Abstract
Oncolytic Newcastle disease virus (NDV) reportedly employs direct fusion of the viral envelope with the plasma membrane and caveolae-dependent endocytosis to enter cells. Here, we show that macropinocytosis and clathrin-mediated endocytosis are involved in NDV entry into a galline embryonic fibroblast cell line. Upon specific inhibition of clathrin assembly, GTPase dynamin, Na+/H+ exchangers, Ras-related C3 botulinum toxin substrate 1, p21 activated kinase 1 or protein kinase C, entry of NDV and its propagation were suppressed. NDV entry into cells triggers Rac1-Pak1 signaling and elicits actin rearrangement and plasma membrane ruffling. Moreover, NDV internalization within macropinosomes and trafficking involve Rab5a-positive vesicles. This is the first report demonstrating that NDV utilizes clathrin-mediated endocytosis and macropinocytosis as alternative endocytic pathways to enter cells. These findings shed new light on the molecular mechanisms underlying NDV entry into cells, and provide potential targets for NDV-mediated therapy in cancer.
Highlights
Newcastle disease virus (NDV) is an avian paramyxovirus
These results indicate that NDV proteins and genomic RNA were probably produced within 2 hpi and completed the replicative cycle within 4 hpi
We investigated whether DF-1 cells utilize an endocytic mechanism for NDV uptake
Summary
Newcastle disease virus (NDV) is an avian paramyxovirus. Its virions are pleomorphic, but mostly spherical with a diameter of approximately 100 nm [1]. It is thought that NDV enters cells after direct fusion with the plasma membrane through a pH-independent mechanism. NDV may enter host cells through a caveolaemediated endocytosis (CavME) pathway [3], as well as through other endocytotic routes that are not yet well understood. The mainly endocytic pathways include clathrinmediated endocytosis (CME), CavME, macropinocytosis and phagocytosis, among others [4]. With CME, clathrin is assembled on the cytoplasmic face of the plasma membrane to form a clathrin-coated pit (CCP). Internalization of the virus-receptor complexes occurs within CCPs, which invaginate toward the cytoplasmic side and dissociate to form clathrin-coated vesicles containing the endocytic cargo, which are delivered into endosomes. Semliki forest virus [5], vesicular stomatitis virus [6, 7], egg drop syndrome virus [8], and human immunodeficiency virus-1 (HIV-1) [9] all use the CME pathway to enter cells
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