Neuroblastoma Cells Depend on CSB for Faithful Execution of Cytokinesis and Survival.
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ABSTRACT:
SUBMITTER: Paccosi E
PROVIDER: S-EPMC8465547 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The number of long-term survivors of high-risk neuroblastoma remains discouraging, with 10-year survival as low as 20%, despite decades of considerable international efforts to improve outcome. Major obstacles remain and include managing resistance to induction therapy, which causes tumor progression and early death in high-risk patients, and managing chemotherapy-resistant relapses, which can occur years after the initial diagnosis. Identifying and validating novel therapeutic targets is essential to improve treatment. Delineating and deciphering specific functions of single histone deacetylases in neuroblastoma may support development of targeted acetylome-modifying therapeutics for patients with molecularly defined high-risk neuroblastoma profiles. We show here that HDAC11 depletion in MYCN-driven neuroblastoma cell lines strongly induces cell death, mostly mediated by apoptotic programs. Genes necessary for mitotic cell cycle progression and cell division were most prominently enriched in at least two of three time points in whole-genome expression data combined from two cell systems, and all nine genes in these functional categories were strongly repressed, including CENPA, KIF14, KIF23 and RACGAP1. Enforced expression of one selected candidate, RACGAP1, partially rescued the induction of apoptosis caused by HDAC11 depletion. High-level expression of all nine genes in primary neuroblastomas significantly correlated with unfavorable overall and event-free survival in patients, suggesting a role in mediating the more aggressive biological and clinical phenotype of these tumors. Our study identified a group of cell cycle-promoting genes regulated by HDAC11, being both predictors of unfavorable patient outcome and essential for tumor cell viability. The data indicate a significant role of HDAC11 for mitotic cell cycle progression and survival of MYCN-amplified neuroblastoma cells, and suggests that HDAC11 could be a valuable drug target.
| S-EPMC5386552 | biostudies-literature
Project description:The number of long-term survivors of high-risk neuroblastoma remains discouraging, with 10-year survival as low as 20%, despite decades of considerable international efforts to improve outcome. Major obstacles remain and include managing resistance to induction therapy, which causes tumor progression and early death in high-risk patients, and managing chemotherapy-resistant relapses, which can occur years after the initial diagnosis. Identifying and validating novel therapeutic targets is essential to improve treatment. Delineating and deciphering specific functions of single histone deacetylases in neuroblastoma may support development of targeted acetylome-modifying therapeutics for patients with molecularly defined high-risk neuroblastoma profiles. We show here that HDAC11 depletion in MYCN-driven neuroblastoma cell lines strongly induces cell death, mostly mediated by apoptotic programs. Genes necessary for mitotic cell cycle progression and cell division were most prominently enriched in at least two of three time points in whole-genome expression data combined from two cell systems, and all nine genes in these functional categories were strongly repressed, including CENPA, KIF14, KIF23 and RACGAP1. Enforced expression of one selected candidate, RACGAP1, partially rescued the induction of apoptosis caused by HDAC11 depletion. High-level expression of all nine genes in primary neuroblastomas significantly correlated with unfavorable overall and event-free survival in patients, suggesting a role in mediating the more aggressive biological and clinical phenotype of these tumors. Our study identified a group of cell cycle-promoting genes regulated by HDAC11, being both predictors of unfavorable patient outcome and essential for tumor cell viability. The data indicates a significant role of HDAC11 for mitotic cell cycle progression and survival of MYCN-amplified neuroblastoma cells, and suggests that HDAC11 could be a valuable drug target.
