scholarly journals Integrator Subunit 4 is a ‘Symplekin-like’ Scaffold that Associates with INTS9/11 to Form the Integrator Cleavage Module

2017 ◽  
Author(s):  
Todd R. Albrecht ◽  
Sergey P. Shevtsov ◽  
Lauren G. Mascibroda ◽  
Natoya J. Peart ◽  
Iain A. Sawyer ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Cajal Body ◽  
Catalytic Core ◽  
Histone Locus Bodies ◽  
Transcriptional Regulatory ◽  
Significant Homology ◽  
Regulatory Complex ◽  
Cleavage And Polyadenylation ◽  
Histone Locus ◽  
Β Sheet

AbstractIntegrator (INT) is a transcriptional regulatory complex associated with RNA polymerase II that is required for the 3’-end processing of both UsnRNAs and enhancer RNAs. Integrator subunits 9 (INTS9) and INTS11 constitute the catalytic core of INT and are paralogues of the cleavage and polyadenylation specificity factors CPSF100 and CPSF73. While CPSF73/100 are known to associate with a third protein called Symplekin, there is no paralog of Symplekin within INT raising the question of how INTS9/11 associate with the other INT subunits. Here, we have identified that INTS4 is a specific and conserved interaction partner of INTS9/11 that does not interact with either subunit individually. Although INTS4 has no significant homology with Symplekin, it possesses N-terminal HEAT repeats similar to Symplekin but also contains a β-sheet rich C-terminal region, both of which are important to bind INTS9/11. We assess three functions of INT including UsnRNA 3’-end processing, maintenance of Cajal body integrity, and formation of histone locus bodies to conclude that INTS4/9/11 are the most critical of the INT subunits for UsnRNA biogenesis. Altogether, these results indicate that INTS4/9/11 compose a heterotrimeric complex that likely represents the Integrator ‘cleavage module’ responsible for its endonucleolytic activity.

scholarly journals Coilin Is Essential for Cajal Body Organization in Drosophila melanogaster

2009 ◽  
Vol 20 (6) ◽  
pp. 1661-1670 ◽  
Author(s):  
Ji-Long Liu ◽  
Zheng'an Wu ◽  
Zehra Nizami ◽  
Svetlana Deryusheva ◽  
T.K. Rajendra ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Survival Motor Neuron ◽  
Cajal Body ◽  
Homology Search ◽  
Insertion Mutant ◽  
Small Nuclear Rna ◽  
Histone Locus Bodies ◽  
Piggybac Insertion ◽  
Smn Protein ◽  
Histone Locus

Cajal bodies (CBs) are nuclear organelles that occur in a variety of organisms, including vertebrates, insects, and plants. They are most often identified with antibodies against the marker protein coilin. Because the amino acid sequence of coilin is not strongly conserved evolutionarily, coilin orthologues have been difficult to recognize by homology search. Here, we report the identification of Drosophila melanogaster coilin and describe its distribution in tissues of the fly. Surprisingly, we found coilin not only in CBs but also in histone locus bodies (HLBs), calling into question the use of coilin as an exclusive marker for CBs. We analyzed two null mutants in the coilin gene and a piggyBac insertion mutant, which leads to specific loss of coilin from the germline. All three mutants are homozygous viable and fertile. Cells that lack coilin also lack distinct foci of other CB markers, including fibrillarin, the survival motor neuron (SMN) protein, U2 small nuclear RNA (snRNA), U5 snRNA, and the small CB-specific (sca) RNA U85. However, HLBs are not obviously affected in coilin-null flies. Thus, coilin is required for normal CB organization in Drosophila but is not essential for viability or production of functional gametes.

scholarly journals RNA polymerase II condensate formation and association with Cajal and histone locus bodies in living human cells

2020 ◽  
Author(s):  
Takashi Imada ◽  
Takeshi Shimi ◽  
Ai Kaiho ◽  
Yasushi Saeki ◽  
Hiroshi Kimura
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Gene Clusters ◽  
Nuclear Bodies ◽  
Histone Genes ◽  
Small Nuclear Rna ◽  
Pol Ii ◽  
Histone Locus Bodies ◽  
Dynamic Structures ◽  
Histone Locus

ABSTRACTIn eukaryotic nuclei, a number of phase-separated nuclear bodies (NBs) are present. RNA polymerase II (Pol II) is the main player in transcription and forms large condensates in addition to localizing at numerous transcription foci. Cajal bodies (CBs) and histone locus bodies (HLBs) are NBs that are involved in transcriptional and post-transcriptional regulation of small nuclear RNA and histone genes. By live-cell imaging using human HCT116 cells, we here show that Pol II condensates (PCs) nucleated near CBs and HLBs, and the number of PCs increased during S phase concomitantly with the activation period of histone genes. Ternary PC–CB– HLB associates were formed via three pathways: nucleation of PCs and HLBs near CBs, interaction between preformed PC–HLBs with CBs, and nucleation of PCs near preformed CB– HLBs. Coilin knockout increased the co-localization rate between PCs and HLBs, whereas the number, nucleation timing, and phosphorylation status of PCs remained unchanged. Depletion of PCs did not affect CBs and HLBs. Treatment with 1,6-hexanediol revealed that PCs were more liquid-like than CBs and HLBs. Thus, PCs are dynamic structures often nucleated following the activation of gene clusters associated with other NBs. (187 words)

scholarly journals Drosophila CG2469 Encodes a Homolog of Human CTR9 and Is Essential for Development

