Development of zebrafish medulloblastoma model by TALENs induced somatic gene inactivation methods
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ABSTRACT:
ORGANISM(S): Danio rerio
PROVIDER: GSE92260 | GEO | 2017/10/31
SECONDARY ACCESSION(S): PRJNA357051
REPOSITORIES: GEO
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Project description:Disruption of miRNA Sequence by TALENs and CRISPR/Cas9 Induces Varied Length of miRNA Production [TALENs]
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Project description:Mechanisms of neuroendocrine tumor (NET) proliferation are poorly understood and therapies that effectively control NET progression and metastatic disease are limited. We found amplification of a putative oncogene, RABL6A, in primary human pancreatic NETs (PNETs) that correlated with high level RABL6A protein expression. Consistent with those results, stable silencing of RABL6A in cultured BON-1 PNET cells revealed that it is essential for their proliferation and survival. Cells lacking RABL6A predominantly arrested in G1 phase with a moderate mitotic block. Pathway analysis of microarray data suggested activation of the p53 and retinoblastoma (Rb1) tumor suppressor pathways in the arrested cells. Specific inactivation of p53 had no effect on the RABL6A knockdown phenotype, indicating RABL6A functions independent of p53 in this setting. By comparison, Rb1 inactivation restored G1 to S phase progression in RABL6A knockdown cells although it was insufficient to override the mitotic arrest and cell death caused by RABL6A loss. Thus, RABL6A promotes G1 progression in PNET cells by inactivating Rb1, an established suppressor of PNET proliferation and development. This work identifies RABL6A as a new negative regulator of Rb1 that is essential for PNET proliferation and survival. We suggest RABL6A is a new potential biomarker and target for anticancer therapy in PNET patients. Total RNA obtained from human BON-1 PNET cells with RABL6A shRNA knockdown compared to BON-1 cells expressing control vector.
2014-07-01 | E-GEOD-53778 | biostudies-arrayexpress
Project description:Development of zebrafish medulloblastoma model by TALENs induced somatic gene inactivation methods
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Project description:Goal is to analyze the expression profile of cells shortly after induction of mitochondrial double stranded breaks. ARPE-19 cells are transfected with constructs coding for mitochondrial TALENs, either active or dead. These constructs target two different locations of mtDNA (ATP8 and Dloop). Gene expression is then compared between untransfected and transfected cells (either active or dead TALENs), or between active TALEN (cells with mitochondrial double stranded breaks) and dead TALEN (expressing the same protein without cutting the mitochondrial genome)
2021-02-24 | GSE164979 | GEO
Project description:Mechanisms of neuroendocrine tumor (NET) proliferation are poorly understood and therapies that effectively control NET progression and metastatic disease are limited. We found amplification of a putative oncogene, RABL6A, in primary human pancreatic NETs (PNETs) that correlated with high level RABL6A protein expression. Consistent with those results, stable silencing of RABL6A in cultured BON-1 PNET cells revealed that it is essential for their proliferation and survival. Cells lacking RABL6A predominantly arrested in G1 phase with a moderate mitotic block. Pathway analysis of microarray data suggested activation of the p53 and retinoblastoma (Rb1) tumor suppressor pathways in the arrested cells. Specific inactivation of p53 had no effect on the RABL6A knockdown phenotype, indicating RABL6A functions independent of p53 in this setting. By comparison, Rb1 inactivation restored G1 to S phase progression in RABL6A knockdown cells although it was insufficient to override the mitotic arrest and cell death caused by RABL6A loss. Thus, RABL6A promotes G1 progression in PNET cells by inactivating Rb1, an established suppressor of PNET proliferation and development. This work identifies RABL6A as a new negative regulator of Rb1 that is essential for PNET proliferation and survival. We suggest RABL6A is a new potential biomarker and target for anticancer therapy in PNET patients.
2014-07-01 | GSE53778 | GEO
Project description:Primitive neuroectodermal tumors of the central nervous system (CNS PNETs) are highly aggressive, poorly differentiated embryonal tumors occurring predominantly in young children. Using DNA methylation and gene expression profiling we have demonstrated that a significant proportion of institutionally diagnosed CNS PNETs display molecular profiles indistinguishable from those of various other well defined CNS tumor entities, facilitating diagnosis and appropiate therapy for children with these tumors. From the remaining fraction of CNS PNETs, we have identified four distinct new CNS tumor entities extending to other neuroepithelial tumors, each associated with a recurrent genetic alteration and particular histopathological and clinical features. These molecular entities, designated âCNS Neuroblastoma with FOXR2 activation (CNS NB FOXR2)â, âCNS Ewing sarcoma family tumor with CIC alteration (CNS EFT CIC)â, âCNS high grade neuroepithelial tumor with MN1 alteration (CNS HGNET MN1)â, and âCNS high grade neuroepithelial tumor with BCOR alteration (CNS HGNET BCOR)â, will enable meaningful clinical trials and the development of therapeutic strategies for patients affected by these poorly differentiated CNS tumors. 182 brain tumor samples were selected for RNA extraction and hybridization on Affymetrix Affymetrix Human Genome U133 Plus 2.0 Arrays.
