Histone H3K4 trimethylation: dynamic interplay with pre-mRNA splicing

2016 ◽  
Vol 94 (1) ◽  
pp. 1-11 ◽  
Author(s):  
James R. Davie ◽  
Wayne Xu ◽  
Genevieve P. Delcuve
Keyword(s):  
Mammalian Cells ◽  
Cpg Island ◽  
Protein Complexes ◽  
Cpg Islands ◽  
Mrna Splicing ◽  
Chromatin Remodelers ◽  
Multiple Protein ◽  
H3k4me3 Mark ◽  
Histone H3k4 ◽  
Enzyme Complexes

Histone H3 lysine 4 trimethylation (H3K4me3) is often stated as a mark of transcriptionally active promoters. However, closer study of the positioning of H3K4me3 shows the mark locating primarily after the first exon at the 5′ splice site and overlapping with a CpG island in mammalian cells. There are several enzyme complexes that are involved in the placement of the H3K4me3 mark, including multiple protein complexes containing SETD1A, SETD1B, and MLL1 enzymes (writers). CXXC1, which is associated with SETD1A and SETD1B, target these enzymes to unmethylated CpG islands. Lysine demethylases (KDM5 family members, erasers) demethylate H3K4me3. The H3K4me3 mark is recognized by several proteins (readers), including lysine acetyltransferase complexes, chromatin remodelers, and RNA bound proteins involved in pre-mRNA splicing. Interestingly, attenuation of H3K4me3 impacts pre-mRNA splicing, and inhibition of pre-mRNA splicing attenuates H3K4me3.

scholarly journals PTE, a novel module to target Polycomb Repressive Complex 1 to the Cyclin D2 oncogene

2017 ◽  
Author(s):  
Sarina R. Cameron ◽  
Soumyadeep Nandi ◽  
Tatyana G. Kahn ◽  
Juan I. Barrasa ◽  
Per Stenberg ◽  
...  
Keyword(s):  
Cpg Island ◽  
Protein Complexes ◽  
Cpg Islands ◽  
Biochemical Properties ◽  
Polycomb Group ◽  
Cyclin D2 ◽  
Polycomb Response Element ◽  
Multiple Protein ◽  
Polycomb Repression ◽  
Polycomb Repressive Complex 1

AbstractPolycomb Group proteins are essential epigenetic repressors. They form multiple protein complexes of which two kinds, PRC1 and PRC2, are indispensable for repression. Although much is known about their biochemical properties, how PRC1 and PRC2 are targeted to specific genes is poorly understood. Here we establish the Cyclin D2 (CCND2) oncogene as a simple model to address this question. We provide the evidence that coordinated recruitment of PRC1 and PRC2 complexes to CCND2 involves a combination of a specialized PRC1 targeting element (PTE) and an adjacent CpG-island, which together act as a human Polycomb Response Element. Chromatin immunoprecipitation analysis of CCND2 in different transcriptional states indicates that histone modifications produced by PRC1 and PRC2 are not sufficient to recruit either of the complexes. However, catalytic activity of PRC2 helps to anchor PRC1 at the PTE. Our analyses suggest that coordinated targeting of PRC1 and PRC2 complexes by juxtaposed AT-rich PTEs and CpG-islands may be a general feature of Polycomb repression in mammals.

scholarly journals The Core Element of a CpG Island Protects Avian Sarcoma and Leukosis Virus-Derived Vectors from Transcriptional Silencing

2008 ◽  
Vol 82 (16) ◽  
pp. 7818-7827 ◽  
Author(s):  
Filip Šenigl ◽  
Jiří Plachý ◽  
Jiří Hejnar
Keyword(s):  
Binding Sites ◽  
Mammalian Cells ◽  
Cpg Island ◽  
Cpg Islands ◽  
Transcriptional Silencing ◽  
Retroviral Vectors ◽  
Core Element ◽  
Promoter Sequences ◽  
Rous Sarcoma ◽  
Sarcoma Virus

ABSTRACT Unmethylated CpG islands are known to keep adjacent promoters transcriptionally active. In the CpG island adjacent to the adenosine phosphoribosyltransferase gene, the protection against transcriptional silencing can be attributed to the short CpG-rich core element containing Sp1 binding sites. We report here the insertion of this CpG island core element, IE, into the long terminal repeat of a retroviral vector derived from Rous sarcoma virus, which normally suffers from progressive transcriptional silencing in mammalian cells. IE insertion into a specific position between enhancer and promoter sequences led to efficient protection of the integrated vector from silencing and gradual CpG methylation in rodent and human cells. Individual cell clones with IE-modified reporter vectors display high levels of reporter expression for a sustained period and without substantial variegation in the cell culture. The presence of Sp1 binding sites is important for the protective effect of IE, but at least some part of the entire antisilencing capacity is maintained in IE with mutated Sp1 sites. We suggest that this strategy of antisilencing protection by the CpG island core element may prove generally useful in retroviral vectors.

scholarly journals Sem1 is a functional component of the nuclear pore complex–associated messenger RNA export machinery

