Electrophoretic Separation of Influenza Virus Ribonucleoproteins

Rees, P. J.; Dimmock, Nigel J.

doi:10.1099/0022-1317-53-1-125

Cited by 36 publications

(25 citation statements)

References 25 publications

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“…In addition, it should be taken into consideration that there may be a threshold concentration of M protein in the system, below which the complex cannot be formed. It should be noted that the ratio of M to NP in the complex which we have obtained resembles the one reported by Rees & Dimmock (1981) for RNP isolated from virions of the FPV Rostock strain, using milder conditions of isolation than in our experiments. These data and our results suggest that M protein-RNP complexes can exist under natural conditions as well.…”

Section: Discussionsupporting

confidence: 52%

In vitro Inhibition of Negative Strand Virus Transcriptase Activity by Proteins Soluble in Acidic Chloroform-Methanol

Mikhejeva¹,

Ghendon²

1983

Journal of General Virology

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SUMMARYThe effect of proteins soluble in acidic chloroform-methanol (ACMS proteins) on the transcriptase activity of virus ribonucleoproteins (RNPs) in vitro has been studied. Experiments with ACMS membrane (M) proteins from type A and B orthomyxoviruses, as well as from vesicular stomatitis virus, showed that inhibition of the viral RNP transcriptase activity occurred when they interacted with M proteins isolated from viruses of a different serotype, or even of a different family. The presence of ACMS proteins capable of inhibiting the transcriptase activity of orthomyxovirus RNP in vitro was also detected in human blood plasma and among proteins produced by human leukocytes. Determination of the minimum concentration of M protein inhibiting the RNP transcriptase activity, and analysis of the fowl plague virus M protein-RNP complex formed in the in vitro system, showed that the M protein was capable of inhibiting RNP transcriptase activity at a M:RNP ratio of 0.1 to 0.2:1.

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Section: Discussionsupporting

confidence: 52%

In vitro Inhibition of Negative Strand Virus Transcriptase Activity by Proteins Soluble in Acidic Chloroform-Methanol

Mikhejeva¹,

Ghendon²

1983

Journal of General Virology

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“…1. Letters a to e refer to RNPs a to e. All counts have been corrected for RNP size, based on the length of RNA contained in each RNP complex (Sleigh et al, 1979;Rees & Dimmock, 1981 a); RNPs a, b, c, d and e contain RNAs 1 to 3, RNA 4, RNAs 5 and 6, RNA 7, and RNA 8 respectively. labelled for 10 min at 4 h post-infection and chased for 60 min was in R N P structures.…”

Section: The Accumulation Of [35s]methionine Into Each Of the Five Simentioning

confidence: 99%

“…At each time post-infection one dish of cells was taken and nuclear and cytoplasmic fractions prepared using the nuclear monolayer technique (Hudson & Dimmock, 1977;Possee et al, 1982). Proteins were analysed by PAGE as described by Cook et aL (•979) using the buffer system of Laemmli (1970) and RNPs were separated on 3 to 4% acrylamide gradient gels (Rees & Dimmock, 1981 a) with a modified electrophoresis buffer containing 0.02 M-NaC1, 0.001 M-EDTA, 0.05 ra-tris-HC1 pH 7.4 and electrophoresis at 40 mA. All RNPs identified by this procedure co-migrated with those from virions and no others were found in infected cells (Rees & Dimmock, 1981 a, b).…”

