The Various Sendai Virus C Proteins Are Not Functionally Equivalent and Exert both Positive and Negative Effects on Viral RNA Accumulation during the Course of Infection

Abstract

Recombinant Sendai viruses were prepared which cannot express their Cprime, C, or Cprime plus C proteins due to mutation of their respective start codons ([Cprime-minus], [C-minus] and [double mutant], respectively). The [Cprime-minus] and [C-minus] stocks were similar to that of wild-type (wt) virus in virus titer and plaque formation, whereas the double-mutant stock had a much-reduced PFU or 50% egg infective dose/particle ratio and produced very small plaques. Relative to the wt virus infection, the [Cprime-minus] and [C-minus] infections of BHK cells resulted in significantly greater accumulation of viral RNAs, consistent with the known inhibitory effects of the Cprime and C proteins. The double-mutant infection, in contrast, was delayed in its accumulation of viral RNAs; however, once accumulation started, overaccumulation quickly occurred, as in the single-mutant infections. Our results suggest that the Cprime and C proteins both provide a common positive function early in infection, so that only the double mutant undergoes delayed RNA accumulation and exhibits the highly debilitated phenotype. Later in infection, the same proteins appear to act as inhibitors of RNA accumulation. In infections of mice, [Cprime-minus] was found to be as virulent as wt virus whereas [C-minus] was highly attenuated. These results suggest that the Cprime and C proteins cannot be functionally equivalent, since C can replace Cprime for virulence in mice whereas Cprime cannot replace C.

Paramyxovirus Accessory Proteins as Interferon Antagonists

“…Similar but distinct conclusions were drawn by Garcin et al (42). They succeeded in generating a series of recombinant viruses that do not express a subset of C proteins or that express the C proteins with a mutation of a single amino acid change, but failed to isolate four-Cknockout virus (69). Not only a double mutant similar to C/C'( -) virus but also C -virus, which was generated by mutations of the initiation codon AUG I 14 to GCG 11 \ lost the inhibitory effect on IFN-~-stimulated induction of Stat I and on establishment of anti-VSV state.…”

“…Although gene expression, replication and cytopathogenicity of recombinant V-knockout (V( -)) virus in cell lines were either potentiated or comparable to those of wild type virus, the V( -) virus was strikingly attenuated in capacity of in vivo replication and pathogenicity (24,58). Knockout of the C gene more severely affected virus growth (50,67,69). Two types of mutants were generated; C/C'( -) virus, which does not express C and C' but does express Y1 and Y2, and 4C( -) virus, which does not express any of four C proteins (50,67).…”

Inhibition of interferon induction and signaling by paramyxoviruses

“…Recombinant viruses defective in C expression have been generated for several species, demonstrating that C is dispensable in cell culture (95,96). However, many of these C-deficient viruses demonstrated altered growth characteristics and attenuation in vitro (84,97,98), and all of them were attenuated in animal models, indicating that C proteins are virulence factors (82,98).…”

Paramyxovirus replication and pathogenesis. Reverse genetics transforms understanding

“…SeV C protein was initially recognised as a downmodulator of viral mRNA synthesis by using synthetic mini-genomes as templates. 68,69 More direct support for this down-regulatory role of C proteins came from the observations that infections of C ( ), C( ) and C/C ( ) viruses created by Latorre et al 66 and our 4C( ) all accumulated significantly more viral transcripts late in infection, compared with the wild-type infection. The C( ) overexpressed Y1 and Y2, and the C/C ( ) overexpressed Y2, but Y1 and Y2 did not appear to be able to replace C and/or C for downregulation of transcription.…”

“…65 The results conclusively demonstrated that C proteins greatly contribute to full replication of SeV but can be categorised as nonessential gene products. Failure to isolate 4C( ) viruses by Latorre et al 66 might be simply due to a low efficiency of their virus rescue system.…”