2017-02-16 | GSE77080 | GEO
Project description:To identify mutations that cause defects in mitosis, a collection of mutants in Saccharomyces cerevisiae was screened by a rapid visual assay for abnormal chromosome segregation. From this screen we identified one mutation, pds1-1 that was independently identified in an alternative screen for mutants that exhibit inviability after transient exposure to nocodazole and precocious disassociation of sister chromatids (Guacci, V., A. Yamamoto, A. Strunnikov, J. Kingsbury, E. Hogan, P. Meluh, and D. Koshland. 1993. CSH Symp. Quant. Biol. 58:677-685; Yamamoto, T.J., G. Li, B. Schaar, I. Szilak, and D.W. Cleveland. 1992. Nature (Lond.). 359:536-539). At 23 degrees C pds1-1 mutants exhibit frequent cell death and a 300-fold increase in chromosome loss compared to wild type. At 37 degrees C pds1-1 cells fail to elongate their spindles during anaphase. This spindle defect of pds1 mutants results from a temperature-sensitive step that occurs around the G1/S boundary about the time of spindle assembly. In the absence of spindle elongation pds1 mutants undergo cytokinesis, leading to the missegregation of both chromosomes and spindle pole bodies. After abnormal cell division pds1-1 mutants also initiate new rounds of DNA replication, spindle pole body duplication, and bud formation. Thus, in the pds1-1 mutant at 37 degrees C, cell cycle progression is uncoupled from the completion of anaphase. A pds1 deletion allele has similar phenotypes to the original allele. Taken together these results suggest that Pds1 protein plays an important role in chromosome segregation at 23 degrees C and an essential role for this process at 37 degrees C. The PDS1 gene encodes a novel 42-kD nuclear protein that has both basic and acidic domains. The level of PDS1 mRNA varies with the cell cycle with maximal accumulation around the G1/S boundary. The stability of Pds1 protein also appears to change during the cell cycle as overproduced Pds1p is stable in S and M but degraded in early G1. Therefore, expression of Pds1p is regulated apparently both transcriptionally and postranslationally during the cell cycle. The phenotypes of pds1 mutants and expression pattern of Pds1p are discussed in the context of other spindle-defective mutants and the knowledge that Pds1 protein is an inhibitor of anaphase (Yamamoto, T.J., G. Li, B. Schaar, I. Szilak, and D.W. Cleveland. 1992. Nature (Lond.). 359:536-539).
| S-EPMC2120769 | biostudies-other
Project description:In response to perturbation of the cell division machinery fission yeast cells activate regulatory networks that ensure the faithful completion of cytokinesis. For instance, when cells are treated with drugs that impede constriction of the actomyosin ring (low doses of Latrunculin A, for example) these networks ensure that cytokinesis is complete before progression into the subsequent mitosis. Here, we identify three previously uncharacterized genes, hif2, set3, and snt1, whose deletion results in hyper-sensitivity to LatA treatment and in increased rates of cytokinesis failure. Interestingly, these genes are orthologous to TBL1X, MLL5, and NCOR2, human genes that encode components of a histone deacetylase complex with a known role in cytokinesis. Through co-immunoprecipitation experiments, localization studies, and phenotypic analysis of gene deletion mutants, we provide evidence for an orthologous complex in fission yeast. Furthermore, in light of the putative role of the complex in chromatin modification, together with our results demonstrating an increase in Set3p levels upon Latrunculin A treatment, global gene expression profiles were generated. While this analysis demonstrated that the expression of cytokinesis genes was not significantly affected in set3Δ backgrounds, it did reveal defects in the ability of the mutant to regulate genes with roles in the cellular response to stress. Taken together, these findings support the existence of a conserved, multi-protein complex with a role in promoting the successful completion of cytokinesis.
