Organization of RNA polymerase II transcription and pre-mRNA splicing within the mammalian cell nucleus

1993 ◽  
Vol 21 (4) ◽  
pp. 918-920 ◽  
Author(s):  
David L. Spector ◽  
Shelley Landon ◽  
Raymond T. O'Keefe
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Mammalian Cell ◽  
Cell Nucleus ◽  
Mrna Splicing ◽  
Rna Polymerase Ii Transcription

scholarly journals Coupling of RNA Polymerase II Transcription Elongation with Pre-mRNA Splicing

2016 ◽  
Vol 428 (12) ◽  
pp. 2623-2635 ◽  
Author(s):  
Tassa Saldi ◽  
Michael A. Cortazar ◽  
Ryan M. Sheridan ◽  
David L. Bentley
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Elongation ◽  
Mrna Splicing ◽  
Rna Polymerase Ii Transcription

scholarly journals Spatial Organization of RNA Polymerase II Revealed by Super-Resolution Imaging of Mammalian Cell Nucleus

2013 ◽  
Vol 104 (2) ◽  
pp. 339a
Author(s):  
Ziqing Zhao ◽  
Rahul Roy ◽  
J. Christof M. Gebhardt ◽  
David M. Suter ◽  
Alec R. Chapman ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Mammalian Cell ◽  
Cell Nucleus ◽  
Spatial Organization ◽  
Super Resolution ◽  
Resolution Imaging

scholarly journals Serine/threonine phosphatase 1 modulates the subnuclear distribution of pre-mRNA splicing factors.

1996 ◽  
Vol 7 (10) ◽  
pp. 1559-1572 ◽  
Author(s):  
T Misteli ◽  
D L Spector
Keyword(s):  
Rna Polymerase Ii ◽  
Mammalian Cell ◽  
Cell Nucleus ◽  
Nuclear Extract ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Cell Nuclei ◽  
Nuclear Speckles ◽  
Permeabilized Cells

HeLa cell nuclei were permeabilized and reconstituted with nuclear extract to identify soluble nuclear factors which play a role in the organization of pre-mRNA splicing factors in the mammalian cell nucleus. Permeabilized nuclei reconstituted with nuclear extract were active in transcription and DNA replication and nuclear speckles containing pre-mRNA splicing factors were maintained over several hours independent of soluble nuclear components. The characteristic rounding up of nuclear speckles in response to inhibition of RNA polymerase II seen in vivo was reproduced in permeabilized cells and was strictly dependent on a catalytic activity present in the nuclear extract. By inhibitor titration experiments and sensitivity to inhibitor 2, this activity was identified as a member of the serine/threonine protein phosphatase 1 family (PP1). Interference with PP1 activity affected the distribution of pre-mRNA splicing factors in transcriptionally active, permeabilized cells, and excess PP1 activity caused increased dephosphorylation of SR proteins in nuclear speckles. These data show that the dynamic reorganization of the mammalian cell nucleus can be studied in permeabilized cells and that PP1 is involved in the rounding up of speckles as well as the overall organization of pre-mRNA splicing factors in the mammalian cell nucleus.

scholarly journals Human prefoldin modulates co-transcriptional pre-mRNA splicing

2020 ◽  
Author(s):  
L Payán-Bravo ◽  
X Peñate ◽  
I Cases ◽  
Y Pareja-Sánchez ◽  
S Fontalva ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Cell Nucleus ◽  
Transcription Elongation ◽  
Mrna Splicing ◽  
Splicing Factor ◽  
Positive Contribution ◽  
Chromatin Dynamics ◽  
Serum Stimulation ◽  
Significant Impairment

