Project description:Duck Tembusu virus (DTMUV) is a newly emerging pathogenic flavivirus that has caused huge economic losses to the duck industry in China since 2010. Moreover, DTMUV can also infect geese, chickens, house sparrows and replicate in in the brain, spleen, liver and kidneys of BALB/c mice, which poses public health concerns. So developing vaccines to combat DTMUV becomes urgent. Baby hamster kidney cell line (BHK-21) is usually employed to produce veterinary vaccine and DTMUV as well as other flaviviruses can be propagated in BHK-21 cells. However, information about mammalian host cell responses to DTMUV infection is limited. In this study, LC–MS/MS coupled to iTRAQ labeling was used to quantitatively identify differentially expressed cellular proteins in DTMUV-infected BHK-21 cells. We identified 192 differentially expressed cellular proteins including 11 upregulated proteins and 8 downregulated at 24 hpostinfection; 25 upregulated proteins and 151 downregulated at 48 h postinfection. Some of these proteins were involved in viral infection.
Project description:<p>Tick-borne encephalitis virus is an enveloped, pathogenic, RNA virus in the family Flaviviridae, genus Flavivirus. Viral particles are formed when the nucleocapsid, consisting of an RNA genome and multiple copies of the capsid protein, buds through the endoplasmic reticulum membrane and acquires the viral envelope and the associated proteins. The coordination of the nucleocapsid components to the sites of assembly and budding are poorly understood. Here, we investigate nucleocapsid assembly by characterizing the interactions of the wild-type and truncated capsid proteins with membranes by using biophysical methods and model membrane systems. We show that capsid protein initially binds membranes via electrostatic interactions with negatively-charged lipids which is followed by membrane insertion. Additionally, we show that membrane-bound capsid protein can recruit viral genomic RNA. We confirm the biological relevance of the biophysical findings by using mass spectrometry to show that purified virions contain negatively-charged lipids. Our results suggest that nucleocapsid assembly is coordinated by negatively-charged membrane patches on the endoplasmic reticulum and that the capsid protein mediates direct contacts between the nucleocapsid and the membrane.</p>
Project description:Small RNA libraries were constructed from total RNA from Jasminum sambac plants exhibiting virus-like symptoms. After sequencing, small RNAs were assembled into contigs with MetaVelvet and assembled contigs were aligned against the NR database of NCBI using BLASTx. Top hits that reported a virus as subject were considered putative viral sequences. Based on such alignments, the whole genome of a virus, we tentatively name Jasmine Virus H was recovered and cloned. Two more small RNA libraries were made in a confirmatory experiment. One from Jasminum sambac and another one from Nicotiana benthamiana plants infected with the newly-cloned virus. The small RNA libraries were aligned against the full-length sequence of Jasmine Virus H to determine the spacial distribution of virus-derived small RNAs along the virus genome.