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.