SPT6-driven error-free DNA repair safeguards genomic stability of glioblastoma cancer stem-like cells
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE125621 | GEO | 2020/07/17
REPOSITORIES: GEO
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Project description:SPT6, encoded by the SUPT6H in humans and Supt6 in mice, respectively, is a conserved histone chaperone that interacts with RNA polymerase II and participates in transcription elongation. However, the question of how SPT6 comes into play in transcriptional activation upon signaling, particularly in mammalian cells, has remained elusive. We investigated the contribution of SPT6 to interferon beta (IFNbeta) induced transcription in mouse NIH3T3 cells. IFNbeta triggers rapid and high level transcription of many IFN-stimulated genes (ISGs). We report here that SPT6 is recruited to ISGs after IFN stimulation. This recruitment was dependent on the interaction with the methyltransferase, NSD2. Further, siRNA-based SPT6 knockdown reduced levels of ISG activation. RNA-Seq analysis showed that SPT6 knockdown diminished about 50% of ISGs whose induction levels were higher than those unaffected by SPT6 knockdown. Under the tested conditions, SPT6 knockdown did not measurably change expression of constitutively expressed genes. This report highlights that SPT6 is recruited in a stimulus-dependent manner and elicits a major impact on signal induced transcription.
2018-05-01 | GSE97302 | GEO
Project description:Glioblastoma cancer-stem like cells (GSCs) display marked resistance to ionizing radiation (IR), a standard of care for glioblastoma patients. Mechanisms underpinning radio-resistance of GSCs remain largely unknown. Chromatin state and the accessibility of DNA lesions to DNA repair machineries are crucial for the maintenance of genomic stability. Understanding the functional impact of chromatin remodeling on DNA repair in GSCs may lay the foundation for advancing the efficacy of radio-sensitizing therapies. Here, we present the results of a high-content siRNA microscopy screen, revealing the transcriptional elongation factor SPT6 to be critical for the genomic stability and self-renewal of GSCs. Mechanistically, SPT6 transcriptionally up-regulates BRCA1 and thereby drives an error-free DNA repair in GSCs. SPT6 loss impairs the self-renewal, genomic stability and tumor initiating capacity of GSCs. Collectively, our results provide mechanistic insights into how SPT6 regulates DNA repair and identify SPT6 as a putative therapeutic target in glioblastoma.
| S-EPMC7501306 | biostudies-literature
Project description:Effect of PAAF1 and Spt6 knock-down on gene expression. Total RNAs were extracted from HeLa-LTR-Luc cells in which PAAF1 or Spt6 had been knock down by a specific siRNA or HeLa-LTR-Luc cells that had been transfected with a control siRNA (“si Neg”). Differentially expressed genes in PAAF1 knock-down cells were identified by microarray. Two biological replicates (A / B) in 3 experimental conditions (si-Neg, si-PAAF1, si-Spt6) were analyzed in a single channel experiment.
2012-05-28 | E-GEOD-32033 | biostudies-arrayexpress
Project description:Effect of PAAF1 and Spt6 knock-down on gene expression. Total RNAs were extracted from HeLa-LTR-Luc cells in which PAAF1 or Spt6 had been knock down by a specific siRNA or HeLa-LTR-Luc cells that had been transfected with a control siRNA (“si Neg”). Differentially expressed genes in PAAF1 knock-down cells were identified by microarray.
2012-05-29 | GSE32033 | GEO
Project description:H3K4me3 plays a critical role in the activation-induced cytidine deaminase (AID)-induced DNA cleavage of switch (S) regions in the immunoglobulin heavy chain (IgH) locus during class-switch recombination (CSR). The histone chaperone complex facilitates chromatin transcription (FACT) is responsible for forming H3K4me3 at AID target loci. Histone chaperone suppressor of Ty6 (Spt6) also participates in regulating H3K4me3 for CSR and for somatic hypermutation (SHM) in AID target loci. H3K4me3 loss was correlated with defects in AID-induced DNA breakage and reduced mutation frequencies in IgH loci, in both S and variable regions, and in non-IgH loci, such as metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and small nucleolar RNA host gene 3 (SNHG3). Global gene expression analysis revealed that Spt6 can act as both a positive and negative transcriptional regulator in B cells, affecting approximately 5% of the genes that includes suppressor of Ty4 (Spt4) and AID. Interestingly, Spt6 regulates CSR and AID expression through two distinct histone modification pathways, H3K4me3 and H3K36me3, respectively. Spt6 is a unique histone chaperone, capable of regulating the histone epigenetic state of both AID targets and the AID locus. CH12F3-2A cells were transfected with control and Spt6 siRNAs; 24h later, cells were stimulated with CIT to induce CSR. Total RNA was extracted from control and Spt6 siRNA treated cells for mRNA expression profiling.
