title: Targeting mitosis-regulating genes in cisplatin-sensitive and -resistant melanoma cells: A live-cell RNAi screen displays differential nucleus-derived phenotypes.
abstract: Chemoresistance in malignant melanoma remains an unresolved clinical issue. In the search for novel molecular targets, a live-cell high-content RNAi screen based on gene expression data was performed in cisplatin-sensitive and cisplatin-resistant MeWo melanoma cells, Mel-28 cells and a melanocyte cell line. Cells were exposed to 91 siRNAs and distinct nucleus-derived phenotypes such as cell division, cell death and migration phenotypes were detected by time-lapse microscopy over 60 h. Using this approach, cisplatin-sensitive and cisplatin-resistant melanoma cells were compared by automated image analysis and visual inspection. In cisplatin-sensitive MeWo melanoma cells, 14 genes were identified that showed distinct phenotype abnormalities after exposure to gene-specific siRNAs. In cisplatin-resistant MeWo cells, five genes were detected. Nine genes were detected whose knock-down led to differential nuclear phenotypes in cisplatin-sensitive and -resistant cells. In Mel-28 cells, nine genes were identified which induced nuclear phenotypes including all eight genes which were identified in cisplatin-resistant MeWo cells. An analogous RNAi screen on melanocytes revealed no detectable phenotype abnormalities after RNAi. Pathway analysis showed in cisplatin-sensitive MeWo cells and Mel-28 cells an enrichment of at least three genes in major mitotic pathways. We hereby show that siRNA screening may help to identify tumor-specific genes leading to phenotype abnormalities. These genes may serve as potential therapeutic targets in the treatment of melanoma.
authors.affiliation: BioQuant, Heidelberg University, Heidelberg, Germany. holger.erfle@bioquant.uni-heidelberg.de.
authors.forename: Holger
authors.initials: H
authors.lastname: Erfle
authors.affiliation: BioQuant, Heidelberg University, Heidelberg, Germany.|Clinic for Vascular and Endovascular Surgery, University Hospital, Frankfurt, Germany.
authors.forename: K
authors.initials: K
authors.lastname: Pashayeva
authors.affiliation: BioQuant and IPMB, University of Heidelberg and DKFZ, Biomedical Computer Vision Group, Heidelberg, Germany.
authors.forename: N
authors.initials: N
authors.lastname: Harder
authors.affiliation: Clinic for Vascular and Endovascular Surgery, University Hospital, Frankfurt, Germany.
authors.forename: L
authors.initials: L
authors.lastname: Zhang
authors.affiliation: BioQuant and IPMB, University of Heidelberg and DKFZ, Biomedical Computer Vision Group, Heidelberg, Germany.
authors.forename: K
authors.initials: K
authors.lastname: Rohr
authors.affiliation: Department of Dermatology, University of Duisburg-Essen, Essen, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Schadendorf
authors.affiliation: Department of Dermatology, University of Würzburg, Würzburg, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Ugurel
authors.affiliation: Clinic for Vascular and Endovascular Surgery, University Hospital, Frankfurt, Germany. michael.keese@kgu.de.
authors.forename: M
authors.initials: M
authors.lastname: Keese
mesh_terms: D045744:Cell Line, Tumor; D002467:Cell Nucleus; D002945:Cisplatin / Q000494:pharmacology*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008545:Melanoma / Q000235:genetics*; D008938:Mitosis / Q000235:genetics*; D010641:Phenotype; D034622:RNA Interference / Q000187:drug effects*; D014162:Transfection
chemical_list: D002945:Cisplatin
keywords: Cisplatin resistance; Melanoma; Mitotic phenotype; Phenotypic RNAi screen