Effect of intragenic rearrangement and changes in the 3' consensus sequence on NSP1 expression and rotavirus replication.
The nonpolyadenylated mRNAs of rotavirus are templates for the synthesis of protein and the segmented double-stranded RNA (dsRNA) genome. During serial passage of simian SA11 rotaviruses in cell culture, two variants emerged with gene 5 dsRNAs containing large (1.1 and 0.5 kb) sequence duplications within the open reading frame (ORF) for NSP1. Due to the sequence rearrangements, both variants encoded only C-truncated forms of NSP1. Comparison of these and other variants encoding defective NSP1 with their corresponding wild-type viruses indicated that the inability to encode authentic NSP1 results in a small-plaque phenotype. Thus, although nonessential, NSP1 probably plays an active role in rotavirus replication in cell culture. In determining the sequences of the gene 5 dsRNAs of the SA11 variants and wild-type viruses, it was unexpectedly found that their 3' termini ended with 5'-UGAACC-3' instead of the 3' consensus sequence 5'-UGACC-3', which is present on the mRNAs of nearly all other group A rotaviruses. Cell-free assays indicated that the A insertion into the 3' consensus sequence interfered with its ability to promote dsRNA synthesis and to function as a translation enhancer. The results provide evidence that the 3' consensus sequence of the gene 5 dsRNAs of SA11 rotaviruses has undergone a mutation causing it to operate suboptimally in RNA replication and in the expression of NSP1 during the virus life cycle. Indeed, just as rotavirus variants which encode defective NSP1 appear to have a selective advantage over those encoding wild-type NSP1 in cell culture, it may be that the atypical 3' end of SA11 gene 5 has been selected for because it promotes the expression of lower levels of NSP1 than the 3' consensus sequence.
Patton
J T
JT
Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, Bethesda, Maryland 20892, USA. jpatton@atlas.niaids.nih.gov
Taraporewala
Z
Z
Chen
D
D
Chizhikov
V
V
Jones
M
M
Elhelu
A
A
Collins
M
M
Kearney
K
K
Wagner
M
M
Hoshino
Y
Y
Gouvea
V
V
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D016384:Consensus Sequence / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D008969:Molecular Sequence Data; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism; D012401:Rotavirus / Q000235:genetics / Q000502:physiology*; D012400:Rotavirus Infections / Q000821:virology; D017422:Sequence Analysis, DNA; D017361:Viral Nonstructural Proteins / Q000235:genetics* / Q000378:metabolism*; D010948:Viral Plaque Assay; D014779:Virus Replication
D018076:DNA, Complementary; D012330:RNA, Double-Stranded; D017361:Viral Nonstructural Proteins; C104282:nsp1 protein, Rotavirus