A Novel Assay to Study Jagged-1 Trafficking

Abstract

The Notch signaling pathway is an evolutionarily conserved cell-signaling system that plays an important role in cell differentiation and other processes. Notch signaling is activated when a ligand binds to the Notch protein at the cell surface, thereby initiating a series of events that leads to modification of gene expression. Notch and one of its ligands, Jagged-1, are found at elevated levels in metastatic prostate cancer, indicating that increased Notch signaling is responsible for metastasis of prostate cancer. If Jagged-1 were prevented from reaching the cell surface in the first place, prostate cancer metastasis could be averted. However, more knowledge about the trafficking of Jagged-1 is required to achieve that goal. This study hopes to develop a novel assay that could aid in the study of protein trafficking. A preliminary quantitative assay showed that the manufactured single chain variable fragment antibody bound to the Jagged-1 receptor at the cell surface and exhibited luciferase activity. Another qualitative assay showed that both receptor and antibody could be visualized fluorescently and that the internalization of the antibody could be tracked visually. These experiments will serve as a starting point for more extensive assays which will further optimize these techniques. This knowledge can then be utilized to combat cancer metastasis by blunting the expression of Jagged-1 at the cell surface.