HPVE6-USP46 Mediated Cdt2 Stabilization Reduces Set8 Mediated H4K20-Methylation to Induce Gene Expression Changes
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ABSTRACT:
PROVIDER: PRJNA789318 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR17242273_1.fastq.gz | Fastqsanger.gz | |||
| SRR17242273_2.fastq.gz | Fastqsanger.gz | |||
| SRR17242274_1.fastq.gz | Fastqsanger.gz | |||
| SRR17242274_2.fastq.gz | Fastqsanger.gz | |||
| SRR17242275_1.fastq.gz | Fastqsanger.gz |
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Project description:Cervical cancer is one of the most common cancers in women. More than 95% of the cervical cancers are caused by the human papilloma viruses. Earlier we had identified a protein target, USP46 that can be inhibited to block the cancer growth. We found that the viral E6 and human USP46 complex increases the levels of Cdt2 in cervical cancers. In this paper we compared real cervical tumors with matched normal tissue from same patients and found that the effects of E6 mediated USP46 activation are indeed reflected as increased levels of Cdt2 and decreased levels of Set8 protein in the real tumors. We also found that the HPV protein E6 alone can stimulate the enzymatic activity of human USP46. By doing this it activates several cellular pathways that are required for the growth of cancers. One such pathway is the EGFR pathway that is normally upregulated in cervical cancers. We find that E6-USP46 contributes to the activation of EGFR by inducing epigenetic changes on the DNA by degrading the Set8 protein. These findings illuminate the role of viral E6 protein in inducing cancers and substantiate the candidature of USP46 as a drug target for HPV induced cancers.
2021-12-20 | GSE191010 | GEO
Project description:Histone methylation has an important role in transcriptional regulation. However, unlike H3K4 and H3K9 methylation, the role of H4K20 mono-methylation (H4K20me-1) in transcriptional regulation remains unclear. Here we show that Wnt3a specifically stimulates H4K20 mono-methylation at TBE (TCF-binding element) via the histone methylase SET8. Additionally, SET8 is crucial for activation of the Wnt reporter gene and target genes in both mammalian cells and zebrafish. Furthermore, SET8 interacts with TCF4/LEF1 directly, and this interaction is regulated by Wnt3a. Therefore, we conclude that SET8 is a Wnt signaling mediator and is recruited by LEF1/TCF4 to regulate the transcription of Wnt-activated genes, possibly via H4K20 mono-methylation at the target gene promoters. Our findings also indicate that H4K20me-1 is a marker for gene transcription activation, at least in canonical Wnt signaling. This chip experiment was carried out to find out how many genes would be regulated by SET8 after Wnt3a CM stimulated HEK293 cells. Total RNA was extracted from HEK293 cells after siRNA transfection (control or SET8 shRNA) for about 40 hours and then induced with Control or Wnt3a CM for 7 hours using TRIzol (Invitrogen) and the RNeasy kit (Qiagen). Three pairs of samples: si-control + control CM / si-control + Wnt3a CM 7 h / si-SET8 + Wnt3a CM. Samples were amplified and labeled using a NimbleGen One-Color DNA Labeling Kit.
2010-12-29 | E-GEOD-24708 | biostudies-arrayexpress
Project description:Ubiquitin-mediated proteolysis play a significant role in various biological processes including transcription, DNA repair and cell cycle progression. The identification of Set8 and Set8b (a splice isoform) histone H4K20 methyl transferase as a substrate for the cullin-based ubiquitin ligase (CRL4-Cdt2) demonstrate that this pathway plays a significant role in promoting cell cycle progression, specifically promoting G2 progression. This study investigate the effect of failure to degrade Set8 in S-phase of the cell cycle via CRL4-Cdt2 on gene expression. We used microarrays to detail the effect of the expression of stable form of Set8b H4K20 mono-methyl transferase (Set8b_deltaPIP2) on gene expression Human osteosarcoma-derived human cells were transduced with retroviruses encoding either wt-Set8b or a mutant of Setb8 which is resistant to degradation via CRL4-Cdt2 ubiquitin ligase complex (Set8b_deltaPIP2) or with an a control pMSCV empty virus. 5 days after transduction, cells were harvested and the RNA was extracted by Trizol (Invitrogen) and hybridized to the affymetrix chips array.
