The Norovirus NS3 Protein Is a Dynamic Lipid- and Microtubule-Associated Protein Involved in Viral RNA Replication

Autor: Ben T Cotton, Kim Y. Green, Stanislav V. Sosnovtsev, Jennifer L. Hyde, Jason M. Mackenzie, Soroush T Sarvestani, Peter A. White
Rok vydání: 2017
Předmět:
0301 basic medicine
Microtubule-associated protein
Viral protein
viruses
030106 microbiology
Immunology
ved/biology.organism_classification_rank.species
Intracellular Space
Viral Nonstructural Proteins
Biology
Virus Replication
medicine.disease_cause
Microtubules
Microbiology
Cell Line
Mice
03 medical and health sciences
Membrane Microdomains
Virology
Chlorocebus aethiops
medicine
Animals
Amino Acid Sequence
Vero Cells
Caliciviridae Infections
RNA
Double-Stranded
NS3
ved/biology
Norovirus
Membrane Proteins
virus diseases
Lipid Metabolism
digestive system diseases
Virus-Cell Interactions
Cell biology
Transport protein
Protein Transport
Cholesterol
030104 developmental biology
Viral replication
Membrane protein
Insect Science
Viral replication complex
Host-Pathogen Interactions
RNA
Viral
Protein Binding
Murine norovirus
Zdroj: Journal of Virology. 91
ISSN: 1098-5514
0022-538X
Popis: Norovirus (NoV) infections are a significant health burden to society, yet the lack of reliable tissue culture systems has hampered the development of appropriate antiviral therapies. Here we show that the NoV NS3 protein, derived from murine NoV (MNV), is intimately associated with the MNV replication complex and the viral replication intermediate double-stranded RNA (dsRNA). We observed that when expressed individually, MNV NS3 and NS3 encoded by human Norwalk virus (NV) induced the formation of distinct vesicle-like structures that did not colocalize with any particular protein markers to cellular organelles but localized to cellular membranes, in particular those with a high cholesterol content. Both proteins also showed some degree of colocalization with the cytoskeleton marker β-tubulin. Although the distribution of MNV and NV NS3s were similar, NV NS3 displayed a higher level of colocalization with the Golgi apparatus and the endoplasmic reticulum (ER). However, we observed that although both proteins colocalized in membranes counterstained with filipin, an indicator of cholesterol content, MNV NS3 displayed a greater association with flotillin and stomatin, proteins known to associate with sphingolipid- and cholesterol-rich microdomains. Utilizing time-lapse epifluorescence microscopy, we observed that the membrane-derived vesicular structures induced by MNV NS3 were highly motile and dynamic in nature, and their movement was dependent on intact microtubules. These results begin to interrogate the functions of NoV proteins during virus replication and highlight the conserved properties of the NoV NS3 proteins among the seven Norovirus genogroups. IMPORTANCE Many mechanisms involved in the replication of norovirus still remain unclear, including the role for the NS3 protein, one of seven nonstructural viral proteins, which remains to be elucidated. This study reveals that murine norovirus (MNV) NS3 is intimately associated with the viral replication complex and dsRNA. We observed that the NS3 proteins of both MNV and Norwalk virus (NV) induce prominent vesicular structures and that this formation is dependent on microtubules and cellular cholesterol. Thus, this study contributes to our understanding of protein function within different Norovirus genogroups and expands a growing knowledge base on the interaction between positive-strand RNA [(+)RNA] viruses and cellular membranes that contribute to the biogenesis of virus-induced membrane organelles. This study contributes to our understanding of viral protein function and the ability of a viral protein to recruit specific cellular organelles and lipids that enable replication.
Databáze: OpenAIRE
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