Fine structure of influenza A virus observed by electron cryo-microscopy.

Abstract

A rapidly frozen vitrified aqueous suspension of influenza A virus was observed by high resolution electron cryomicroscopy. The influenza particles were grouped into small (diameter < 150 nm) spherical particles with well organized interiors, large spherical ones with less internal organization, and framentous ones. Envelopes of most of the large virus particles were phospholipid bilayers, and the chromatography fraction containing these large particles was largely devoid of viral activity. The envelopes of most of the fiamentous and small spherical virus particles, on the other hand, gave a strange contrast which could be ascribed to a combination of a thin outer lipid monolayer and a 7.2 nm thick protein-containing inner layer. These latter particles represented most of the viral activity in the preparation. Densitometric traces of the near in-focus images confimed these structural differences. Some viral envelope structures apparently intermediate between these two distinct types of membrane were also detected. A structural model of intact biologically active influenza virus particles was formulated from these results, together with computer simulations.

The M1 and NP proteins of influenza A virus form homo- but not heterooligomeric complexes when coexpressed in BHK-21 cells.

“…Membranes were isolated from transfected cells by floatation in a sucrose gradient as described above, incubated with (i) 2 M KCl\10 mM Tris-HCl (pH 7n4) for 1 h at 25 mC or (ii) 100 mM Na # CO $ for 30 min at 0 mC (Chong & Rose, 1993 ;Fujiyoshi et al, 1994). Treated membranes were adjusted to 80 % (w\v) sucrose, and subjected to a new membrane flotation centrifugation.…”

“…This implies that they have a central function in the assembly process of the virus. In the virion the M1 proteins form a continuous layer at the internal surface of the envelope thereby enclosing the eight viral ribonucleoproteins (RNPs) (Murti et al, 1992 ;Fujiyoshi et al, 1994). The NPs, on the other hand, form the backbone of the RNPs (Pons et al, 1969 ;Kingsbury et al, 1987 ;Ruigrok & Baudin, 1995).…”

Formation of virus-like particles from human cell lines exclusively expressing influenza neuraminidase

“…Influenza virions are pleomorphic, although generally their shape is roughly spherical with a diameter of ,150 nm. However, larger (100-400 nm) influenza virions are also generated, as are filamentous forms (Fujiyoshi et al, 1994).…”

Structural organization of a filamentous influenza A virus

“…In addition, ultrastructural changes that are essential to membrane fusion and virion disassembly during cell entry remain to be described. Electron cryomicroscopy of influenza virions that are vitrified by rapid plungefreezing (14)(15)(16)(17) shows contrast from virion structures themselves and not from stain distributions, which can also distort or disrupt virus structure. However, an understanding of the structural organization of influenza virus has remained difficult because of its pleomorphic nature.…”