Endocytosis of gene delivery vectors: from clathrin-dependent to lipid raft-mediated endocytosis.

The ideal nonviral vector delivers its nucleic acid cargo to a specific intracellular target. Vectors enter cells mainly through endocytosis and are distributed to various intracellular organelles. Recent advances in microscopy, lipidomics, and proteomics confirm that the cell membrane is composed of clusters of lipids, organized in the form of lipid raft domains, together with non-raft domains that comprise a generally disordered lipid milieu. The binding of a nonviral vector to either region can determine the pathway for its endocytic uptake and subsequent intracellular itinerary. Given this model of the cell membrane structure, endocytic pathways should be reclassified in relation to lipid rafts. In this review, we attempt to assess the currently recognized endocytic pathways in mammalian cells. The endocytic pathways are classified in relation to the membrane regions that make up the primary endocytic vesicles. This review covers the well-recognized clathrin-mediated endocytosis (CME), phagocytosis, and macropinocytosis in addition to the less addressed pathways that take place in lipid rafts. These include caveolae-mediated, flotillin-dependent, GTPase regulator associated with focal adhesion kinase-1 (GRAF1)-dependent, adenosine diphosphate-ribosylation factor 6 (Arf6)-dependent, and RhoA-dependent endocytic pathways. We summarize the regulators associated with each uptake pathway and methods for interfering with these regulators are discussed. The fate of endocytic vesicles resulting from each endocytic uptake pathway is highlighted.