title: SH3 domain-mediated recruitment of host cell amphiphysins by alphavirus nsP3 promotes viral RNA replication.
abstract: Among the four non-structural proteins of alphaviruses the function of nsP3 is the least well understood. NsP3 is a component of the viral replication complex, and composed of a conserved aminoterminal macro domain implicated in viral RNA synthesis, and a poorly conserved carboxyterminal region. Despite the lack of overall homology we noted a carboxyterminal proline-rich sequence motif shared by many alphaviral nsP3 proteins, and found it to serve as a preferred target site for the Src-homology 3 (SH3) domains of amphiphysin-1 and -2. Nsp3 proteins of Semliki Forest (SFV), Sindbis (SINV), and Chikungunya viruses all showed avid and SH3-dependent binding to amphiphysins. Upon alphavirus infection the intracellular distribution of amphiphysin was dramatically altered and colocalized with nsP3. Mutations in nsP3 disrupting the amphiphysin SH3 binding motif as well as RNAi-mediated silencing of amphiphysin-2 expression resulted in impaired viral RNA replication in HeLa cells infected with SINV or SFV. Infection of Balb/c mice with SFV carrying an SH3 binding-defective nsP3 was associated with significantly decreased mortality. These data establish SH3 domain-mediated binding of nsP3 with amphiphysin as an important host cell interaction promoting alphavirus replication.
authors: Institute of Biotechnology, University of Helsinki, Helsinki, Finland.
authors: Maarit
authors: M
authors: Neuvonen
authors: Arunas
authors: A
authors: Kazlauskas
authors: Miika
authors: M
authors: Martikainen
authors: Ari
authors: A
authors: Hinkkanen
authors: Tero
authors: T
authors: Ahola
authors: Kalle
authors: K
authors: Saksela
mesh_terms: D000524:Alphavirus / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002646:Chikungunya virus / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008024:Ligands; D051379:Mice; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D012367:RNA, Viral / Q000096:biosynthesis*; D012672:Semliki forest virus / Q000235:genetics; D012845:Sindbis Virus / Q000235:genetics; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D014779:Virus Replication / Q000235:genetics*; D018909:src Homology Domains / Q000235:genetics
chemical_list: D008024:Ligands; D009419:Nerve Tissue Proteins; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; C089383:nsp3 protein, alphavirus; C075637:amphiphysin