| S-EPMC3275627 | biostudies-literature
Project description:Multiple myeloma (MM) cells suffer from baseline proteotoxicity as the result of an imbalance between the load of misfolded proteins awaiting proteolysis and the capacity of the ubiquitin-proteasome system to degrade them. This intrinsic vulnerability is at the base of MM sensitivity to agents that perturb proteostasis, such as proteasome inhibitors (PIs), the mainstay of modern-day myeloma therapy. De novo and acquired PI resistance are important clinical limitations that adversely affect prognosis. The molecular mechanisms underpinning PI resistance are only partially understood, limiting the development of drugs that can overcome it. The transcription factor NRF1 is activated by the aspartic protease DNA damage inducible 1 homolog 2 (DDI2) upon proteasome insufficiency and governs proteasome biogenesis. In this article, we show that MM cells exhibit baseline NRF1 activation and are dependent upon DDI2 for survival. DDI2 knockout (KO) is cytotoxic for MM cells, both in vitro and in vivo. Protein structure-function studies show that DDI2 KO blocks NRF1 cleavage and nuclear translocation, causing impaired proteasome activity recovery upon irreversible proteasome inhibition and, thereby, increasing sensitivity to PIs. Add-back of wild-type, but not of catalytically dead DDI2, fully rescues these phenotypes. We propose that DDI2 is an unexplored promising molecular target in MM by disrupting the proteasome stress response and exacerbating proteotoxicity.
| S-EPMC8791580 | biostudies-literature
Project description:Mutations in the CSA and CSB genes are causative of Cockayne syndrome neurological disorder. Since the identification of the indispensable functions of these two proteins in transcription-coupled repair and restoring RNA synthesis following DNA damage, the paradoxical milder clinical spectrum in CS-A patients has been puzzling. In this study we compared the effect of a CSA and a CSB defect at the levels of the cell and the intact organism. We showed that CSA-deficient zebrafish embryos exhibited modest hypersensitive to UV damage than CSB depletion. We found that loss of CSA can effectively release aggregation of mutant crystallin proteins in vitro. We described the distinct effect of CSA and CSB on neuritogenesis and elucidated the differentiated gene expression pathways regulated by these two proteins. Our data demonstrate convergent and divergent roles for CSA and CSB in DNA repair and transcription regulation and provide explanations for the reported differences between CS-A and CS-B patients.
2019-03-29 | GSE129002 | GEO
Project description:Neuroblastoma (NB) is the most common extracranial solid tumour in children. NB is highly heterogeneous and is comprised of a mixture of neuroblastic cancer cells and stromal cells. We previously reported that N-type cells (neuroblastic cells) and S-type cells (substrate-adherent cells) in the SK-N-SH cell line shared almost identical genetic backgrounds. Sublines of N- and S-type cells were isolated from an early passage (P35) of SK-N-SH. Sequencing analysis revealed that all sublines harboured the anaplastic lymphoma kinase (ALK) F1174L mutation, indicating that they were tumour derived. Surprisingly, over 74% resembled S-type cells. In coculture experiments, S-type cells protected N-type cells from apoptosis induced by the oncogenic ALK inhibitor TAE684. Western blotting analyses showed that ALK, protein kinase A (AKT) and STAT3 signalling were stimulated in the cocultures. Furthermore, the conditioned medium from S-type cells activated these downstream signalling molecules in the N-type cells. The activation of STAT3 in the N-type cells was ALK-independent, while AKT was regulated by the ALK activation status. To identify the responsible soluble factors, we used a combination of transcriptomic and proteomic analysis and found that plasminogen activator inhibitor 1, secreted protein acidic and cysteine rich, periostin and galectin-1 were potential mediators of STAT3 signalling. The addition of recombinant proteins to the tumour cells treated with the ALK inhibitor partially enhanced cell viability. Overall, the tumour-derived S-type cells prevented apoptosis in the N-type cells via ALK-independent STAT3 activation triggered by secreted factors. The inhibition of these factors in combination with ALK inhibition could provide a new direction for targeted therapies to treat high-risk NB.
