A Novel Techniques to Follow Consequences of Exogenous Factors, Including Therapeutic Drugs, on Living Human Breast Epithelial Cells
Abstract
Monitoring fluorescently tagged proteins in live cells enables the observation of spatial and temporal events in a way that would otherwise not be possible. We are using this approach to examine normal and tumor human mammary epithelial cells (growing in 2D and 3D cultures) before and after addition of exogenous agents. We used confocal microscopy to examine living cells containing proteins tagged with green fluorescent protein (GFP). We examined GFP-tagged cytoskeletal proteins, those proteins responsible for cell shape and transport within the cell, and GFP-tagged proteins of the Wnt signaling pathway, a pathway involved in cancer. Normal cells demonstrate slow, directional movements across the dish whereas tumor cells demonstrate rapid and exaggerated, random movements. Cells containing actin-GFP show that tumor cells, unlike normal cells, have excessive mediated ruffles, filopodia, and microspikes. Cells containing GFP-tagged beta-catenin, a Wnt signaling protein, become rounded and demonstrate large aggregates of beta-catenin in the cell nucleus, consistent with reports that beta-catenin can activate gene transcription. The cells with beta-catenin-GFP also demonstrate numerous protrusions ("blebs") emanating form the cell surface. These protrusions are rapidly extending and contracting as the cells make random movements around the dish.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jul 01, 2000
- Accession Number
- ADA386854
Entities
Organizations
- University of California, Berkeley