2009 ◽  
Vol 184 (6) ◽  
pp. 833-846 ◽  
Author(s):  
Marius Boulos Faza ◽  
Stefan Kemmler ◽  
Sonia Jimeno ◽  
Cristina González-Aguilera ◽  
Andrés Aguilera ◽  
...  
Keyword(s):  
Nuclear Pore Complex ◽  
Messenger Rna ◽  
Mammalian Cells ◽  
Genome Instability ◽  
Protein Complexes ◽  
Mrna Export ◽  
Nuclear Pore ◽  
Multiple Protein ◽  
Rna Export ◽  
Biochemical Analyses

The evolutionarily conserved protein Sem1/Dss1 is a subunit of the regulatory particle (RP) of the proteasome, and, in mammalian cells, binds the tumor suppressor protein BRCA2. Here, we describe a new function for yeast Sem1. We show that sem1 mutants are impaired in messenger RNA (mRNA) export and transcription elongation, and induce strong transcription-associated hyper-recombination phenotypes. Importantly, Sem1, independent of the RP, is functionally linked to the mRNA export pathway. Biochemical analyses revealed that, in addition to the RP, Sem1 coenriches with components of two other multisubunit complexes: the nuclear pore complex (NPC)-associated TREX-2 complex that is required for transcription-coupled mRNA export, and the COP9 signalosome, which is involved in deneddylation. Notably, targeting of Thp1, a TREX-2 component, to the NPC is perturbed in a sem1 mutant. These findings reveal an unexpected nonproteasomal function of Sem1 in mRNA export and in prevention of transcription-associated genome instability. Thus, Sem1 is a versatile protein that might stabilize multiple protein complexes involved in diverse pathways.

Multiple protein-binding sites in the 5'-flanking region regulate c-fos expression

1986 ◽  
Vol 6 (12) ◽  
pp. 4305-4316
Author(s):  
M Z Gilman ◽  
R N Wilson ◽  
R A Weinberg
Keyword(s):  
Binding Sites ◽  
Mammalian Cells ◽  
Promoter Activity ◽  
Protein Complexes ◽  
Protein Binding Sites ◽  
Nuclear Extracts ◽  
Multiple Protein ◽  
Flanking Region ◽  
Fos Expression

We tested sequences flanking the mouse c-fos gene for the ability to form specific DNA-protein complexes with factors present in crude nuclear extracts prepared from mammalian cells. Three such complexes were detected. One complex formed in a region necessary for the induction of c-fos expression by serum growth factors. Two additional complexes formed at sequences that contribute to basal c-fos promoter activity in vivo. These complexes represent three novel sequence-specific DNA-binding activities which appear to participate in the regulation of c-fos transcription.

scholarly journals Multiple protein-binding sites in the 5'-flanking region regulate c-fos expression.

1986 ◽  
Vol 6 (12) ◽  
pp. 4305-4316 ◽  
Author(s):  
M Z Gilman ◽  
R N Wilson ◽  
R A Weinberg
Keyword(s):  
Binding Sites ◽  
Mammalian Cells ◽  
Promoter Activity ◽  
Protein Complexes ◽  
Protein Binding Sites ◽  
Nuclear Extracts ◽  
Multiple Protein ◽  
Flanking Region ◽  
Fos Expression

We tested sequences flanking the mouse c-fos gene for the ability to form specific DNA-protein complexes with factors present in crude nuclear extracts prepared from mammalian cells. Three such complexes were detected. One complex formed in a region necessary for the induction of c-fos expression by serum growth factors. Two additional complexes formed at sequences that contribute to basal c-fos promoter activity in vivo. These complexes represent three novel sequence-specific DNA-binding activities which appear to participate in the regulation of c-fos transcription.

scholarly journals DNA Methylation of Human Choline Kinase Alpha Promoter-Associated CpG Islands in MCF-7 Cells

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 853
Author(s):  
Siti Aisyah Faten Mohamed Sa’dom ◽  
Sweta Raikundalia ◽  
Shaharum Shamsuddin ◽  
Wei Cun See Too ◽  
Ling Ling Few
Keyword(s):  
Dna Methylation ◽  
Transcription Factor ◽  
Binding Site ◽  
Promoter Activity ◽  
Cytosine Methylation ◽  
Cpg Island ◽  
Cpg Islands ◽  
Site Directed Mutagenesis ◽  
Choline Kinase ◽  
Mcf 7

