title: Different types of nsP3-containing protein complexes in Sindbis virus-infected cells.
abstract: Alphaviruses represent a serious public health threat and cause a wide variety of diseases, ranging from severe encephalitis, which can result in death or neurological sequelae, to mild infection, characterized by fever, skin rashes, and arthritis. In the infected cells, alphaviruses express only four nonstructural proteins, which function in the synthesis of virus-specific RNAs and in modification of the intracellular environment. The results of our study suggest that Sindbis virus (SINV) infection in BHK-21 cells leads to the formation of at least two types of nsP3-containing complexes, one of which was found in association with the plasma membrane and endosome-like vesicles, while the second was coisolated with cell nuclei. The latter complexes could be solubilized only with the cytoskeleton-destabilizing detergent. Besides viral nsPs, in the mammalian cells, both complexes contained G3BP1 and G3BP2 (which were found in different ratios), YBX1, and HSC70. Rasputin, an insect cell-specific homolog of G3BP1, was found in the nsP3-containing complexes isolated from mosquito cells, which was suggestive of a high conservation of the complexes in the cells of both vertebrate and invertebrate origin. The endosome- and plasma membrane-associated complexes contained a high concentration of double-stranded RNAs (dsRNAs), which is indicative of their function in viral-RNA synthesis. The dsRNA synthesis is likely to efficiently proceed on the plasma membrane, and at least some of the protein-RNA complexes would then be transported into the cytosol in association with the endosome-like vesicular organelles. These findings provide new insight into the mechanism of SINV replication and virus-host cell interactions.
authors.affiliation: Department of Microbiology and Immunology, University of Texas Medical Branch, 301 University Boulevard, Galveston, TX 77555-1019, USA.
authors.forename: Rodion
authors.initials: R
authors.lastname: Gorchakov
authors.forename: Natalia
authors.initials: N
authors.lastname: Garmashova
authors.forename: Elena
authors.initials: E
authors.lastname: Frolova
authors.forename: Ilya
authors.initials: I
authors.lastname: Frolov
mesh_terms: D018354:Alphavirus Infections / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D009033:Culicidae; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D046912:Multiprotein Complexes / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012845:Sindbis Virus / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D005456:Fluorescent Dyes; D046912:Multiprotein Complexes; D012330:RNA, Double-Stranded; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; C089383:nsp3 protein, alphavirus; D012321:DNA-Directed RNA Polymerases