Recombinant Measles Viruses with Mutations in the C, V, or F Gene Have Altered Growth Phenotypes In Vivo

摘要: Measles virus (MV), a negative-strand RNA virus, was originally propagated in tissue culture by Enders and Peebles 1954 (15). However, investigation of the molecular determinants MV replication has been difficult, functions number genes control regions remain unknown. The rescue infectious from transfected cDNA clone enabled construction first recombinant mutant MVs (39) greatly facilitated such studies. viruses examined to date grow identically parent Vero cells (38, 39, 43), suggesting that mutated are dispensable for this cell line. effects these mutations on vivo not understood, part because there is no small-animal model measles. We have previously used human thymus xenograft, SCID-hu thy/liv model, characterize pathologic changes induced vaccine wild-type strains known differ virulence (2). Thymus implants created coinoculation liver under renal capsule SCID mouse. give rise thymic microenvironment, provide source hematopoietic precursors populate developing (33). Three four months postengraftment, structurally functionally normal formed. This study other lack models, including immunodeficiency (HIV) (4, 22), cytomegalovirus (5, 30), varicella-zoster (31, 32). The implant relevant examining since infects natural disease. antigens viral cytopathic effect found at autopsy following acute infection humans (50) necropsy after experimental primates (42). In vitro, primary cultures stromal support growth (34). addition, phenotypes faithfully reproduced Infection with minimally passaged patient isolate Chicago-89 (Chi-1) strain results high levels macrophages 3 days massive thymocyte death. contrast, an attenuated Moraten slow causes little death. We utilized system investigate role 5′ untranslated region F gene (F 5′UTR), C protein, V protein vivo. Among paramyxoviruses, only mRNAs morbilliviruses contain long 460- 580-nucleotide GC-rich between transcription start site methionine initiation codon. predicted extensive secondary structure (41). Experiments define its function suggest 5′UTR acts as focusing factor, directing translation second clustered AUGs (8). affects efficiency translation. DNA vaccination studies, required effective anti-F antibody response mice therefore presumably expression (47). It also rinderpest canine distemper F-protein immortalized using vaccinia virus-encoded T7 polymerase express mRNA (16). partial deletion increased (10), rabbit reticulocyte lysates, inhibits (16), can inhibit translation. The small (∼20-kDa), positively charged encoded within P open reading frame (ORF) paramyxoviruses but rubulaviruses. expresses single alternative downstream initiator ORF (3). Sendai (SeV) infection, nested set proteins, (C′, C, Y1, Y2) different sites common COOH termini expressed (13, 18, 36). A studies failed identify purified virions, leading designation nonstructural protein. recent SeV amounts associated nucleocapsid virions (51). infected cells, nucleus cytoplasm, where it colocalizes nucleocapsids clear. Studies antigenomic synthesis (6, 45), perhaps through direct binding L, catalytic subunit (19). Whether regulates similar manner unknown. The ∼40-kDa all except respiratory syncytial parainfluenza types 1 3. paramyxovirus morbillivirus families, synthesized result slippage distinct which leads pseudotemplated insertion G residue (9, 12, 48). Thus, amino terminus (231 acids MV) identical P, acidic, highly phosphorylated. carboxy unique (68 MV), conserved among contains cysteine-rich zinc finger domain binds (27). distributed diffusely cytoplasm does colocalize (12, 49). Analysis suggests genome soluble nucleoprotein (20). coimmunoprecipitation appears bind cellular proteins (28), relevance interactions yet clear. The rubulaviruses encode proteins. Rubulavirus structural associate (35, 44). They directly (44, 46) shorter, basic termini. simian 5, (26, 37, 40). Like those MV, rubulavirus cysteine rich (35). MV mutants or abrogating (C− V− mutants) like unaltered variety lines (14). To determine whether play vivo, we studied model. mutation two- fivefold increase characterized. may be accessory elements efficient peak titers were lower than parent. Production delayed prolonged absence V, while excess resulted more rapid replication. Data showing survival despite production large late additional pathogenesis thymus.