PAX3-FOXO1 drives targetable cell state-dependent metabolic vulnerabilities in rhabdomyosarcoma
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE286950 | GEO | 2025/09/02
REPOSITORIES: GEO
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Project description:Rhabdomyosarcomas (RMS) are rare but very aggressive tumours of childhood that arise as a consequence of regulatory disruption of the growth and differentiation pathways of myogenic precursor cells. Based on morphology, two major RMS subtypes can be identified: embryonal RMS (ERMS) and alveolar (ARMS). To better understand the global function of miRNA in RMS, we analyzed the expression profile of 8 different RMS cell lines (3 PAX3/FOXO1 positive and 5 PAX3/FOXO1 negative RMS) using the mirVana miRNA Probe Set V1 (Ambion). The miRNA microarray platform was able to distinguish PAX3/FOXO1 positive (RH4, RH30, RH28) and negative RMS (RD, CCA, SMS-CTR, RH36, RH18) cell lines through the expression pattern of about 90 miRNAs. Since the translocation positive RMS patients fared worse than the negative counterpart our results demonstrated the potential of miRNA expression profiling to classify different RMS subtypes, in agreement to previous gene expression studies, and set the basis for a further functional characterization of selected miRNAs implicated in RMS pathogenesis and in the different clinical behaviour and aggressiveness of the two RMS subtypes. We decided to study miRNAs with the greatest difference in expression between PAX3/FOXO1 positive and negative RMS:miR-23a, miR-27a and miR-199a.
2015-05-04 | GSE52677 | GEO
Project description:To investigate the role of TGF-M-NM-21-regulated miRNAs in the progression of RMS,we performed comprehensive miRMA microarray analysis on RNA derived from typical RMS cell lines and TGF-M-NM-21 knock-down cell lines. We identified a novel set of TGF-M-NM-21-related miRNAs. Total RNA was isolated from TGF-M-NM-21-knock down rhabdmyosarcoma cell lines and controls. Three-condition experiment: shRNA-TGF-M-NM-21/RD cells vs. shRNA-Control/RD cells, shRNA-TGF-M-NM-21/SMS-CTR cells vs. shRNA-Control/ SMS-CTR cells, and shRNA-TGF-M-NM-21/RH28 cells vs. shRNA-Control/RH28 cells. Biological replicates: 1 RD cells stably transfected with shRNA-TGF-M-NM-21- pSUPER gfp-neo, 1SMS-CTR cells stably transfected with shRNA-TGF-M-NM-21- pSUPER gfp-neo, 1RH28 cells stably transfected with shRNA-TGF-M-NM-21- pSUPER gfp-neo, 1RD cells stably transfected with shRNA-Control- pSUPER gfp-neo, 1SMS-CTR cells stably transfected with shRNA-Control- pSUPER gfp-neo, and 1RH28 cells stably transfected with shRNA-Control- pSUPER gfp-neo, independently grown and harvested. One replicate per array.
2012-09-13 | E-GEOD-40843 | biostudies-arrayexpress
Project description:Background: Alveolar rhabdomyosarcoma (ARMS) has a high propensity to metastasize, leading to its aggressiveness and a poor survival rate among those with the disease. More than 80% of aggressive ARMSs harbor a PAX3-FKHR fusion transcription factor, which regulates cell migration and promotes metastasis, most likely by regulating the fusion protein's transcriptional targets. Therefore, identifying druggable transcription targets of PAX3-FKHR that are also downstream effectors of PAX3-FKHR-mediated cell migration and metastasis may lead to novel therapeutic approaches for treating ARMS. Methods: To identify genes whose expression is directly affected by the level of PAX3-FKHR in an ARMS cellular-context, we first developed an ARMS cell line in which PAX3-FKHR is stably down-regulated, and showed that stably downregulating PAX3-FKHR in ARMS cells significantly decreased the cells' motility. We used microarrays analysis to identify genes whose expression level decreased when PAX3-FKHR was downregulated. We used mutational analysis, promoter reporter assays, and electrophoretic mobility shift assays to determine whether PAX3-FKHR binds directly and specifically to the promoter region of the target gene. We used siRNA and pharmacologic inhibitor to downregulate the target gene of PAX3-FKHR and investigated the effect of such downregulation on cell motility, Results: We found that When PAX3-FKHR was downregulated, the expression of carnitine palmitoyltransferase 1A (CPT1A) decreased. We showed that PAX3-FKHR binds directly and specifically to a paired-domain binding-site in the CPT1A promoter region, indicating that CPT1A is a novel direct transcriptional target of PAX3-FKHR. Furthermore, downregulating CPT1Adecreased cell motility in ARMS cells, indicating that CPT1A is a downstream effector of PAX3-FKHR-mediated cell migration and metastasis. Conclusions: Taken together, we have identified CPT1A as a novel direct transcriptional target of PAX3-FKHR and revealed the novel function of CPT1A in promoting cell motility. CPT1A may represent a novel therapeutic target for the treatment of ARMS. Identification of transcription targets of PAX3-FKHR in human alveolar rhabdomyosarcoma: By stably knocking down PAX3-FKHR in human alveolar rhabdomyosarcoma cell line Rh30 and comparing the gene expression profile of the knockdown clone (KD) to the parental Rh30, transcription targets of PAX3-FKHR have been identified. Gene expression in parental Rh30 cells was compared to that in either Rh30 cells stably expressing shRNA targeting PAX3-FKHR (KD) or control non-targeting shRNA (CON), in duplicate.