2010-09-18 | E-GEOD-24170 | biostudies-arrayexpress
Project description:Histone methylation has an important role in transcriptional regulation. However, unlike H3K4 and H3K9 methylation, the role of H4K20 mono-methylation (H4K20me-1) in transcriptional regulation remains unclear. Here we show that Wnt3a specifically stimulates H4K20 mono-methylation at TBE (TCF-binding element) via the histone methylase SET8. Additionally, SET8 is crucial for activation of the Wnt reporter gene and target genes in both mammalian cells and zebrafish. Furthermore, SET8 interacts with TCF4/LEF1 directly, and this interaction is regulated by Wnt3a. Therefore, we conclude that SET8 is a Wnt signaling mediator and is recruited by LEF1/TCF4 to regulate the transcription of Wnt-activated genes, possibly via H4K20 mono-methylation at the target gene promoters. Our findings also indicate that H4K20me-1 is a marker for gene transcription activation, at least in canonical Wnt signaling. This chip experiment was carried out to find out how many genes would be regulated by SET8 after Wnt3a CM stimulated HEK293 cells.
2010-12-29 | GSE24708 | GEO
Project description:Ubiquitin-mediated proteolysis play a significant role in various biological processes including transcription, DNA repair and cell cycle progression. The identification of Set8 and Set8b (a splice isoform) histone H4K20 methyl transferase as a substrate for the cullin-based ubiquitin ligase (CRL4-Cdt2) demonstrate that this pathway plays a significant role in promoting cell cycle progression, specifically promoting G2 progression. This study investigate the effect of failure to degrade Set8 in S-phase of the cell cycle via CRL4-Cdt2 on gene expression.
2010-09-18 | GSE24170 | GEO
Project description:E6 from high-risk strains of HPV is well known to transform cells by deregulating p53. We reported that in HPV transformed cell-lines E6 from high-risk HPV can recruit the USP46 deubiquitinase to substrates such as Cdt2 and stabilize the latter, and that USP46 is important for growth of HPV induced tumors in xenografts. Here we show that in cervical cancer biopsies the stabilization of Cdt2 in the HPV-induced cancers leads to the decrease of a CRL4-Cdt2 substrate, the histone H4K20 mono-methyltransferase Set8, and decrease in H4K20me1 or H4K20me3 that can be detected by immunohistochemistry. In HPV-transformed cancer cell lines in vitro, knockdown of E6 decreases Cdt2 and increases Set8. Co-knockdown of Set8 shows that some of the gene expression changes produced by E6 knockdown is due to the increase of Set8. EGFR and EGFR regulated genes were identified in this set of genes. Turning to the mechanism by which E6 stabilizes Cdt2, we find that a purified E6:USP46 complex has significantly more de-ubiquitinase activity in vitro than USP46 alone, demonstrating that E6 can directly interact with USP46 in the absence of other proteins and that it can substitute for the known activators of USP46, UAF1 and WDR20. Deletion mapping of Cdt2 shows that there are three discrete, but redundant, parts of the substrate that are essential for stabilization by E6: USP46. The helix-loop-helix region or the WD40 repeat driven beta-propeller structure of Cdt2 are dispensable for the stabilization implying that interaction with DDB1 (and the rest of the CRL4 complex) or with the substrate of the CRL4-Cdt2 E3 ligase is not necessary for E6:USP46 to interact with and stabilize Cdt2. The identification of 50 amino acid stretches in the 731 amino acid Cdt2 protein as being important for the stabilization by E6 underlines the specificity of the process. In summary, E6 activates the deubiquitinase activity of USP46, stabilizes Cdt2 utilizing multiple sites on Cdt2, and leads to degradation of Set8 and changes in gene-expression in HPV-transformed cells.
