Respiratory synctial virus interactions with host-cell RNA-processing structures and proteins

Thesis (Ph.D.)--Boston UniversityRespiratory syncytial virus (RSV) is a negative-strand RNA virus that causes significant pneumonia-related morbidity and mortality worldwide. There are currently neither vaccines nor effective therapies for RSV. As with other viruses, RSV mRNAs are translated using host-cell machinery, rendering the virus subject to cellular factors that regulate mRNA homeostasis. Stress granules (SGs) and processing bodies (p-bodies) are inter-dependent, stress-response cytoplasmic structures involved in mRNA triage and degradation, respectively. We hypothesized that RSV has evolved to manipulate cellular stress responses in order to facilitate optimal virus propagation. While wild-type (wt) RSV induced SGs in approximately 1% of infected cells, a mutant version of RSV whose Tr region was replaced with an inverted LeC sequence (LeC virus) induced SG formation in approximately 50% to 70% of infected cells. A 12U to A substitution relative to the 5' end of the LeC virus abrogated SG induction. Mixed-infection studies showed that wt RSV was able to prevent LeC-mediated SG induction. Unlike Sendai virus, RSV-mediated prevention of SG formation was independent of SG-associated t-cell intracellular antigen related (TIAR) protein. RSV infection altered neither the number nor distribution of p-bodies; however, p-body-associated decapping protein 1 (dcp1) was phosphorylated throughout RSV infection via the extracellular signal-regulated kinase (ERK) 1/2 pathway. RSV-mediated dcp1 phosphorylation was limited to serine 315, serine 319, and threonine 321. Dcp1 phosphorylation occurred in response to some, but not all, environmental stresses, and dcp1 was not phosphorylated during infection with HIV-1, measles, mumps, or canine distemper virus. Overexpression of dcp1 significantly attenuated RSV cytopathic effects, and preliminary data suggested that dcp1 phosphorylation regulated RSV-induced interleukin-8 production. Finally, an antibody toward cellular SG- and p-body-associated, RNA-binding protein p54 was able to recognize a subset of RSV nucleoprotein (N). p54 and RSV N contain a similar amino acid sequence motif, suggesting that they may have similar or competing activities that are important during RSV replication. Taken together, our results demonstrate that RSV can manipulate cellular RNA-processing structures and proteins to facilitate viral propagation