Abstract
Mammalian reoviruses replicate in a broad range of hosts, cells, and tissues. These viruses display strain-dependent variation in tropism for different types of cells in vivo and ex vivo. Early steps in the reovirus life cycle, attachment, entry, and disassembly, have been identified as pivotal points of virus-cell interaction that determine the fate of infection, either productive or abortive. However, in studies of the differential capacity of reovirus strains type 1 Lang and type 3 Dearing to replicate in Madin-Darby canine kidney (MDCK) cells, we found that replication efficiency is regulated at a late point in the viral life cycle following primary transcription and translation. Results of genetic studies using recombinant virus strains show that reovirus tropism for MDCK cells is primarily regulated by replication protein μ2 and further influenced by the viral RNA-dependent RNA polymerase protein, λ3, depending on the viral genetic background. Furthermore, μ2 residue 347 is a critical determinant of replication efficiency in MDCK cells. These findings indicate that components of the reovirus replication complex are mediators of cell-selective viral replication capacity at a post-entry step. Thus, reovirus cell tropism may be determined at early and late points in the viral replication program.
Highlights
Reovirus Strain-specific Replication in Madin-Darby canine kidney (MDCK) Cells—Previous studies revealed a strain-specific capacity of reovirus type 1 Lang (T1) and type 3 Dearing (T3) to replicate in MDCK cells [38]
The effect of M1 on viral replication was unidirectional; the T3 M1 gene segment alone did not diminish T1 yield. These results indicate that the 2 protein regulates reovirus replication efficiency in MDCK cells, but the effects of 2 on replication are subject to modulation by other viral determinants
Our primary findings emerging from this study are the following: 1) 2 controls efficiency of reovirus growth in MDCK cells; 2) residue 347 is the primary determinant of 2-mediated viral replication potential in these cells; 3) polymerase protein 3 is a coregulator of viral replication in a strain-dependent manner; and 4) 2 protein appears to regulate a later step in the reovirus replication program subsequent to primary rounds of viral transcription and translation
Summary
Secondary viral mRNAs transcribed from new dsRNA templates fuel subsequent rounds of protein synthesis The source of these late transcripts is presumed to be subviral particles. Reovirus replication and assembly are thought to occur within viral intracytoplasmic inclusions [10], which strictly depend on nonstructural proteins NS and NS and structural protein 2 for proper formation and function in reovirus-infected cells [11, 12]. Reovirus Cell Tropism and Replication Protein 2 tectable within 4 h post-infection, lack a delimiting membrane, contain viral proteins, dsRNA and virion particles at various stages of morphogenesis, and are composed of highly ordered arrays of mature virions at late times of infection [13,14,15,16,17,18,19]. Fluorescence microscopy performed on a variety of cell types has shown that T1 and T3 form filamentous and globular inclusions, respectively [11, 20, 23, 24], which is explained by a differential capacity of T1 and T3 2 proteins to bind microtubules and anchor inclusions to the cytoskeleton [24]
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