| S-EPMC8750077 | biostudies-literature
Project description:Mono-methylation of Lysine 20 of histone H4 (H4K20me1) is catalyzed by Set8 and thought to play important roles in many aspects of genome function that are mediated by H4K20me-binding proteins. We interrogated this model in a developing animal by comparing in parallel the transcriptomes of Set8null, H4K20R/A, and l(3)mbt mutant Drosophila melanogaster. We found that the gene expression profiles of H4K20A and H4K20R larvae are markedly different than Set8null larvae despite similar reductions in H4K20me1. Set8null mutant cells have a severely disrupted transcriptome and fail to proliferate in vivo, but these phenotypes are not recapitulated by mutation of H4K20 indicating that the developmental defects of Set8null animals are largely due to H4K20me1-independent effects on gene expression. Further, the H4K20me1 binding protein L(3)mbt is recruited to the transcription start sites of most genes independently of H4K20me even though genes bound by L(3)mbt have high levels of H4K20me1. Moreover, both Set8 and L(3)mbt bind to purified H4K20R nucleosomes in vitro. We conclude that gene expression changes in Set8null and H4K20 mutants cannot be explained by loss of H4K20me1 or L(3)mbt binding to chromatin, and therefore that H4K20me1 does not play a large role in gene expression.
2024-06-07 | GSE268821 | GEO
Project description:Mono-methylation of Lysine 20 of histone H4 (H4K20me1) is catalyzed by Set8 and thought to play important roles in many aspects of genome function that are mediated by H4K20me-binding proteins. We interrogated this model in a developing animal by comparing in parallel the transcriptomes of Set8null, H4K20R/A, and l(3)mbt mutant Drosophila melanogaster. We found that the gene expression profiles of H4K20A and H4K20R larvae are markedly different than Set8null larvae despite similar reductions in H4K20me1. Set8null mutant cells have a severely disrupted transcriptome and fail to proliferate in vivo, but these phenotypes are not recapitulated by mutation of H4K20 indicating that the developmental defects of Set8null animals are largely due to H4K20me1-independent effects on gene expression. Further, the H4K20me1 binding protein L(3)mbt is recruited to the transcription start sites of most genes independently of H4K20me even though genes bound by L(3)mbt have high levels of H4K20me1. Moreover, both Set8 and L(3)mbt bind to purified H4K20R nucleosomes in vitro. We conclude that gene expression changes in Set8null and H4K20 mutants cannot be explained by loss of H4K20me1 or L(3)mbt binding to chromatin, and therefore that H4K20me1 does not play a large role in gene expression.
2024-06-07 | GSE268819 | GEO
Project description:Mono-methylation of Lysine 20 of histone H4 (H4K20me1) is catalyzed by Set8 and thought to play important roles in many aspects of genome function that are mediated by H4K20me-binding proteins. We interrogated this model in a developing animal by comparing in parallel the transcriptomes of Set8null, H4K20R/A, and l(3)mbt mutant Drosophila melanogaster. We found that the gene expression profiles of H4K20A and H4K20R larvae are markedly different than Set8null larvae despite similar reductions in H4K20me1. Set8null mutant cells have a severely disrupted transcriptome and fail to proliferate in vivo, but these phenotypes are not recapitulated by mutation of H4K20 indicating that the developmental defects of Set8null animals are largely due to H4K20me1-independent effects on gene expression. Further, the H4K20me1 binding protein L(3)mbt is recruited to the transcription start sites of most genes independently of H4K20me even though genes bound by L(3)mbt have high levels of H4K20me1. Moreover, both Set8 and L(3)mbt bind to purified H4K20R nucleosomes in vitro. We conclude that gene expression changes in Set8null and H4K20 mutants cannot be explained by loss of H4K20me1 or L(3)mbt binding to chromatin, and therefore that H4K20me1 does not play a large role in gene expression.
2024-06-07 | GSE268820 | GEO
Project description:In mammalian cells, SET8 mediated Histone H4 Lys 20 monomethylation (H4K20me1) has been implicated in regulating mitotic condensation, DNA replication, DNA damage response, and gene expression. Here we show SET8, the only known enzyme for H4K20me1 is post-translationally poly ADP-ribosylated by PARP1 on lysine residues. PARP1 interacts with SET8 in a cell cycle- dependent manner. Poly ADP-ribosylation on SET8 renders it catalytically compromised and it undergoes degradation via ubiquitylation pathway. Knockdown of PARP1 shifted the relative dynamic equilibrium of H4K20me2 to H4K20me3 in cells. Overexpression or knockdown of PARP1 led to aberrant H4K20me1 domains genome-wide, impacting Wnt signaling pathways genes and transcription factor binding site enrichment. Therefore, SET8 mediated chromatin remodeling in mammalian cells are influenced by poly ADP-ribosylation by PARP1.
2022-11-17 | GSE188744 | GEO