Coronavirus replication complex formation utilizes components of cellular autophagy.
The coronavirus mouse hepatitis virus (MHV) performs RNA replication on double membrane vesicles (DMVs) in the cytoplasm of the host cell. However, the mechanism by which these DMVs form has not been determined. Using genetic, biochemical, and cell imaging approaches, the role of autophagy in DMV formation and MHV replication was investigated. The results demonstrated that replication complexes co-localize with the autophagy proteins, microtubule-associated protein light-chain 3 and Apg12. MHV infection induces autophagy by a mechanism that is resistant to 3-methyladenine inhibition. MHV replication is impaired in autophagy knockout, APG5-/-, embryonic stem cell lines, but wild-type levels of MHV replication are restored by expression of Apg5 in the APG5-/-cells. In MHV-infected APG5-/-cells, DMVs were not detected; rather, the rough endoplasmic reticulum was dramatically swollen. The results of this study suggest that autophagy is required for formation of double membrane-bound MHV replication complexes and that DMV formation significantly enhances the efficiency of replication. Furthermore, the rough endoplasmic reticulum is implicated as the possible source of membranes for replication complexes.
Prentice
Erik
E
Department of Microbiology and Immunology, Elizabeth B. Lamb Center for Pediatric Research, Vanderbilt University Medical Center, Nashville, Tennessee 37221, USA.
Jerome
W Gray
WG
Yoshimori
Tamotsu
T
Mizushima
Noboru
N
Denison
Mark R
MR
D000225:Adenine / Q000031:analogs & derivatives* / Q000737:chemistry; D000818:Animals; D001343:Autophagy; D000071192:Autophagy-Related Protein 12; D015153:Blotting, Western; D002455:Cell Division; D002460:Cell Line; D017934:Coronavirus / Q000235:genetics / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology; D018870:Endoplasmic Reticulum, Rough / Q000378:metabolism; D051379:Mice; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D010802:Phylogeny; D011485:Protein Binding; D021381:Protein Transport; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013234:Stem Cells; D013997:Time Factors
C491971:Atg12 protein, mouse; D000071192:Autophagy-Related Protein 12; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins; C025946:3-methyladenine; D000225:Adenine