AbstractPrefoldin is a heterohexameric complex conserved from archaea to humans that plays a cochaperone role during the cotranslational folding of actin and tubulin monomers. Additional functions of prefoldin in the cell nucleus have been described, including a positive contribution to transcription elongation and chromatin dynamics in yeast. Here we show that prefoldin depletion provoked transcriptional alterations across the genome of human cells. Severe pre-mRNA splicing defects were also detected, particularly under serum stimulation conditions. We found a significant impairment of co-transcriptional splicing during transcription elongation, which explains why the expression of long genes with a high number of introns was affected the most. We detected prefoldin binding to the gene body of transcribed genes and found that its absence caused significant decrease in the levels of Ser2 phosphorylation of the RNA polymerase II carboxy-terminal domain. Moreover, lack of prefoldin reduced the association of the splicing factor U2AF65 with chromatin, an association that is known to be dependent on Ser2 phosphorylation. Altogether the reported results indicate that human prefoldin is able to modulate gene expression by influencing phosphorylation of elongating RNA polymerase II, and thereby regulating co-transcriptional splicing efficiency.Significance StatementPrefoldin is a protein complex conserved from archaea to humans. It is known that human prefoldin facilitates folding of cytoskeleton monomers in the cytoplasm, but its role in the cell nucleus is unclear. We found prefoldin bound to chromatin in transcribed genes and detected accumulation of unspliced pre-mRNA precursors when prefoldin was depleted, particularly in serum-activated cells. We also found that lack of prefoldin decreases the levels of a phosphorylated form of RNA polymerase II that is the best-known marker of transcription elongation. This defect, in turn, impairs recruitment of the splicing factor U2AF65, provoking the uncoupling of pre-mRNA splicing from transcription. These results show that prefoldin is, in addition to a cytoplasmic chaperone, a modulator of co-transcriptional pre-mRNA processing.

scholarly journals Nuclear mRNA accumulation causes nucleolar fragmentation in yeast mtr2 mutant.

1994 ◽  
Vol 5 (11) ◽  
pp. 1253-1263 ◽  
Author(s):  
T Kadowaki ◽  
M Hitomi ◽  
S Chen ◽  
A M Tartakoff
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Nuclear Protein ◽  
Tail Length ◽  
Mrna Splicing ◽  
Rrna Synthesis ◽  
Polya Tail ◽  
Mrna Accumulation ◽  
Rna Accumulation ◽  
Rna Polymerase Ii Transcription

We have identified a set of genes that affect mRNA transport (mtr) from the nucleus to the cytoplasm of Saccharomyces cerevisiae. One of these genes, MTR2, has been cloned and shown to encode a novel 21-kDa nuclear protein that is essential for vegetative growth. MTR2 shows limited homology to a protein implicated in plasmid DNA transfer in Escherichia coli. PolyA+RNA accumulates within the nucleus of mtr2-1 in two to three foci at 37 degrees C. mRNA, tRNA, and rRNA synthesis continue as do pre-mRNA splicing, tRNA processing, and rRNA export at 37 degrees C. Under these conditions the polyA tail length increases, and protein synthesis is progressively inhibited. Nucleolar antigens also redistribute to two to three nuclear foci at 37 degrees C, and this redistribution depends on ongoing transcription by RNA polymerase II. Surprisingly, these foci coincide with the sites of polyA+RNA accumulation. Comparable colocalization and dependance on RNA polymerase II transcription is seen for the mtr1-1 mutant. The disorganization of the nucleolus thus depends on mRNA accumulation in these mutants. We discuss the possible functions of MTR2 and the yeast nucleolus in mRNA export.

Organization of transcription and pre-mRNA splicing within the mammalian cell nucleus

1995 ◽  
Vol 53 ◽  
pp. 768-769
Author(s):  
D.L. Spector ◽  
S. Huang ◽  
S. Kaurin
Keyword(s):  
Rna Polymerase Ii ◽  
Cell Nucleus ◽  
Functional Organization ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Interchromatin Granule Clusters ◽  
Interchromatin Granule ◽  
Nuclear Regions ◽  
Relationship Of ◽  
Perichromatin Fibrils

We have been interested in the organization of RNA polymerase II transcription and pre-mRNA splicing within the cell nucleus. Several models have been proposed for the functional organization of RNA within the eukaryotic nucleus and for the relationship of this organization to the distribution of pre-mRNA splicing factors. One model suggests that RNAs which must be spliced are capable of recruiting splicing factors to the sites of transcription from storage and/or reassembly sites. When one examines the organization of splicing factors in the nucleus in comparison to the sites of chromatin it is clear that splicing factors are not localized in coincidence with heterochromatin (Fig. 1). Instead, they are distributed in a speckled pattern which is composed of both perichromatin fibrils and interchromatin granule clusters. The perichromatin fibrils are distributed on the periphery of heterochromatin and on the periphery of interchromatin granule clusters as well as being diffusely distributed throughout the nucleoplasm. These nuclear regions have been previously shown to represent initial sites of incorporation of 3H-uridine.

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