A novel role for flotillin-1 in H-Ras-regulated breast cancer aggressiveness

Abstract

Elevated expression and aberrant activation of Ras have been implicated in breast cancer aggressiveness. H-Ras, but not N-Ras, induces breast cell invasion. A crucial link between lipid rafts and H-Ras function has been suggested. This study sought to identify the lipid raft protein(s) responsible for H-Ras-induced tumorigenicity and invasiveness of breast cancer. We conducted a comparative proteomic analysis of lipid raft proteins from invasive MCF10A human breast epithelial cells engineered to express active H-Ras and non-invasive cells expressing active N-Ras. Here, we identified a lipid raft protein flotillin-1 as an important regulator of H-Ras activation and breast cell invasion. Flotillin-1 was required for epidermal growth factor-induced activation of H-Ras, but not that of N-Ras, in MDA-MB-231 triple-negative breast cancer (TNBC) cells. Flotillin-1 knockdown inhibited the invasiveness of MDA-MB-231 and Hs578T TNBC cells in vitro and in vivo. In xenograft mouse tumor models of these TNBC cell lines, we showed that flotillin-1 played a critical role in tumor growth. Using human breast cancer samples, we provided clinical evidence for the metastatic potential of flotillin-1. Membrane staining of flotillin-1 was positively correlated with metastatic spread (p = 0.013) and inversely correlated with patient disease-free survival rates (p = 0.005). Expression of flotillin-1 was associated with H-Ras in breast cancer, especially in TNBC (p < 0.001). Our findings provide insight into the molecular basis of Ras isoform-specific interplay with flotillin-1, leading to tumorigenicity and aggressiveness of breast cancer. What's new? We identified a lipid raft protein flotillin-1 as an important regulator of H-Ras activation and breast cancer aggressiveness. Flotillin-1 was required for epidermal growth factor-induced activation of H-Ras, but not that of N-Ras, in triple-negative breast cancer (TNBC) cells. Flotillin-1 knockdown inhibited the invasiveness and tumorigenicity of TNBC cells in vivo. Using human breast cancer samples, we provide clinical evidence for the tumorigenic and metastatic potential of flotillin-1 and its association with H-Ras. Oncogenic activation of H-Ras is a common feature of breast cancer, with the mutant protein serving a leading role in tumorigenesis and also likely contributing to the induction of tumor invasion and metastasis. In this study, the lipid raft protein flotillin-1 was found to be a key regulator of H-Ras activation and tumor aggressiveness in breast cancer. In triple-negative breast cancer (TNBC) cells, flotillin-1 was required for epidermal growth factor-induced H-Ras activation, and in vivo, flotillin-1 knockdown inhibited TNBC invasiveness and tumorigenicity. In patient samples, flotillin-1 membrane staining correlated positively with metastasized disease.