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Human parainfluenza virus type 3: messenger RNAs, polypeptide coding assignments, intergenic sequences, and genetic
Abstract:
cDNA clones of mRNAs for the major nucleocapsid protein (NP), the nucleocapsid P protein plus the nonstructural C protein (P+C), and the matrix protein (M) of human parainfluenza virus type 3 (PF3) were identified by hybrid arrest and hybrid selection of in vitro translation. Previously, cDNA clones were identified and sequenced for the hemagglutinin-neuraminidase glycoprotein (HN) and the fusion glycoprotein (F) mRNAs (N. Elango, J. E. Coligan, R. C. Jambou, and S. Venkatesan, J. Virol. 57:481-489, 1986; M. K. Spriggs, R. A. Olmsted, S. Venkatesan, J. E. Coligan, and P. L. Collins, Virology 152:241-251, 1986). Synthetic oligonucleotides, designed from nucleotide sequences of the cDNAs, were used to direct dideoxynucleotide sequencing of gene junctions in PF3 genomic RNA (vRNA). From sequencing of vRNA, a sixth viral gene was detected and identified as the large nucleocapsid protein (L) gene by hybridization of a synthetic oligonucleotide to intracellular PF3 mRNAs separated by gel electrophoresis. The order of the six PF3 genes on vRNA was 3'-NP-P+C-M-F-HN-L-5'. The five intergenic regions consisted of the trinucleotide 3'-GAA. The PF3 genes initiated with semiconserved 10-nucleotide gene-start sequences and terminated with semiconserved 12-nucleotide gene-end sequences. The M gene terminated with an aberrant gene-end sequence; analysis of intracellular mRNA showed that this aberrant sequence correlated with a disproportionately high accumulation of readthrough mRNA. These studies showed that PF3 encodes six unique mRNAs (NP, P+C, M, F, HN, and L) that encode seven proteins (NP, P, C, M, F, HN, and L) and provided evidence of a close relationship between PF3 and Sendai (murine parainfluenza type 1) viruses.
Insights
This study identifies six genes in human parainfluenza virus type 3 (PF3), detailing their order and regulatory sequences. These findings reveal the virus
Area of Science:
- Virology
- Molecular Biology
- Genomics
Background:
- Human parainfluenza virus type 3 (PF3) is a significant respiratory pathogen.
- Previous research had identified cDNA clones for several PF3 mRNAs, including those for the hemagglutinin-neuraminidase glycoprotein (HN) and fusion glycoprotein (F).
Purpose of the Study:
- To identify and characterize the complete set of PF3 viral genes and their organization on the viral RNA (vRNA).
- To elucidate the gene order, intergenic regions, and gene-termination signals of PF3.
- To investigate potential readthrough transcription events.
Main Methods:
- Hybrid arrest and hybrid selection of in vitro translation were used to identify cDNA clones for nucleocapsid protein (NP), P+C, and matrix protein (M) mRNAs.
- Synthetic oligonucleotides and dideoxynucleotide sequencing were employed to analyze gene junctions in PF3 genomic RNA (vRNA).
- Oligonucleotide hybridization to separated intracellular PF3 mRNAs identified the sixth viral gene (L).
Main Results:
- Six unique PF3 mRNAs were identified, encoding seven proteins: NP, P, C, M, F, HN, and L.
- The gene order on the vRNA was established as 3'-NP-P+C-M-F-HN-L-5'.
- Intergenic regions contained the trinucleotide 3'-GAA, and genes initiated with conserved gene-start and terminated with conserved gene-end sequences, with an aberrant termination signal in the M gene.
Conclusions:
- The complete gene content and organization of PF3 were elucidated.
- The findings provide insights into PF3 gene expression regulation, including transcription initiation, termination, and potential readthrough.
- A close evolutionary relationship between PF3 and Sendai viruses was suggested.