Abstract 5197: Comparative proteome analysis reveals flotillin-1 as an H-Ras-interacting lipid raft protein critical for breast epithelial cell invasion

Abstract Metastatic spread is the major cause of death from breast cancer. The objective of our study is to unveil signal transduction pathways specific to breast epithelial cell invasion and migration apart from cell proliferation/transformation. We previously showed that the active H-Ras, but not N-Ras, induces invasive/migratory phenotypes of MCF10A human breast epithelial cells via activation of Rac1/p38 signaling leading to MMP-2 up-regulation, while both H-Ras and N-Ras induce cell proliferation and transformation. In the present study, we hypothesized that the Ras isoform-specific signaling pathway is initiated by differential involvement of lipid raft proteins. To address this, we characterized proteome profiles of lipid raft proteins from three cell lines: normal MCF10A; N-Ras-transformed, non-invasive MCF10A; and H-Ras-transformed, invasive MCF10A cells. Among twenty-four lipid raft proteins that were up-regulated in an H-Ras-specific manner, we tested whether flotillin-1 plays a causative role for H-Ras-mediated MCF10A cell invasion/migration. Here, we show that flotillin-1 interacts with H-Ras and H-Ras signaling increases flotillin-1 expression, suggesting a signaling amplification loop between flotillin-1 and H-Ras. Importantly, siRNA-mediated flotillin-1 knockdown significantly reduced H-Ras-mediated motility/invasion accompanied with down-regulation of Rac1/p38 and MMP-2. We also show that flotillin-1 co-localizes with H-Ras in the plasma membrane, while the interaction between flotillin-1 and N-Ras was negligible. Taken together, this study identified flotillin-1 as an H-Ras-interacting lipid raft protein critical for activation of H-Ras-specific signaling resulting invasive phenotype in breast epithelial cells. Consistently, flotillin-1 mRNA was significantly higher in malignant breast cancer tissues, supporting our hypothesis that flotillin-1 may contribute to breast carcinoma invasion. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5197.