scholarly journals Determination of the orientation of an integral membrane protein and sites of glycosylation by oligonucleotide-directed mutagenesis: influenza B virus NB glycoprotein lacks a cleavable signal sequence and has an extracellular NH2-terminal region.

1986 ◽  
Vol 6 (12) ◽  
pp. 4317-4328 ◽  
Author(s):  
M A Williams ◽  
R A Lamb
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Signal Sequence ◽  
Integral Membrane Protein ◽  
Terminal Region ◽  
Influenza B Virus ◽  
Influenza B ◽  
Directed Mutagenesis ◽  
Cell Fractionation ◽  
B Virus

The membrane orientation of the NB protein of influenza B virus, a small (Mr, approximately 18,000) glycoprotein with a single internal hydrophobic domain, was investigated by biochemical and genetic means. Cell fractionation and protein solubility studies indicate NB is an integral membrane protein, and NB has been shown to be a dimer under nonreducing conditions. Treatment of infected-cell surfaces with proteinase K and endoglycosidase F and immunoprecipitation with a site-specific antibody suggests that the 18-amino-acid NH2-terminal region of NB is exposed at the cell surface. Oligonucleotide-directed mutagenesis to eliminate each of the four potential sites of N-linked glycosylation and expression of the mutant NB proteins in eucaryotic cells suggest that the two sites adjacent to the NH2 terminus are glycosylated. This provides further evidence that NB, which lacks a cleavable NH2-terminal signal sequence, has an exposed NH2 terminus at the cell surface.

Determination of the orientation of an integral membrane protein and sites of glycosylation by oligonucleotide-directed mutagenesis: influenza B virus NB glycoprotein lacks a cleavable signal sequence and has an extracellular NH2-terminal region

1986 ◽  
Vol 6 (12) ◽  
pp. 4317-4328
Author(s):  
M A Williams ◽  
R A Lamb
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Signal Sequence ◽  
Integral Membrane Protein ◽  
Terminal Region ◽  
Influenza B Virus ◽  
Influenza B ◽  
Directed Mutagenesis ◽  
Cell Fractionation ◽  
B Virus

The membrane orientation of the NB protein of influenza B virus, a small (Mr, approximately 18,000) glycoprotein with a single internal hydrophobic domain, was investigated by biochemical and genetic means. Cell fractionation and protein solubility studies indicate NB is an integral membrane protein, and NB has been shown to be a dimer under nonreducing conditions. Treatment of infected-cell surfaces with proteinase K and endoglycosidase F and immunoprecipitation with a site-specific antibody suggests that the 18-amino-acid NH2-terminal region of NB is exposed at the cell surface. Oligonucleotide-directed mutagenesis to eliminate each of the four potential sites of N-linked glycosylation and expression of the mutant NB proteins in eucaryotic cells suggest that the two sites adjacent to the NH2 terminus are glycosylated. This provides further evidence that NB, which lacks a cleavable NH2-terminal signal sequence, has an exposed NH2 terminus at the cell surface.

scholarly journals Influenza B virus BM2 protein is an oligomeric integral membrane protein expressed at the cell surface

Virology ◽  
2003 ◽  
Vol 306 (1) ◽  
pp. 7-17 ◽  
Author(s):  
Reay G Paterson ◽  
Makoto Takeda ◽  
Yuki Ohigashi ◽  
Lawrence H Pinto ◽  
Robert A Lamb
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Integral Membrane Protein ◽  
Influenza B Virus ◽  
Influenza B ◽  
B Virus

scholarly journals Influenza B Virus BM2 Protein Is Transported through the trans-Golgi Network as an Integral Membrane Protein

2003 ◽  
Vol 77 (19) ◽  
pp. 10630-10637 ◽  
Author(s):  
Shinji Watanabe ◽  
Masaki Imai ◽  
Yoshiro Ohara ◽  
Takato Odagiri
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Matrix Protein ◽  
Integral Membrane Protein ◽  
Influenza B Virus ◽  
Influenza B ◽  
Hydrophobic Domain ◽  
Cytoplasmic Transport ◽  
B Virus ◽  
Golgi Network