2013-02-01 | E-GEOD-35334 | biostudies-arrayexpress
Project description:Spt6 is a histone chaperone that associates with RNA polymerase II and deposits nucleosomes in the wake of transcription. Although Spt6 has an essential function in nucleosome deposition, it is not known whether this function is regulated by post-translational modification. Here, we report that casein kinase II (CKII) phosphorylation of Spt6 directs nucleosome reassembly at the 5’ ends of a broad range of genes to prevent aberrant antisense transcription and enforce transcriptional directionality. Mechanistically, we show that interaction of Spt6 with Spn1 – a constitutive binding partner required for chromatin reassembly and full recruitment of Spt6 to genes is positively regulated by CKII phosphorylation of Spt6. Together, our study defines a previously unknown function for CKII phosphorylation in transcription, and further, highlights the importance of post-translational modification as a mechanism to fine-tune the functions of histone chaperones.
2018-11-30 | GSE122620 | GEO
Project description:SPT6 is a histone chaperone that tightly binds RNA polymerase II (POL2) during transcription elongation. However, its primary role in POL2 transcription is uncertain. We used targeted protein degradation to rapidly deplete SPT6 in human cells and analyzed defects in POL2 behavior by a multi-omics approach and mathematical modeling. Our data indicate that SPT6 is a crucial factor for POL2 processivity and is therefore required for the productive transcription of protein-coding genes. Unexpectedly, SPT6 also has a vital role in POL2 termination, as acute depletion induced readthrough transcription for thousands of genes. Long-term depletion of SPT6 induced cryptic intragenic transcription, as observed earlier in yeast. However, this phenotype was not observed upon acute SPT6 depletion and therefore can be attributed to accumulated epigenetic perturbations in the absence of SPT6. In conclusion, targeted protein degradation of SPT6 allowed the temporal discrimination of its function as an epigenetic safeguard and POL2 elongation factor.
2021-06-28 | PXD025243 | Pride
Project description:The histone chaperone Spt6 is involved in promoting elongation of RNA polymerase II (RNAPII), maintaining chromatin structure, regulating co-transcriptional histone modifications, and controlling mRNA processing. These diverse functions of Spt6 are partly mediated through its interactions with RNAPII and other factors in the transcription elongation complex. In this study, we used mass spectrometry to characterize the differences in RNAPII interacting factors between wild-type cells and those depleted for Spt6, leading to the identification of proteins that depend on Spt6 for their interaction with RNAPII. In all, eight samples were processed - four genotypes (1. SPT6, RPB3-untagged; 2. SPT6, RPB3-tagged; 3. spt6-1004, RPB3-untagged; 4. spt6-1004; RPB3-tagged) in biological duplicates.
2021-12-05 | PXD027708 | Pride
Project description:SPT6-driven error-free DNA repair safeguards genomic stability of glioblastoma cancer stem-like cells
| PRJNA516926 | ENA
Project description:In vitro studies identified various factors including P-TEFb, SEC, SPT6, PAF1, DSIF, and NELF functioning at different stages of transcription elongation driven by RNA polymerase II (RNA Pol II). What remains unclear is how these factors cooperatively regulate pause/release and productive elongation in the context of living cells. Using an acute 5 protein-depletion approach, prominent release and a subsequent increase in mature transcripts, whereas long genes fail to yield mature transcripts due to a loss of processivity. Mechanistically, loss of SPT6 results in loss of PAF1 complex (PAF1C) from RNA Pol II, leading to NELF-bound RNA Pol II release into the gene bodies. Furthermore, SPT6 and/or PAF1 depletion impairs heat shock-induced pausing, pointing to a role for SPT6 in regulating RNA Pol II pause/release through the recruitment of PAF1C during the early elongation.
2022-08-09 | PXD033771 | Pride