Giuseppe
Emeritus
J Virol. 2008 Nov 5. [Epub ahead of print]
Studies of an influenza A virus temperature sensitive mutant identify a 'late' role for NP in the formation of infectious virions.
Noton SL, Simpson-Holley M, Medcalf E, Wise HM, Hutchinson EC, McCauley JW, Digard P. - Division of Virology, Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QP, United Kingdom; and Institute for Animal Health, Compton Laboratory, Berkshire RG20 7NN, United Kingdom.
The influenza A virus nucleoprotein (NP) is a single-strand RNA binding protein that encapsidates the virus genome and has essential functions in viral RNA synthesis.
Here, we report the characterisation of a temperature sensitive (ts) NP mutant (US3) originally generated in fowl plague virus (A/chicken/Rostock/34).
Sequence analysis revealed a single mutation, M239L, in NP, consistent with earlier mapping studies assigning the ts lesion to segment 5.
Introduction of this mutation into A/PR/8/34 virus by reverse genetics produced a ts phenotype, confirming the identity of the lesion.
Despite an approximately 100-fold drop in viral titre at the non permissive temperature, the mutant US3 polypeptide supported wild type (WT) levels of genome transcription, replication and protein synthesis, indicating a 'late' stage defect in function of the NP polypeptide.
Nucleocytoplasmic trafficking of the US3 NP was also normal, and the virus actually assembled and released around six-fold more virus particles than the WT virus, with normal vRNA content.
However, the particle
fu ratio of these virions was 50-fold higher than that of WT virus and many particles exhibited an abnormal morphology.
Reverse genetics studies in which the A/PR/8/34 segment 7 was swapped with sequences from other strains of virus revealed a profound incompatibility between the M239L mutation and the A/Udorn/72 M1 gene, suggesting the ts mutation affects M1-NP interactions.
Thus we have identified a 'late' acting defect in NP that, separate from its function in RNA synthesis, indicates a role for the polypeptide in virion assembly, most likely involving M1 as a partner.
PMID: 18987140 [PubMed - as supplied by publisher]
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Studies of an influenza A virus temperature sensitive mutant identify a 'late' role for NP in the formation of infectious virions.
Noton SL, Simpson-Holley M, Medcalf E, Wise HM, Hutchinson EC, McCauley JW, Digard P. - Division of Virology, Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QP, United Kingdom; and Institute for Animal Health, Compton Laboratory, Berkshire RG20 7NN, United Kingdom.
The influenza A virus nucleoprotein (NP) is a single-strand RNA binding protein that encapsidates the virus genome and has essential functions in viral RNA synthesis.
Here, we report the characterisation of a temperature sensitive (ts) NP mutant (US3) originally generated in fowl plague virus (A/chicken/Rostock/34).
Sequence analysis revealed a single mutation, M239L, in NP, consistent with earlier mapping studies assigning the ts lesion to segment 5.
Introduction of this mutation into A/PR/8/34 virus by reverse genetics produced a ts phenotype, confirming the identity of the lesion.
Despite an approximately 100-fold drop in viral titre at the non permissive temperature, the mutant US3 polypeptide supported wild type (WT) levels of genome transcription, replication and protein synthesis, indicating a 'late' stage defect in function of the NP polypeptide.
Nucleocytoplasmic trafficking of the US3 NP was also normal, and the virus actually assembled and released around six-fold more virus particles than the WT virus, with normal vRNA content.
However, the particle
fu ratio of these virions was 50-fold higher than that of WT virus and many particles exhibited an abnormal morphology.Reverse genetics studies in which the A/PR/8/34 segment 7 was swapped with sequences from other strains of virus revealed a profound incompatibility between the M239L mutation and the A/Udorn/72 M1 gene, suggesting the ts mutation affects M1-NP interactions.
Thus we have identified a 'late' acting defect in NP that, separate from its function in RNA synthesis, indicates a role for the polypeptide in virion assembly, most likely involving M1 as a partner.
PMID: 18987140 [PubMed - as supplied by publisher]
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