| S-EPMC8334291 | biostudies-literature
Project description:Cockayne syndrome is a segmental progeria most often caused by mutations in the CSB gene encoding a SWI/SNF-like ATPase required for transcription-coupled DNA repair (TCR). Over 43 Mya before marmosets diverged from humans, a piggyBac3 (PGBD3) transposable element integrated into intron 5 of the CSB gene. As a result, primate CSB genes now generate both CSB protein and a conserved CSB-PGBD3 fusion protein in which the first 5 exons of CSB are alternatively spliced to the PGBD3 transposase. We show by microarray analysis that expression of the fusion protein alone in CSB-null UV-sensitive syndrome cells (UVSS1KO) cells induces an interferon-like response that resembles both the innate antiviral response and the prolonged interferon response normally maintained by unphosphorylated STAT1 (U-STAT1); moreover, as might be expected based on conservation of the fusion protein, this potentially cytotoxic interferon-like response is largely reversed by coexpression of functional CSB protein. Interestingly, expression of CSB and the CSB-PGBD3 fusion protein together, but neither alone, upregulates the insulin growth factor binding protein IGFBP5 and downregulates IGFBP7, suggesting that the fusion protein may also confer a metabolic advantage, perhaps in the presence of DNA damage. Finally, we show that the fusion protein binds in vitro to members of a dispersed family of 900 internally deleted piggyBac elements known as MER85s, providing a potential mechanism by which the fusion protein could exert widespread effects on gene expression. Our data suggest that the CSB-PGBD3 fusion protein is important in both health and disease, and could play a role in Cockayne syndrome.
2014-03-20 | GSE56049 | GEO
Project description:BackgroundThe precision of the metaphase-anaphase transition ensures stable genetic inheritance. The spindle checkpoint blocks anaphase onset until the last chromosome biorients at metaphase plate, then the bonds between sister chromatids are removed and disjoined chromatids segregate to the spindle poles. But, how sister separation is triggered is not fully understood.Principal findingsWe identify PIASgamma as a human E3 sumo ligase required for timely and efficient sister chromatid separation. In cells lacking PIASgamma, normal metaphase plates form, but the spindle checkpoint is activated, leading to a prolonged metaphase block. Sister chromatids remain cohered even if cohesin is removed by depletion of hSgo1, because DNA catenations persist at centromeres. PIASgamma-depleted cells cannot properly localize Topoisomerase II at centromeres or in the cores of mitotic chromosomes, providing a functional link between PIASgamma and Topoisomerase II.ConclusionsPIASgamma directs Topoisomerase II to specific chromosome regions that require efficient removal of DNA catenations prior to anaphase. The lack of this activity activates the spindle checkpoint, protecting cells from non-disjunction. Because DNA catenations persist without PIASgamma in the absence of cohesin, removal of catenations and cohesin rings must be regulated in parallel.
| S-EPMC1762334 | biostudies-literature
Project description:Cockayne syndrome is a segmental progeria most often caused by mutations in the CSB gene encoding a SWI/SNF-like ATPase required for transcription-coupled DNA repair (TCR). Over 43 Mya before marmosets diverged from humans, a piggyBac3 (PGBD3) transposable element integrated into intron 5 of the CSB gene. As a result, primate CSB genes now generate both CSB protein and a conserved CSB-PGBD3 fusion protein in which the first 5 exons of CSB are alternatively spliced to the PGBD3 transposase. We show by microarray analysis that expression of the fusion protein alone in CSB-null UV-sensitive syndrome cells (UVSS1KO) cells induces an interferon-like response that resembles both the innate antiviral response and the prolonged interferon response normally maintained by unphosphorylated STAT1 (U-STAT1); moreover, as might be expected based on conservation of the fusion protein, this potentially cytotoxic interferon-like response is largely reversed by coexpression of functional CSB protein. Interestingly, expression of CSB and the CSB-PGBD3 fusion protein together, but neither alone, upregulates the insulin growth factor binding protein IGFBP5 and downregulates IGFBP7, suggesting that the fusion protein may also confer a metabolic advantage, perhaps in the presence of DNA damage. Finally, we show that the fusion protein binds in vitro to members of a dispersed family of 900 internally deleted piggyBac elements known as MER85s, providing a potential mechanism by which the fusion protein could exert widespread effects on gene expression. Our data suggest that the CSB-PGBD3 fusion protein is important in both health and disease, and could play a role in Cockayne syndrome. 12 samples total; 4 gene expression conditions in triplicate; 1 condition is a tag-only negative control
2014-03-20 | E-GEOD-56049 | biostudies-arrayexpress