Real-time analysis of clathrin-mediated endocytosis during cell migration

Abstract

Simultaneous dual-color total-internal-reflection fluorescence microscopy (TIR-FM) was performed to analyze the internalization and distribution of markers for clathrin-mediated endocytosis (clathrin, dynamin1, dynamin2 and transferrin) in migrating cells. In MDCK cells, which endogenously express dynamin2, the dynamin2-EGFP fluorescence demonstrated identical spatial and temporal behavior as clathrin both prior to and during internalization. By contrast, in the same cells, the neuronal dynamin1 only localized with clathrin just prior to endocytosis. In migrating cells, each endocytic marker was polarized towards the leading edge, away from the lagging edge. These observations suggest a re-evaluation of the functional differences between dynamin1 and dynamin2, and of the role of clathrin-mediated endocytosis in cell migration.