2016 ◽  
Vol 6 (12) ◽  
pp. 3849-3857 ◽  
Author(s):  
Dhananjay Chaturvedi ◽  
Mayu Inaba ◽  
Shane Scoggin ◽  
Michael Buszczak
Keyword(s):  
Germ Cell ◽  
Sequence Similarity ◽  
Cell Loss ◽  
Germline Stem Cell ◽  
Cell Nuclei ◽  
Transcriptional Initiation ◽  
Histone Locus Bodies ◽  
Paf1 Complex ◽  
Histone Locus ◽  
Drosophila Cells

Abstract Conserved from yeast to humans, the Paf1 complex participates in a number of diverse processes including transcriptional initiation and polyadenylation. This complex typically includes five proteins: Paf1, Rtf1, Cdc73, Leo1, and Ctr9. Previous efforts identified clear Drosophila homologs of Paf1, Rtf1, and Cdc73 based on sequence similarity. Further work showed that these proteins help to regulate gene expression and are required for viability. To date, a Drosophila homolog of Ctr9 has remained uncharacterized. Here, we show that the gene CG2469 encodes a functional Drosophila Ctr9 homolog. Both human and Drosophila Ctr9 localize to the nuclei of Drosophila cells and appear enriched in histone locus bodies. RNAi knockdown of Drosophila Ctr9 results in a germline stem cell loss phenotype marked by defects in the morphology of germ cell nuclei. A molecular null mutation of Drosophila Ctr9 results in lethality and a human cDNA CTR9 transgene rescues this phenotype. Clonal analysis in the ovary using this null allele reveals that loss of Drosophila Ctr9 results in a reduction of global levels of histone H3 trimethylation of lysine 4 (H3K4me3), but does not compromise the maintenance of stem cells in ovaries. Given the differences between the null mutant and RNAi knockdown phenotypes, the germ cell defects caused by RNAi likely result from the combined loss of Drosophila Ctr9 and other unidentified genes. These data provide further evidence that the function of this Paf1 complex component is conserved across species.

scholarly journals The Essential WD Repeat Protein Swd2 Has Dual Functions in RNA Polymerase II Transcription Termination and Lysine 4 Methylation of Histone H3

2004 ◽  
Vol 24 (7) ◽  
pp. 2932-2943 ◽  
Author(s):  
Hailing Cheng ◽  
Xiaoyuan He ◽  
Claire Moore
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Chromatin Modification ◽  
Histone H3 ◽  
Temperature Sensitive ◽  
Repeat Protein ◽  
Tightly Coupled ◽  
Cleavage And Polyadenylation ◽  
Wd Repeat

ABSTRACT Swd2, an essential WD repeat protein in Saccharomyces cerevisiae, is a component of two very different complexes: the cleavage and polyadenylation factor CPF and the Set1 methylase, which modifies lysine 4 of histone H3 (H3-K4). It was not known if Swd2 is important for the function of either of these entities. We show here that, in extract from cells depleted of Swd2, cleavage and polyadenylation of the mRNA precursor in vitro are completely normal. However, temperature-sensitive mutations or depletion of Swd2 causes termination defects in some genes transcribed by RNA polymerase II. Overexpression of Ref2, a protein previously implicated in snoRNA 3′ end formation and Swd2 recruitment to CPF, can rescue the growth and termination defects, indicating a functional interaction between the two proteins. Some swd2 mutations also significantly decrease global H3-K4 methylation and cause other phenotypes associated with loss of this chromatin modification, such as loss of telomere silencing, hydroxyurea sensitivity, and alterations in repression of INO1 transcription. Even though the two Swd2-containing complexes are both localized to actively transcribed genes, the allele specificities of swd2 defects suggest that the functions of Swd2 in mediating RNA polymerase II termination and H3-K4 methylation are not tightly coupled.

scholarly journals Identification of a mouse protein whose homolog in Saccharomyces cerevisiae is a component of the CCR4 transcriptional regulatory complex.