Choline kinase (CK) is the enzyme catalyzing the first reaction in CDP-choline pathway for the biosynthesis of phosphatidylcholine. Higher expression of the α isozyme of CK has been implicated in carcinogenesis, and inhibition or downregulation of CKα (CHKA) is a promising anticancer approach. This study aimed to investigate the regulation of CKα expression by DNA methylation of the CpG islands found on the promoter of this gene in MCF-7 cells. Four CpG islands have been predicted in the 2000 bp promoter region of ckα (chka) gene. Six CpG island deletion mutants were constructed using PCR site-directed mutagenesis method and cloned into pGL4.10 vectors for promoter activity assays. Deletion of CpG4C region located between –225 and –56 significantly increased the promoter activity by 4-fold, indicating the presence of important repressive transcription factor binding site. The promoter activity of methylated full-length promoter was significantly lower than the methylated CpG4C deletion mutant by 16-fold. The results show that DNA methylation of CpG4C promotes the binding of the transcription factor that suppresses the promoter activity. Electrophoretic mobility shift assay analysis showed that cytosine methylation at MZF1 binding site in CpG4C increased the binding of putative MZF1 in nuclear extract. In conclusion, the results suggest that DNA methylation decreased the promoter activity by promoting the binding of putative MZF1 transcription factor at CpG4C region of the ckα gene promoter.

scholarly journals Tafazzins from Drosophila and mammalian cells assemble in large protein complexes with a short half-life

Mitochondrion ◽  
2015 ◽  
Vol 21 ◽  
pp. 27-32 ◽  
Author(s):  
Yang Xu ◽  
Ashim Malhotra ◽  
Steven M. Claypool ◽  
Mindong Ren ◽  
Michael Schlame
Keyword(s):  
Half Life ◽  
Mammalian Cells ◽  
Protein Complexes ◽  
Large Protein ◽  
Short Half Life

scholarly journals A Functional Interaction between the Carboxy-Terminal Domain of RNA Polymerase II and Pre-mRNA Splicing

1997 ◽  
Vol 136 (1) ◽  
pp. 5-18 ◽  
Author(s):  
Lei Du ◽  
Stephen L. Warren
Keyword(s):  
Rna Polymerase Ii ◽  
Mammalian Cells ◽  
Mrna Splicing ◽  
Splicing Factors ◽  
Functional Interaction ◽  
Carboxy Terminal Domain ◽  
Localization Signal ◽  
Nuclear Domains ◽  
Carboxy Terminal

In the preceding study we found that Sm snRNPs and SerArg (SR) family proteins co-immunoprecipitate with Pol II molecules containing a hyperphosphorylated CTD (Kim et al., 1997). The association between Pol IIo and splicing factors is maintained in the absence of pre-mRNA, and the polymerase need not be transcriptionally engaged (Kim et al., 1997). The latter findings led us to hypothesize that a phosphorylated form of the CTD interacts with pre-mRNA splicing components in vivo. To test this idea, a nested set of CTD-derived proteins was assayed for the ability to alter the nuclear distribution of splicing factors, and to interfere with splicing in vivo. Proteins containing heptapeptides 1-52 (CTD52), 1-32 (CTD32), 1-26 (CTD26), 1-13 (CTD13), 1-6 (CTD6), 1-3 (CTD3), or 1 (CTD1) were expressed in mammalian cells. The CTD-derived proteins become phosphorylated in vivo, and accumulate in the nucleus even though they lack a conventional nuclear localization signal. CTD52 induces a selective reorganization of splicing factors from discrete nuclear domains to the diffuse nucleoplasm, and significantly, it blocks the accumulation of spliced, but not unspliced, human β-globin transcripts. The extent of splicing factor disruption, and the degree of inhibition of splicing, are proportional to the number of heptapeptides added to the protein. The above results indicate a functional interaction between Pol II's CTD and pre-mRNA splicing.

scholarly journals U1 small nuclear RNAs with altered specificity can be stably expressed in mammalian cells and promote permanent changes in pre-mRNA splicing.

1993 ◽  
Vol 13 (5) ◽  
pp. 2666-2676 ◽  
Author(s):  
J B Cohen ◽  
S D Broz ◽  
A D Levinson
Keyword(s):  
Splice Site ◽  
Mammalian Cells ◽  
Mrna Processing ◽  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
Splice Sites ◽  
Gene Complementation ◽  
Small Nuclear Rnas ◽  
Nuclear Rna ◽  
Mutant Genes

Pre-mRNA 5' splice site activity depends, at least in part, on base complementarity to U1 small nuclear RNA. In transient coexpression assays, defective 5' splice sites can regain activity in the presence of U1 carrying compensatory changes, but it is unclear whether such mutant U1 RNAs can be permanently expressed in mammalian cells. We have explored this issue to determine whether U1 small nuclear RNAs with altered specificity may be of value to rescue targeted mutant genes or alter pre-mRNA processing profiles. This effort was initiated following our observation that U1 with specificity for a splice site associated with an alternative H-ras exon substantially reduced the synthesis of the potentially oncogenic p21ras protein in transient assays. We describe the development of a mammalian complementation system that selects for removal of a splicing-defective intron placed within a drug resistance gene. Complementation was observed in proportion to the degree of complementarity between transfected mutant U1 genes and different defective splice sites, and all cells selected in this manner were found to express mutant U1 RNA. In addition, these cells showed specific activation of defective splice sites presented by an unlinked reporter gene. We discuss the prospects of this approach to permanently alter the expression of targeted genes in mammalian cells.

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