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1.
Proc Natl Acad Sci U S A ; 119(10): e2119676119, 2022 03 08.
Article in English | MEDLINE | ID: mdl-35235462

ABSTRACT

Lymphocytic choriomeningitis virus (LCMV) is a rodent-borne zoonotic arenavirus that causes congenital abnormalities and can be fatal for transplant recipients. Using a genome-wide loss-of-function screen, we identify host factors required for LCMV entry into cells. We identify the lysosomal mucin CD164, glycosylation factors, the heparan sulfate biosynthesis machinery, and the known receptor alpha-dystroglycan (α-DG). Biochemical analysis revealed that the LCMV glycoprotein binds CD164 at acidic pH and requires a sialylated glycan at residue N104. We demonstrate that LCMV entry proceeds by the virus switching binding from heparan sulfate or α-DG at the plasma membrane to CD164 prior to membrane fusion, thus identifying additional potential targets for therapeutic intervention.


Subject(s)
Lymphocytic choriomeningitis virus/physiology , Virus Internalization , A549 Cells , CRISPR-Cas Systems , Endolyn/physiology , Gene Editing , HEK293 Cells , HeLa Cells , Host-Pathogen Interactions , Humans , Hydrogen-Ion Concentration , Lymphocytic choriomeningitis virus/pathogenicity , Membrane Fusion , Virulence Factors
2.
PLoS Pathog ; 5(4): e1000394, 2009 Apr.
Article in English | MEDLINE | ID: mdl-19390604

ABSTRACT

Many viruses that enter cells by clathrin-dependent endocytosis are significantly larger than the dimensions of a typical clathrin-coated vesicle. The mechanisms by which viruses co-opt the clathrin machinery for efficient internalization remain uncertain. Here we examined how clathrin-coated vesicles accommodate vesicular stomatitis virus (VSV) during its entry into cells. Using high-resolution imaging of the internalization of single viral particles into cells expressing fluorescent clathrin and adaptor molecules, we show that VSV enters cells through partially clathrin-coated vesicles. We found that on average, virus-containing vesicles contain more clathrin and clathrin adaptor molecules than conventional vesicles, but this increase is insufficient to permit full coating of the vesicle. We further show that virus-containing vesicles depend upon the actin machinery for their internalization. Specifically, we found that components of the actin machinery are recruited to virus-containing vesicles, and chemical inhibition of actin polymerization trapped viral particles in vesicles at the plasma membrane. By analysis of multiple independent virus internalization events, we show that VSV induces the nucleation of clathrin for its uptake, rather than depending upon random capture by formation of a clathrin-coated pit. This work provides new mechanistic insights into the process of virus internalization as well as uptake of unconventional cargo by the clathrin-dependent endocytic machinery.


Subject(s)
Actins/metabolism , Clathrin-Coated Vesicles/virology , Vesiculovirus/pathogenicity , Virus Internalization , Clathrin/analysis , Endocytosis , Microscopy, Fluorescence
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