ABSTRACT A bicistronic mRNA transcribed from the influenza B virus RNA segment 7 encodes two viral proteins, matrix protein M1 and uncharacterized small protein BM2. In the present study, we focused on the cytoplasmic transport and cellular membrane association of BM2. Immunofluorescence studies of virus-infected cells indicated that BM2 accumulated at the Golgi apparatus immediately after synthesis and then was transported to the plasma membrane through the trans-Golgi network. Localization of a set of BM2 deletion mutants revealed that the N-terminal half of BM2 (residues 2 to 50) was crucial for its transport; in particular, the deletion of residues 2 to 23, deduced to be a transmembrane domain, resulted in diffused distribution of the protein throughout the entire cell. Sucrose gradient flotation and biochemical analyses of the membrane showed that BM2 was tightly associated with cellular membranes as an integral membrane protein. Oligomerization of BM2 was demonstrated by coprecipitation of differentially epitope-tagged BM2 proteins. Taken together, these results strongly suggest that BM2 is integrated into the plasma membrane at the N-terminal hydrophobic domain as fourth membrane protein, in addition to hemagglutinin, neuraminidase, and NB, of the influenza B virus.

Lipocortin 1 (annexin 1) in patches associated with the membrane of a lung adenocarcinoma cell line and in the cell cytoplasm

1998 ◽  
Vol 111 (10) ◽  
pp. 1405-1418 ◽  
Author(s):  
V. Traverso ◽  
J.F. Morris ◽  
R.J. Flower ◽  
J. Buckingham
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Lung Adenocarcinoma ◽  
Western Blotting ◽  
Signal Sequence ◽  
Membrane Surface ◽  
Integral Membrane Protein ◽  
Subcellular Fractionation ◽  
Cell Fractionation ◽  
Electron Microscopic

Lipocortin 1 (annexin I) is a calcium- and phospholipid-binding annexin protein which can be externalised from cells despite the lack of a signal sequence. To determine its cellular distribution lipocortin 1 in A549 human lung adenocarcinoma cells was localised by light- and electron-microscopic immunocytochemistry and by cell fractionation and western blotting. Lipocortin 1 immunoreactivity is concentrated in prominent patches associated with the plasma membrane. The intensity of these patches varied with the confluence and duration of the culture and was not detectably diminished by an EDTA wash before fixation. Tubulin and cytokeratin 8 were colocalized with lipocortin 1 in the patches. Within the cells lipocortin 1 was distributed throughout the cytoplasm. Electron microscopy revealed prominent immunoreactivity along the plasma membrane with occasional large clusters of gold particles in contact with the membrane surface of the cells; within the cytoplasm the membrane of some vesicle/vacuole structures and some small electron-dense bodies was immunoreactive, but no immunogold particles were associated with the multilamellar bodies. Subcellular fractionation, extraction and western blotting showed that lipocortin 1 in the membrane pellet was present as two distinct fractions; one, intimately associated with the lipid bilayer, which behaved like an integral membrane protein and one loosely attached which behaved like a peripheral membrane protein. The results show that a substantial amounts of lipocortin 1 is concentrated in focal structures associated with and immediately beneath the plasma membrane. These might form part of the mechanism by which lipocortin 1 is released from the cells.

Polylactosaminoglycan modification of a small integral membrane glycoprotein, influenza B virus NB

1988 ◽  
Vol 8 (3) ◽  
pp. 1186-1196
Author(s):  
M A Williams ◽  
R A Lamb
Keyword(s):  
Molecular Weight ◽  
Cell Surface ◽  
Gel Filtration ◽  
Cell Types ◽  
Influenza B Virus ◽  
Influenza B ◽  
Membrane Glycoprotein ◽  
Intact Cells ◽  
B Virus ◽  
Carbohydrate Chains

The structure of the carbohydrate components of NB, the small integral membrane glycoprotein of influenza B virus, was investigated. The carbohydrate chains of NB are processed from the high-mannose form (NB18) to a heterogeneous form of much higher molecular weight, designated NBp. Selection of this carbohydrate-containing form of NB with Datura stramonium lectin, its susceptibility to digestion by endo-beta-galactosidase, and determination of the size of NBp glycopeptides by gel filtration chromatography suggested that the increase in molecular weight is due to processing to polylactosaminoglycan. Investigation of the polypeptides produced by influenza B/Lee/40 virus infection of several cell types and another strain of influenza B virus suggested that the signal for modification to polylactosaminoglycan is contained in NB. Expression of mutants of NB lacking either one or both of the normal N-terminal sites of asparagine-linked glycosylation indicated that both carbohydrate chains are modified to contain polylactosaminoglycan. NBp and a small amount of unprocessed NB18 are expressed at the infected-cell surface, as determined by digestion of the surfaces of intact cells with various endoglycosidases. Unglycosylated NB, expressed either in influenza B virus-infected cells treated with tunicamycin or in cells expressing the NB mutant lacking both N-linked glycosylation sites, was expressed at the cell surface, indicating that NB does not require carbohydrate addition for transport.

scholarly journals The Functional Study of the N-Terminal Region of Influenza B Virus Nucleoprotein

PLoS ONE ◽  
2015 ◽  
Vol 10 (9) ◽  
pp. e0137802 ◽  
Author(s):  
Ming Liu ◽  
Mandy Ka-Han Lam ◽  
Qinfen Zhang ◽  
Ruth Elderfield ◽  
Wendy S. Barclay ◽  
...  
Keyword(s):  
Terminal Region ◽  
Functional Study ◽  
Influenza B Virus ◽  
Influenza B ◽  
B Virus

scholarly journals Polylactosaminoglycan modification of a small integral membrane glycoprotein, influenza B virus NB.

