Showing papers by "Karla Kirkegaard published in 2001"

Poliovirus 3A protein limits interleukin-6 (IL-6), IL-8, and beta interferon secretion during viral infection.

Abstract: During viral infections, the host secretory pathway is crucial for both innate and acquired immune responses. For example, the export of most proinflammatory and antiviral cytokines, which recruit lymphocytes and initiate antiviral defenses, requires traffic through the host secretory pathway. To investigate potential effects of the known inhibition of cellular protein secretion during poliovirus infection on pathogenesis, cytokine secretion from cells infected with wild-type virus and with 3A-2, a mutant virus carrying an insertion in viral protein 3A which renders the virus defective in the inhibition of protein secretion, was tested. We show here that cells infected with 3A-2 mutant virus secrete greater amounts of cytokines interleukin-6 (IL-6), IL-8, and beta interferon than cells infected with wild-type poliovirus. Increased cytokine secretion from the mutant-infected cells can be attributed to the reduced inhibition of host protein secretion, because no significant differences between 3A-2- and wild-type-infected cells were observed in the inhibition of viral growth, host cell translation, or the ability of wild-type- or 3A-2-infected cells to support the transcriptional induction of beta interferon mRNA. We surmise that the wild-type function of 3A in inhibiting ER-to-Golgi traffic is not required for viral replication in tissue culture but, by altering the amount of secreted cytokines, could have substantial effects on pathogenesis within an infected host. The global inhibition of protein secretion by poliovirus may reflect a general mechanism by which pathogens that do not require a functional protein secretory apparatus can reduce the native immune response and inflammation associated with infection.

Oligomeric structures of poliovirus polymerase are important for function.

Abstract: Central to the replication of poliovirus and other positive-strand RNA viruses is the virally encoded RNA-dependent RNA polymerase. Previous biochemical studies have suggested that direct polymerase– polymerase interactions might be important for polymerase function, and the structure of poliovirus polymerase has revealed two regions of extensive polymerase–polymerase interaction. To explore potential functional roles for the structurally observed polymerase–polymerase interactions, we have performed RNA binding and extension studies of mutant polymerase proteins in solution, disulfide cross-linking studies, mutational analyses in cells, in vitro activity analyses and RNA substrate modeling studies. The results of these studies indicate that both regions of polymerase–polymerase interaction observed in the crystals are indeed functionally important and, furthermore, reveal specific functional roles for each. One of the two regions of interaction provides for efficient substrate RNA binding and the second is crucial for forming catalytic sites. These studies strongly support the hypothesis that the polymerase– polymerase interactions discovered in the crystal structure provide an exquisitely detailed structural context for poliovirus polymerase function and for poliovirus RNA replication in cells.

Inhibitors of viral infection

Abstract: The present invention provides a viral RNA-dependent RNA polymerase pharmacophore which is characterized by binding to a conserved interface binding surfaces of a viral RNA-dependent RNA polymerase.

Viral vectors useful in induction of humoral or cellular immunity

Abstract: The invention features recombinant viral vectors that take advantage of the activity of picornaviral protein 3A in modulating cytokine secretion and antigen presentation on MHC Class I (MHC I), which in turn provides for modulation of a Th1-mediated immune response to the transfected host cell Specifically, a recombinant viral vector comprising a sequence encoding picornaviral protein 3A provides for decreased antigen presentation on MHC I and a decreased incidence of Th1-mediated immune response, while a recombinant picornaviral vector that is deficient in protein 3A production provides for increased cytokine secretion, increased antigen presentation on MHC I and increased Th1-mediated immune response towards the transfected host cell The invention also features methods of inducing humoral or cellular immunity using the vectors