transcriptional control can be complex. Recent evidence from the relatively simple eukaryote yeast suggests that complex signalling pathways may be circumvented through the direct interaction between individual metabolites and regulators of RNA polymerase II-mediated transcription. Biochemical and structural analyses are beginning to unravel these elegant genetic control elements.

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