1988 ◽  
Vol 8 (3) ◽  
pp. 1186-1196 ◽  
Author(s):  
M A Williams ◽  
R A Lamb
Keyword(s):  
Molecular Weight ◽  
Cell Surface ◽  
Gel Filtration ◽  
Cell Types ◽  
Influenza B Virus ◽  
Influenza B ◽  
Membrane Glycoprotein ◽  
Intact Cells ◽  
B Virus ◽  
Carbohydrate Chains

The structure of the carbohydrate components of NB, the small integral membrane glycoprotein of influenza B virus, was investigated. The carbohydrate chains of NB are processed from the high-mannose form (NB18) to a heterogeneous form of much higher molecular weight, designated NBp. Selection of this carbohydrate-containing form of NB with Datura stramonium lectin, its susceptibility to digestion by endo-beta-galactosidase, and determination of the size of NBp glycopeptides by gel filtration chromatography suggested that the increase in molecular weight is due to processing to polylactosaminoglycan. Investigation of the polypeptides produced by influenza B/Lee/40 virus infection of several cell types and another strain of influenza B virus suggested that the signal for modification to polylactosaminoglycan is contained in NB. Expression of mutants of NB lacking either one or both of the normal N-terminal sites of asparagine-linked glycosylation indicated that both carbohydrate chains are modified to contain polylactosaminoglycan. NBp and a small amount of unprocessed NB18 are expressed at the infected-cell surface, as determined by digestion of the surfaces of intact cells with various endoglycosidases. Unglycosylated NB, expressed either in influenza B virus-infected cells treated with tunicamycin or in cells expressing the NB mutant lacking both N-linked glycosylation sites, was expressed at the cell surface, indicating that NB does not require carbohydrate addition for transport.

scholarly journals Analysis of the desialidation process of the haemagglutinin protein of influenza B virus: the host-dependent desialidation step

2002 ◽  
Vol 83 (7) ◽  
pp. 1729-1734 ◽  
Author(s):  
C. Luo ◽  
E. Nobusawa ◽  
K. Nakajima
Keyword(s):  
Cell Surface ◽  
Mdck Cells ◽  
Influenza B Virus ◽  
Influenza B ◽  
Dependent Manner ◽  
Cos Cells ◽  
Trans Golgi Network ◽  
B Virus ◽  
Golgi Network ◽  
Ha Protein

It was reported previously that haemadsorption by the haemagglutinin (HA) protein of influenza B virus required that the protein must undergo desialidation. When MDCK and COS cells were infected with influenza B/Kanagawa/73 virus in the presence of a neuraminidase (NA) inhibitor, Zanamivir, haemadsorption on MDCK cells was inhibited but that on COS cells was not. The activity of the NA protein of the two types of infected cells was similar and both were inhibited by Zanamivir in a dose-dependent manner. A comparison of the desialidation of the HA protein was made on MDCK and COS cells in the presence of bacterial NA and both cells were found to have similar sensitivity. On the accumulation of the HA and NA proteins in the trans-Golgi network of MDCK cells by means of low-temperature treatment, desialidation of the HA protein in the presence of Zanamivir was detected by two-dimensional gel electrophoresis. Because this agent was reported to be unable to penetrate cells, these data suggest that, in MDCK cells, desialidation of the HA protein occurs on the cell surface but, in COS cells, the HA and NA proteins might accumulate in the trans-Golgi network, thus allowing NA desialidation before their migration to the cell surface.

scholarly journals Recurring Influenza B Virus Infections in Seals

2013 ◽  
Vol 19 (3) ◽  
pp. 511-512 ◽  
Author(s):  
Rogier Bodewes ◽  
Danny Morick ◽  
Gerrie de Mutsert ◽  
Nynke Osinga ◽  
Theo Bestebroer ◽  
...  
Keyword(s):  
Influenza B Virus ◽  
Influenza B ◽  
Virus Infections ◽  
B Virus
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