2016-02-25 | E-GEOD-73038 | biostudies-arrayexpress
Project description:Purpose: MicroRNAs (miRNAs) have been recognized as crucial players in plant growth, development, production and responses to biotic and abiotic stresses, while only limited number of mutants can be used to dissect their roles. Here we used TALENs (Transcription Activator-Like Effector Nucleases) and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) to introduce base pair mutations in MIRNA genes, resulting in creation of miRNA mutants and elucidating the effect of mutations on the miRNA biogenesis in rice and Arabidopsis. Methods: Total RNAs were prepared using a Trizol reagent-based method. The plant materials were homogenized and reconstituted in Trizol solution. The solution in turn was extracted by chloroform. Isopropanol was used to precipitate the total RNA. The whole process was performed under RNase free environment. DEPC-treated water was used to dissolve RNA. The total RNA underwent miRNA isolation using Qiagen miRNA purification kit according to the manufacturer’s manual. Small RNA libraries were made using 5 µg total RNA with NEBNext Small RNA library Prep Set, as described, and sequenced on the Illumina HiSeq 2500 platform.
2019-04-25 | GSE130239 | GEO
Project description:Background Retinoblastoma is a pediatric eye cancer associated with RB1 loss or MYCN amplification (RB1+/+MYCNA). There are controversies concerning the existence of molecular subtypes within RB1-/- retinoblastoma. To test whether these molecular subtypes exist, we performed molecular profiling. Methods Genome-wide mRNA expression profiling was performed on 76 primary human retinoblastomas. Expression profiling was complemented by genome-wide DNA profiling and clinical, histopathological, and ex vivo drug sensitivity data. Findings RNA and DNA profiling identified major variability between retinoblastomas. While gene expression differences between RB1+/+MYCNA and RB1-/- tumors seemed more dichotomous, differences within the RB1-/- tumors were gradual. Tumors with high expression of a photoreceptor gene signature were highly differentiated, smaller in volume and diagnosed at younger age compared to tumors with low photoreceptor signature expression. Tumors with lower photoreceptor expression showed increased expression of genes involved in M-phase and mRNA and ribosome synthesis and increased frequencies of somatic copy number alterations. Interpretation Molecular, clinical and histopathological differences between RB1-/- tumors are best explained by tumor progression, reflected by a gradual loss of differentiation and photoreceptor expression signature. Since copy number alterations were more frequent in tumors with less photoreceptorness, genomic alterations might be drivers of tumor progression. Fresh frozen material from 76 primary human retinoblastoma samples were profiled with Affymetrix human genome u133 plus 2.0 PM microarray
2015-07-01 | E-GEOD-59983 | biostudies-arrayexpress
Project description:Central nervous system primitive neuroectodermal tumors (CNS PNET) and medulloblastomas are both embryonal tumors that predominantly occur in children. We used microarrays to analyse a cohort of CNS PNETs and medulloblastomas to identify gene expression related to tumor subgroups. RNA extracted from 23 frozen tumor samples, plus a commercial fetal brain sample, was analysed using Affymetrix U133 plus 2.0 arrays. Tumour subgroups, based on gene expression, were identified using clustering methods.
2013-05-01 | E-GEOD-19404 | biostudies-arrayexpress
Project description:The canonical model of small cell lung cancer (SCLC) depicts tumors arising from dual inactivation of TP53 and RB1. However, many genomic studies have persistently identified tumors with no RB1 mutations. Here, we examined RB1 protein expression and function in SCLC. RB1 expression was examined by immunohistochemical analysis of 62 human SCLC tumors. These studies showed that ~14% of SCLC tumors expressed abundant RB1 protein, which is associated with neuroendocrine (NE) gene expression and is enriched in YAP1 expression, but no other lineage proteins that stratify SCLC. SCLC cells and xenograft tumors with RB1 protein expression were sensitive to growth inhibition by the CDK4/6 inhibitor palbociclib, and this inhibition was shown to be dependent on RB1 expression by CRISPR knockout. Furthermore, a patient with biopsy-validated wt RB1 SCLC who received the CDK4/6 inhibitor abemaciclib demonstrated a dramatic decrease in mutant TP53 ctDNA allelic fraction from 62.1% to 0.4% and decreased tumor mass on CT scans. Importantly, immunohistochemistry of the diagnostic biopsy specimen showed RB1 positivity. Finally, we identified a transcriptomics-based RB1 loss-of-function signature that discriminates between SCLC cells with or without RB1 protein expression and validated it in the patient who was responsive to abemaciclib, suggesting its potential use to predict CDK4/6 inhibitor response in SCLC patients. Our study demonstrates that RB1 protein is an actionable target in a subgroup of SCLC, a cancer that exhibits no currently targetable mutations.
2022-11-14 | GSE217646 | GEO