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Dissection of mammalian orthoreovirus µ2 reveals a self-associative domain required for binding to microtubules but not to factory matrix protein µNS


Eichwald, Catherine; Kim, Jonghwa; Nibert, Max L (2017). Dissection of mammalian orthoreovirus µ2 reveals a self-associative domain required for binding to microtubules but not to factory matrix protein µNS. PLoS ONE, 12(9):e0184356.

Abstract

Mammalian orthoreovirus protein μ2 is a component of the viral core particle. Its activities include RNA binding and hydrolysis of the γ-phosphate from NTPs and RNA 5´-termini, suggesting roles as a cofactor for the viral RNA-dependent RNA polymerase, λ3, first enzyme in 5´-capping of viral plus-strand RNAs, and/or prohibitory of RNA-5´-triphosphate-activated antiviral signaling. Within infected cells, μ2 also contributes to viral factories, cytoplasmic structures in which genome replication and particle assembly occur. By associating with both microtubules (MTs) and viral factory matrix protein μNS, μ2 can anchor the factories to MTs, the full effects of which remain unknown. In this study, a protease-hypersensitive region allowed μ2 to be dissected into two large fragments corresponding to residues 1-282 and 283-736. Fusions with enhanced green fluorescent protein revealed that these amino- and carboxyl-terminal regions of μ2 associate in cells with either MTs or μNS, respectively. More exhaustive deletion analysis defined μ2 residues 1-325 as the minimal contiguous region that associates with MTs in the absence of the self-associating tag. A region involved in μ2 self-association was mapped to residues 283-325, and self-association involving this region was essential for MT-association as well. Likewise, we mapped that μNS-binding site in μ2 relates to residues 290-453 which is independent of μ2 self-association. These findings suggest that μ2 monomers or oligomers can bind to MTs and μNS, but that self-association involving μ2 residues 283-325 is specifically relevant for MT-association during viral factories formation.

Abstract

Mammalian orthoreovirus protein μ2 is a component of the viral core particle. Its activities include RNA binding and hydrolysis of the γ-phosphate from NTPs and RNA 5´-termini, suggesting roles as a cofactor for the viral RNA-dependent RNA polymerase, λ3, first enzyme in 5´-capping of viral plus-strand RNAs, and/or prohibitory of RNA-5´-triphosphate-activated antiviral signaling. Within infected cells, μ2 also contributes to viral factories, cytoplasmic structures in which genome replication and particle assembly occur. By associating with both microtubules (MTs) and viral factory matrix protein μNS, μ2 can anchor the factories to MTs, the full effects of which remain unknown. In this study, a protease-hypersensitive region allowed μ2 to be dissected into two large fragments corresponding to residues 1-282 and 283-736. Fusions with enhanced green fluorescent protein revealed that these amino- and carboxyl-terminal regions of μ2 associate in cells with either MTs or μNS, respectively. More exhaustive deletion analysis defined μ2 residues 1-325 as the minimal contiguous region that associates with MTs in the absence of the self-associating tag. A region involved in μ2 self-association was mapped to residues 283-325, and self-association involving this region was essential for MT-association as well. Likewise, we mapped that μNS-binding site in μ2 relates to residues 290-453 which is independent of μ2 self-association. These findings suggest that μ2 monomers or oligomers can bind to MTs and μNS, but that self-association involving μ2 residues 283-325 is specifically relevant for MT-association during viral factories formation.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Institute of Virology
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:2017
Deposited On:19 Sep 2017 15:12
Last Modified:19 Sep 2017 15:12
Publisher:Public Library of Science (PLoS)
ISSN:1932-6203
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1371/journal.pone.0184356
PubMed ID:28880890

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