The NSs protein of Bunyamwera virus (Bunyaviridae) is an antiapoptotic interferon antagonist involved in silencing host protein expression by interfering with mRNA synthesis. Here, we show that the ability to inhibit both host transcription and the interferon response is linked to interaction of NSs with the MED8 component of Mediator, a protein complex necessary for mRNA production. The interacting domain on NSs was mapped to the C-terminal region, which contains amino acids conserved among orthobunyavirus NSs proteins. A recombinant virus in which the interacting domain in NSs was deleted had strongly reduced ability to inhibit host protein expression and was unable to inhibit the interferon response. This study provides further information on the mechanisms by which bunyavirus nonstructural proteins are involved in pathogenesis.

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The family Bunyaviridae contains over 350 named isolates, classified into five genera: Orthobunyavirus, Hantavirus, Nairovirus, Phlebovirus and Tospovirus. The Orthobunyavirus genus contains some 170 isolates that are mainly transmitted by mosquitoes and are responsible for a range of disease syndromes in humans including self-limiting febrile illness, encephalitis and haemorrhagic fever. The viruses have a tripartite, negative-sense RNA genome. Analyses of viruses in four serogroups (Bunyamwera, California, Group C and Simbu) showed that the smallest (S) RNA segment encodes the nucleocapsid protein (N) and a non-structural protein called (NSs). The NSs protein of Bunyamwera virus (BUNV) has been shown to play a role in shut-off of host cell protein synthesis in mammalian cells, but no protein shut-off is observed in BUNVinfected mosquito cells (Aedes albopictus C6/36 cells). Protein shut-off in infected mammalian cells is achieved by global inhibition of RNA polymerase II-mediated transcription and enables the virus to overcome the host innate immune response. As innate defence mechanisms constitute a significant barrier to virus infection of different hosts, NSs would appear to play a key role in determining the zoonotic capacity of orthobunyaviruses.

Artificial minigenomes are powerful tools for studying the replication and transcription of negative-strand RNA viruses. Bunyamwera virus (BUN; genus Orthobunyavirus, family Bunyaviridae) is an arbovirus that shows fundamental biological differences when replicating in mammalian versus mosquito cells. To study BUN RNA synthesis in mosquito cells, we developed a bacteriophage T7 RNA polymerase-based minireplicon system similar to that described previously for mammalian cells. An Aedes albopictus C6/36-derived mosquito cell line stably expressing T7 RNA polymerase was established. Viral proteins and artificial minigenomes (containing Renilla luciferase as a reporter) were transcribed and expressed in these cells from transfected T7 promotercontaining plasmids. Transcription of the minigenome required two viral proteins, the nucleocapsid protein N and the RNA-dependent RNA polymerase L, a situation similar to that in mammalian cells. However, unlike the situation in mammalian cells, the viral polymerase was not inhibited by the viral nonstructural protein NSs. We also report that promoter strength is different for vertebrate versus invertebrate cells. The development of this system opens the way for a detailed comparison of bunyavirus replication in cells of disparate phylogeny.

performed with DBP in a radioactively contaminated electrodynamic balance yielded similar results except that mass losses as large as 75% and charge losses as large as 63% occurred. The mean value of q-/qRL was 0.73 for these data, which is not statistically different from the previous result. It is likely that the electric fields used to suspend the droplets affect the stability phenomenon, that random thermal fluctuations in local surface charge density can contribute only in a minor way to the instability, and that surface contamination can alter the fragmentation of the exploding droplet.Differential scanning calorimetry (DSC) and 'H NMR spectroscopy were used to study various lyotropic liquid crystalline phases of sodium bis(2-ethylhexyl)sulfosuccinate (Aerosol OT or AOT) and water. At 25 "C the lamellar liquid crystalline phase contains between surfactant bilayers bulklike water ("water l"), which has a melting point Tlm = 0 "C and an enthalpy of melting AHlm = 80 cal/g. Close to the bilayers, the lamellar liquid crystalline phase contains interfacial water, "water 2", which melts at TZm = -4 f 2 "C and has AH2" = 35 f 8 cal/g of water. After most of water 1 and 2 freezes, the surfactant and the remaining liquid water undergo a phase transition to a nonlamellar liquid crystalline phase. In this phase, "water 3" melts at T3"' = -9 1 "C and has AH3m = 12 f 5 cal/g of water. When water is in ultrathin (510 A) films in lamellar liquid crystal, then water 1 is absent and water 2 can be supercooled to -45 "C or lower. The surfactant has substantial rotational mobility even when the water is frozen at temperatures from -35 to -10 "C. Similar phase and thermal behavior is observed when initially isotropic aqueous micellar solutions of AOT are frozen or melted.

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