Exocytosis and Endocytosis

Volume 440 of the series Methods in Molecular Biology pp 371-387

Probing E-Cadherin Endocytosis by Morpholino-Mediated Rab5 Knockdown in Zebrafish

  • Florian UlrichAffiliated withMax Planck Institute for Molecular Cell Biology and Genetics
  • , Carl-Philipp HeisenbergAffiliated withMax Planck Institute for Molecular Cell Biology and Genetics

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The controlled internalization of membrane receptors and lipids is crucial for cells to control signaling pathways and interact with their environment. During clathrin-mediated endocytosis, membrane constituents are transported via endocytic vesicles into early endosomes, from which they are further distributed within the cell. The small guanosine triphosphatase (GTPase) Rab5 is both required and sufficient for the formation of these early endosomes and can be used to experimentally address endocytic processes. Recent evidence shows that endocytic turnover of E-cadherin regulates the migration of mesendodermal cells during zebrafish gastrulation by modulating their adhesive interactions with neighboring cells. This in turn leads to effective and synchronized movement within the embryo. In this review, we discuss techniques to manipulate E-cadherin endocytosis by morpholino-mediated knockdown of rab5 during zebrafish gastrulation. We describe the use of antibodies specifically directed against zebrafish E-cadherin to detect its intracellular localization and of in situ hybridization and primary cell culture to reveal patterns of cell migration and adhesion, respectively.


Cell migration endocytosis endosomes gastrulation mesendoderm morpholino small GTPase rab5 zebrafish