Abstract
Protein production, genomic RNA replication, and virion assembly during infection by picornaviruses like human rhinovirus and poliovirus take place in the cytoplasm of infected human cells, making them the quintessential cytoplasmic pathogens. However, a growing body of evidence suggests that picornavirus replication is promoted by a number of host proteins localized normally within the host cell nucleus. To systematically identify such nuclear proteins, we focused on those that appear to re-equilibrate from the nucleus to the cytoplasm during infection of HeLa cells with human rhinovirus via quantitative protein mass spectrometry. Our analysis revealed a highly selective re-equilibration of proteins with known mRNA splicing and transport-related functions over nuclear proteins of all other functional classes. The multifunctional splicing factor proline and glutamine rich (SFPQ) was identified as one such protein. We found that SFPQ is targeted for proteolysis within the nucleus by viral proteinase 3CD/3C, and a fragment of SFPQ was shown to migrate to the cytoplasm at mid-to-late times of infection. Cells knocked down for SFPQ expression showed significantly reduced rhinovirus titers, viral protein production, and viral RNA accumulation, consistent with SFPQ being a pro-viral factor. The SFPQ fragment that moved into the cytoplasm was able to bind rhinovirus RNA either directly or indirectly. We propose that the truncated form of SFPQ promotes viral RNA stability or replication, or virion morphogenesis. More broadly, our findings reveal dramatic changes in protein compartmentalization during human rhinovirus infection, allowing the virus to systematically hijack the functions of proteins not normally found at its cytoplasmic site of replication.
Highlights
Viruses of the Picornaviridae are characterized by a positive polarity, single-stranded RNA genome of 7–10 kb within a non-enveloped icosahedral capsid
We explored the dynamics of host cell protein relocalization from the nucleus to the cytoplasm during an infection by human rhinovirus using quantitative mass spectrometry, confocal imaging, and Western blot analysis
HRV16 infection induces a coordinated redistribution of proteins into the cytoplasm of infected cells
Summary
Viruses of the Picornaviridae are characterized by a positive polarity, single-stranded RNA genome of 7–10 kb within a non-enveloped icosahedral capsid. Genome replication is promoted by elements of secondary structure present at the termini of both positive- and negative-stranded viral RNA. These structures serve as the interaction sites for cellular RNA-binding proteins that are thought to promote intermolecular architectures conducive to this process [4,5,6]. RNA synthesis initially yields RNA intermediates of negative polarity, which serve as templates for the production of genomic RNA. Nascent RNA genomes can serve as templates for further rounds of translation and RNA replication and, upon production of sufficient viral protein, are encapsidated to yield mature, infectious virions. The resulting viral progeny exit the cell via lysis and/or non-lytic release within extracellular vesicles [7]
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