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SiRNA-mediated knockdown of MITF and PAX3 in metastatic melanoma cell lines

ABSTRACT: The transcription factors PAX3 and MITF are required for the development of the neural crest and melanocyte lineage, and both proteins play important roles in melanoma cell growth and survival. PAX3 transcriptionally activates MITF expression during neural crest development, but the relationship between these transcription factors during melanocyte development and in melanoma cells is currently poorly understood. This study aimed to further our understanding of the interaction between transcriptional networks controlled by PAX3 and MITF by assessing the effect of siRNA-mediated knockdown of PAX3 and MITF in metastatic melanoma cell lines. The goals of this study were to determine (i) if PAX3 is required for maintaining expression of MITF in melanoma and melanocyte cell lines; (ii) whether PAX3 and MITF independently, or redundantly, influence growth and survival in melanoma cell lines; and (iii) to investigate the respective roles of PAX3 and MITF expression in melanoma cell differentiation. Microarrays were used to measure global changes in transcript expression in response to siRNA-mediated knockdown of PAX3 or MITF compared to non-targeting controls in two metastatic melanoma cells lines.

ORGANISM(S): Homo sapiens

PROVIDER: GSE16249 | GEO | 2010/01/02

SECONDARY ACCESSION(S): PRJNA115189

REPOSITORIES: GEO

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siRNA-mediated knockdown of MITF and PAX3 in metastatic melanoma cell lines

Project description:The transcription factors PAX3 and MITF are required for the development of the neural crest and melanocyte lineage, and both proteins play important roles in melanoma cell growth and survival. PAX3 transcriptionally activates MITF expression during neural crest development, but the relationship between these transcription factors during melanocyte development and in melanoma cells is currently poorly understood. This study aimed to further our understanding of the interaction between transcriptional networks controlled by PAX3 and MITF by assessing the effect of siRNA-mediated knockdown of PAX3 and MITF in metastatic melanoma cell lines. The goals of this study were to determine (i) if PAX3 is required for maintaining expression of MITF in melanoma and melanocyte cell lines; (ii) whether PAX3 and MITF independently, or redundantly, influence growth and survival in melanoma cell lines; and (iii) to investigate the respective roles of PAX3 and MITF expression in melanoma cell differentiation. Microarrays were used to measure global changes in transcript expression in response to siRNA-mediated knockdown of PAX3 or MITF compared to non-targeting controls in two metastatic melanoma cells lines. RNA was isolated from two different metastatic melanoma cell lines 30 hours after one of four different treaments: (i) transfection with siRNA targeting PAX3; or (ii) transfection with siRNA targeting MITF; or (iii) or transfection with siRNA targeting luciferase (non-targeting negative control); or (iv) treatment with media only (control). Therefore, eight samples were used for gene expression profiling by using GeneChip arrays, with one replicate per cell line per treatment.

2010-01-01 | E-GEOD-16249 | biostudies-arrayexpress

DNA Promoter Hypermethylation of Melanocyte Lineage Genes Determines Melanoma Phenotype

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Homo sapiens

Project description:siRNA-mediated knockdown of MITF and PAX3 in metastatic melanoma cell lines

| PRJNA115189 | ENA

MITF deficiency and oncogenic GNAQ each promote proliferation programs in zebrafish melanocyte lineage cells

Project description:Uveal melanoma (UM) is the most common primary malignancy of the eye in adults, but lacks any FDA-approved therapy for the deadly metastatic disease. Thus, there is great need to dissect the driving mechanisms for UM, and develop strategies to evaluate potential therapeutics. Using a zebrafish model, we previously identified MITF, the master melanocyte transcription factor, as a tumor suppressor in GNAQQ209L-driven UM. Here, we show that zebrafish mitfa-deficient GNAQQ209L-driven tumors significantly up-regulate neural crest markers, and higher expression of a melanoma-associated neural crest signature correlates with poor UM patient survival. We further determined how the mitfa-null state, as well as expression of GNAQQ209L, YAPS127A;S381A, or BRAFV600E oncogenes, impacts melanocyte lineage cells before they acquire the transformed state. Specifically, examination 5 days post-fertilization showed that mitfa-deficiency is sufficient to upregulate pigment progenitor and neural crest markers, while GNAQQ209L expression promotes a proliferative phenotype that is further enhanced by YAPS127A;S381A co-expression. Finally, we show that this oncogene-induced proliferative phenotype can be used to screen chemical inhibitors for their efficacy against the UM pathway. Overall, this study establishes that a neural crest signature correlates with poor UM survival, and describes an in vivo assay for pre-clinical trials of potential UM therapeutics.

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Chromatin remodellers Brg1 and Bptf are required for normal gene expression and progression of oncogenic Braf-driven mouse melanoma [ChIP-seq]

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Feed-forward Cysteine Regulation Maintains Melanoma Differentiation State and Limits Metastatic Spread

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