1995 ◽  
Vol 15 (7) ◽  
pp. 3487-3495 ◽  
Author(s):  
M P Draper ◽  
C Salvadore ◽  
C L Denis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Yeast Cells ◽  
Significant Similarity ◽  
Eukaryotic Transcription ◽  
C Elegans ◽  
Mouse Protein ◽  
Transcriptional Regulatory ◽  
Evolutionarily Conserved ◽  
Regulatory Complex

The CCR4 protein from Saccharomyces cerevisiae is a component of a multisubunit complex that is required for the regulation of a number of genes in yeast cells. We report here the identification of a mouse protein (mCAF1 [mouse CCR4-associated factor 1]) which is capable of interacting with and binding to the yeast CCR4 protein. The mCAF1 protein was shown to have significant similarity to proteins from humans, Caenorhabditis elegans, Arabidopsis thaliana, and S. cerevisiae. The yeast gene (yCAF1) had been previously cloned as the POP2 gene, which is required for expression of several genes. Both yCAF1 (POP2) and the C. elegans homolog of CAF1 were shown to genetically interact with CCR4 in vivo, and yCAF1 (POP2) physically associated with CCR4. Disruption of the CAF1 (POP2) gene in yeast cells gave phenotypes and defects in transcription similar to those observed with disruptions of CCR4, including the ability to suppress spt10-enhanced ADH2 expression. In addition, yCAF1 (POP2) when fused to LexA was capable of activating transcription. mCAF1 could also activate transcription when fused to LexA and could functionally substitute for yCAF1 in allowing ADH2 expression in an spt10 mutant background. These data imply that CAF1 is a component of the CCR4 protein complex and that this complex has retained evolutionarily conserved functions important to eukaryotic transcription.

scholarly journals Interaction ofBTG1and p53-regulatedBTG2Gene Products with mCaf1, the Murine Homolog of a Component of the Yeast CCR4 Transcriptional Regulatory Complex

1998 ◽  
Vol 273 (35) ◽  
pp. 22563-22569 ◽  
Author(s):  
Jean-Pierre Rouault ◽  
Déborah Prévôt ◽  
Cyril Berthet ◽  
Anne-Marie Birot ◽  
Marc Billaud ◽  
...  
Keyword(s):  
Transcriptional Regulatory ◽  
Murine Homolog ◽  
Regulatory Complex

scholarly journals CDK-Regulated Phase Separation Seeded by Histone Genes Ensures Precise Growth and Function of Histone Locus Bodies

2020 ◽  
Vol 54 (3) ◽  
pp. 379-394.e6 ◽  
Author(s):  
Woonyung Hur ◽  
James P. Kemp ◽  
Marco Tarzia ◽  
Victoria E. Deneke ◽  
William F. Marzluff ◽  
...  
Keyword(s):  
Phase Separation ◽  
Histone Genes ◽  
Histone Locus Bodies ◽  
And Function ◽  
Histone Locus

scholarly journals Adenovirus E1A specifically blocks SWI/SNF-dependent transcriptional activation.

1996 ◽  
Vol 16 (10) ◽  
pp. 5737-5743 ◽  
Author(s):  
M E Miller ◽  
B R Cairns ◽  
R S Levinson ◽  
K R Yamamoto ◽  
D A Engel ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Cellular Transformation ◽  
Transcriptional Coactivator ◽  
Gene Encoding ◽  
Adenovirus E1a ◽  
Transcriptional Regulatory ◽  
Genes Encoding ◽  
Regulatory Complex ◽  
Activation Pathways ◽  
Coactivator P300

Expression of the adenovirus E1A243 oncoprotein in Saccharomyces cerevisiae produces a slow-growth phenotype with accumulation of cells in the G1 phase of the cell cycle. This effect is due to the N-terminal and CR1 domains of E1A243, which in rodent cells are involved in triggering cellular transformation and also in binding to the cellular transcriptional coactivator p300. A genetic screen was undertaken to identify genes required for the function of E1A243 in S. cerevisiae. This screen identified SNF12, a gene encoding the 73-kDa subunit of the SWI/SNF transcriptional regulatory complex. Mutation of genes encoding known members of the SWI/SNF complex also led to loss of E1A function, suggesting that the SWI/SNF complex is a target of E1A243. Moreover, expression of E1A in wild-type cells specifically blocked transcriptional activation of the INO1 and SUC2 genes, whose activation pathways are distinct but have a common requirement for the SWI/SNF complex. These data demonstrate a specific functional interaction between E1A and the SWI/SNF complex and suggest that a similar interaction takes place in rodent and human cells.

scholarly journals Structural mechanism of Myb–MuvB assembly

2018 ◽  
Vol 115 (40) ◽  
pp. 10016-10021 ◽  
Author(s):  
Keelan Z. Guiley ◽  
Audra N. Iness ◽  
Siddharth Saini ◽  
Sarvind Tripathi ◽  
Joseph S. Lipsick ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Biochemical Analysis ◽  
Adaptor Protein ◽  
Cancer Cell Proliferation ◽  
Cycle Progression ◽  
Structural Mechanism ◽  
Transcriptional Regulatory ◽  
Regulatory Complex ◽  
Cycle Dependent

The MuvB transcriptional regulatory complex, which controls cell-cycle-dependent gene expression, cooperates with B-Myb to activate genes required for the G2 and M phases of the cell cycle. We have identified the domain in B-Myb that is essential for the assembly of the Myb–MuvB (MMB) complex. We determined a crystal structure that reveals how this B-Myb domain binds MuvB through the adaptor protein LIN52 and the scaffold protein LIN9. The structure and biochemical analysis provide an understanding of how oncogenic B-Myb is recruited to regulate genes required for cell-cycle progression, and the MMB interface presents a potential therapeutic target to inhibit cancer cell proliferation.

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