2012-05-15 | E-GEOD-35862 | biostudies-arrayexpress
Project description:Exosomes were isolated by differential centrifugation from the fusion negative human embryonal rhabdomyosarcoma (ERMS) cell lines (JR1, Rh36, and RD) and fusion positive alveolar RMS (ARMS) cell lines (Rh30 and Rh41) and characterized by western blot for exosomal markers. miRNA content of the RMS-derived exosomes was determined using the Affymetrix GeneChip miRNA 3.0 array and analyzed to specify differentially deregulated (either enriched or depleted) miRNA relative to cellular miRNA from the respective ERMS and ARMS cell lines. Characterization of the miRNA content of RMS exosome is needed to better understand the mechanism by which these particles exert their physiologic effects, notably on proliferation, migration, invasion, and metastasis.
2016-12-01 | E-MTAB-5321 | biostudies-arrayexpress
Project description:We report the genome-wide maps of PAX3-FKHR binding sites. Chromatin immunoprecipitation was performed against PAX3-FKHR positive (Rh4) and PAX3-FKHR negative (RD) rhabdomyosarcoma cells with a monoclonal antibody (pFM2) specific for the fusion region of PAX3-FKHR. We obtained 4 million sequence tags for both input and ChIP DNA that aligned to the human genome. We identified 1,463 binding sites from ChIP-seq of Rh4 cells, none of which appeared from ChIP-seq of fusion negative RD cells. The PAX3-FKHR binding sites were found to associate with 1,072 genes in RMS cells. The data shows that PAX3-FKHR binds to the same sites as PAX3, at the enhancers for MYF5, FGFR4, and the MYOD core enhancer previously shown to be regulated by PAX3. Moreover, our dataset has the precision for rapid identification and validation of novel and specific sequences required for the enhancer activity of MYOD and FGFR4. The genome wide analysis reveals that the PAX3-FKHR sites are: 1) mostly distal to transcription start sites; 2) conserved; 3) enriched for PAX3 motifs; and 4) strongly associated with genes over-expressed in PAX3-FKHR positive RMS cells and tumors. There is little evidence in our dataset for PAX3-FKHR binding at the promoters. In one instance, we show two intronic enhancer elements for MET, rather than at the previously described promoter. The genome-wide analysis further illustrates a strong association between PAX3 and E-box motifs in these binding sites, suggestive of a common co-regulation for many target genes. The map of PAX3-FKHR binding sites provides new links for PAX3 and PAX3-FKHR functions and new targets for RMS therapy. Examination of PAX3-FKHR binding sites in translocation-positive rhabdomyosarcoma cells via ChIP-seq with an antibody specific for the fusion protein.
2010-08-17 | E-GEOD-19063 | biostudies-arrayexpress
Project description:KDM4B shRNA knockdown in Rh30 cells or QC6352 treatment of Rh30 or Rh41 cells We used microarrays to detail the global programme of gene expression underlying cellularisation and identified distinct classes of up-regulated genes during this process.
2022-05-01 | GSE158394 | GEO
Project description:We report the genome-wide maps of PAX3-FKHR binding sites. Chromatin immunoprecipitation was performed against PAX3-FKHR positive (Rh4) and PAX3-FKHR negative (RD) rhabdomyosarcoma cells with a monoclonal antibody (pFM2) specific for the fusion region of PAX3-FKHR. We obtained 4 million sequence tags for both input and ChIP DNA that aligned to the human genome. We identified 1,463 binding sites from ChIP-seq of Rh4 cells, none of which appeared from ChIP-seq of fusion negative RD cells. The PAX3-FKHR binding sites were found to associate with 1,072 genes in RMS cells. The data shows that PAX3-FKHR binds to the same sites as PAX3, at the enhancers for MYF5, FGFR4, and the MYOD core enhancer previously shown to be regulated by PAX3. Moreover, our dataset has the precision for rapid identification and validation of novel and specific sequences required for the enhancer activity of MYOD and FGFR4. The genome wide analysis reveals that the PAX3-FKHR sites are: 1) mostly distal to transcription start sites; 2) conserved; 3) enriched for PAX3 motifs; and 4) strongly associated with genes over-expressed in PAX3-FKHR positive RMS cells and tumors. There is little evidence in our dataset for PAX3-FKHR binding at the promoters. In one instance, we show two intronic enhancer elements for MET, rather than at the previously described promoter. The genome-wide analysis further illustrates a strong association between PAX3 and E-box motifs in these binding sites, suggestive of a common co-regulation for many target genes. The map of PAX3-FKHR binding sites provides new links for PAX3 and PAX3-FKHR functions and new targets for RMS therapy.
2010-08-17 | GSE19063 | GEO
Project description:In SMS-CTR cells, the overexpression of CASZ1b in SMS-CTR cells led to a modification of regional chromation accessibility.
2019-12-26 | GSE126140 | GEO
Project description:In SMS-CTR cells, we identified genomewide binding sites of CASZ1b. The overexpression of CASZ1b in SMS-CTR cells led to a regional epigenetic modification.
2019-12-26 | GSE126142 | GEO
Project description:Genetic and shRNA-mediated inhibition of SIX1 expression in RMS cells induces myogenic differentiation and impedes RMS tumor growth. To elucidate the mechanism by which SIX1 loss activates a differentiation program, we performed SIX1, MYOD1, and H3K27ac ChIPseq in two SIX1 knockdown SMS-CTR cell lines and one control SMS-CTR cell line to profile changes in transcriptional activity and myogenic transcription factor binding in fusion-negative Rhabdomyosarcoma.
2022-02-07 | GSE173151 | GEO