RESUMEN

We performed a kinome-wide siRNA screen and identified 70 kinases altering cell migration in A549 lung cancer cells. In particular, ribosomal S6 kinase 1 (RSK1) silencing increased, whereas RSK2 and RSK4 downregulation inhibited cell motility. In a secondary collagen-based three-dimensional invasion screen, 38 of our hits cross-validated, including RSK1 and RSK4. In two further lung cancer cell lines, RSK1 but not RSK4 silencing showed identical modulation of cell motility. We therefore selected RSK1 for further investigation. Bioinformatic analysis followed by co-immunoprecipitation-based validation revealed that the actin regulators VASP and Mena interact with RSK1. Moreover, RSK1 phosphorylated VASP on T278, a site regulating its binding to actin. In addition, silencing of RSK1 enhanced the metastatic potential of these cells in vivo using a zebrafish model. Finally, we investigated the relevance of this finding in human lung cancer samples. In isogenically matched tissue, RSK1 was reduced in metastatic versus primary lung cancer lesions. Moreover, patients with RSK1-negative lung tumours showed increased number of metastases. Our results suggest that the findings of our high-throughput in vitro screen can reliably identify relevant clinical targets and as a proof of principle, RSK1 may provide a biomarker for metastasis in lung cancer patients.

Asunto(s)

Neoplasias Pulmonares/genética , Interferencia de ARN , ARN Interferente Pequeño/genética , Proteínas Quinasas S6 Ribosómicas 90-kDa/genética , Animales , Sitios de Unión , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Embrión no Mamífero/embriología , Embrión no Mamífero/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Microscopía Fluorescente , Metástasis de la Neoplasia , Trasplante de Neoplasias , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Fosforilación , Unión Proteica , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteínas Quinasas S6 Ribosómicas 90-kDa/metabolismo , Treonina/genética , Treonina/metabolismo , Trasplante Heterólogo , Pez Cebra/embriología