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confidence: 99%

Kinetics of Synthesis of Influenza Virus Ribonucleoprotein Structures

Rees

Dimmock

1982

Journal of General Virology

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SUMMARYThe synthesis of influenza virus ribonucleoprotein structures (RNPs) in infected chick embryo cells was analysed by polyacrylamide gel electrophoresis (PAGE) in the presence of sodium deoxycholate which resolves the RNPs into five size classes. A relatively small proportion of total RNPs accumulated in the nucleus but free NP protein was found there in large amounts over the period 1.5 to 4 h post-infection. In contrast, by 4 h post-infection, all cytoplasmic NP was complexed into RNP structures. At early times, during a 15 min pulse of [aSSlmethionine, nearly all the newly synthesized NP was incorporated into RNPs but by 4 h the majority of pulse-labelled NP was present as free protein. However, the proportion of free NP:NP in RNPs remained constant over the 1.5 to 4 h post-infection period, indicating that there was a delay before the NP synthesized later in infection was assembled into RNP structures. Individual RNP size classes were predominantly cytoplasmic and accumulated at similar rates but were not produced in equimolar amounts. The rates of synthesis of individual RNPs were in general agreement with their rates of accumulation with the remarkable exception of RNP d (containing RNA 7, the matrix protein gene). This was synthesized nearly 10-fold faster but accumulated at the same rate as the other RNPs. Possibly RNP d is more rapidly degraded than the other RNPs.The influenza virus genome consists of eight separate segments of single-stranded negative-sense RNA (RNA-, McGeoch et al., 1976;Palese & Schulman, 1976). In the infected cell these RNA segments are transcribed into two forms of complementary plus-sense RNA (RNA+): a truncated copy which serves as messenger RNA (mRNA) to direct the synthesis of virus proteins and a complete transcript (cRNA) which serves as template for the synthesis of negative-sense virus RNA (Hay et al., 1977a, b). Where the various forms of virus-coded RNA are synthesized is unclear, but there is evidence that the cell nucleus is the probable site of transcription of virus mRNA (Armstrong & Barry, 1974;Taylor et al., 1977;Mark et al., 1978Mark et al., , 1979 Barrett et al., 1979).Within the virion the RNA segments are complexed with nucleoprotein (NP; Pons et al., 1969) to form separate ribonucleoprotein (RNP) complexes (Duesberg, 1969; Compans et al., 1972). In extracts from infected cells, RNPs containing both RNA + and RNA-have been found (Pons, 1971(Pons, , 1975 and it seems reasonable to suppose that all intracellular virus-induced RNA may be present in RNP complexes. Apart from trace amounts of other virus proteins the principal component of intracellular RNPs is the NP protein (Caliguiri & Gerstein, 1978; Rees & Dimmock, 1981 b).A number of studies has been made of the kinetics of synthesis and accumulation of virus-specific RNA (Scholtissek & Rott, 1970; Hay et al., 1977a, Barrett et al., 1978Mark et al., 1978) and proteins (Skehel, 1972(Skehel, , 1973 Meier-Ewert & Compans, 1974;Lamb & Choppin, 1976; Inglis et al., 1976 Inglis et al., , 1978 Inglis & Mahy,...

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“…M1 may also be involved in the inhibition of viral transcription in the late stages of infection and regulation of the switch from replication to viral assembly (36,59,60). M1 binds RNA (41,49,57,59), vRNPs (34,38,41,57), and lipids (7,15); dimerizes with other M1 molecules (16,41); and interacts with both the HA and neuraminidase proteins (1,11). It is also involved in export to the cytoplasmic membrane, virus assembly, and budding (12,19,31).…”

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Characterization of the 1918 “Spanish” Influenza Virus Matrix Gene Segment

Reid

Fanning

Janczewski

et al. 2002

J Virol

101

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The coding region of influenza A virus RNA segment 7 from the 1918 pandemic virus, consisting of the open reading frames of the two matrix genes M1 and M2, has been sequenced. While this segment is highly conserved among influenza virus strains, the 1918 sequence does not match any previously sequenced influenza virus strains. The 1918 sequence matches the consensus over the M1 RNA-binding domains and nuclear localization signal and the highly conserved transmembrane domain of M2. Amino acid changes that correlate with high yield and pathogenicity in animal models were not found in the 1918 strain. Phylogenetic analyses suggest that both genes were mammalian adapted and that the 1918 sequence is very similar to the common ancestor of all subsequent human and classical swine matrix segments. The 1918 sequence matches other mammalian strains at 4 amino acids in the extracellular domain of M2 that differ consistently between avian and mammalian strains, suggesting that the matrix segment may have been circulating in human strains for at least several years before 1918.

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Electrophoretic Separation of Influenza Virus Ribonucleoproteins

Cited by 36 publications

References 25 publications

In vitro Inhibition of Negative Strand Virus Transcriptase Activity by Proteins Soluble in Acidic Chloroform-Methanol

In vitro Inhibition of Negative Strand Virus Transcriptase Activity by Proteins Soluble in Acidic Chloroform-Methanol

Kinetics of Synthesis of Influenza Virus Ribonucleoprotein Structures

Characterization of the 1918 “Spanish” Influenza Virus Matrix Gene Segment

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