Integration of a virus membrane protein intothe lipid bilayer of target cells as a prerequisite for immune cytolysis. Specific cytolysis after virosome- target cell fusion

Abstract

Structural requirements for membrane antigens on target cells to mediate immune cytolysis were studied in a model system with purified membrane proteins from Semliki Forest virus (SFV). These SFV spike proteins were isolated in the form of detergent- and lipid-free protein micelles (29S complexes) or, after reconstitution into lipid vesicles, in the form of virosomes. Both the 29S complexes and the virosomes were found to bind well to murine tumor cells (P815 or Eb). When these cells, however, were used as target cells in complement-dependent lysis or in antibody-dependent cell- mediated cytotoxicity assays in the presence of anti-SFV serum, they were not lysed, although they effectively bound the antibody and consumed complement. The same tumor cells infected with SFV served as positive controls in both assays. Different results were obtained when inactivated Sendai virus was added as a fusion reagent to the cells coated with either virosomes or 29S complexes. Under these conditions the virosome-coated cells became susceptible to SFV- specific lysis, whereas the 29S complex-coated cells remained resistant. Evidence that the susceptibility to lysis ofvirosome-coated cells was dependent on active fusion and, therefore, integration of the viral antigens into the lipid bilayer of the target cells was derived from control experiments with enzyme-treated Sendai virus preparations. The 29S complexes and the virosomes partially and selectively blocked the target cell lysis by anti-H-2 sera but not by anti-non-H-2 sera confirming our previous finding that major histocompatibility antigens serve as receptors for SFV. The general significance of these findings for mechanisms of immune cytolysis is dicussed.

Antibody Response to Spike Protein Vaccines Prepared from Semliki Forest Virus

“…The cytolysis of target cells was assayed in a 51Cr release test performed in round-bottom microtitre plates (Linbro, IS-MRC 96) (Morein et al, 1979). As target cells, the tumour cell line P815 (a mastocytoma cell) derived from DBA/2 mice was used.…”

Purification of Measles Virus Glycoproteins and their Integration into Artificial Lipid Membranes

“…A number of such studies using virosomes, especially relating to the generation of cytotoxic T cells, have been reported with Sendai virus (Finberg et al, 1978), influenza virus (Koszinowski et al, 1980), SFV (Morein et al, 1979) and vesicular stomatitis virus (Loh et al, 1979) in murine systems. However, the role of measles virus in human disease, the known immunosuppressive effects of the whole virus on lymphocytes (Joseph et al, 1975) and the ability of measles virus to replicate in B lymphocytes, and various T lymphocyte subsets (Joseph et al, 1975;Huddlestone et al, 1979Huddlestone et al, , 1980, will make such analyses in the human situation of particular interest.…”

Recognition of viral glycoproteins by influenza A-specific cross-reactive cytolytic T lymphocytes.

“…Second, the HA and N glycoprotein of the human influenza virus A/Victoria were separated from the other virion components and reconstituted into artificial lipid bilayers (24,25) to give vesicles free of M protein and NP. Target cells were then prepared by incubating these vesicles with cells in the presence of fusion-active Sendai virus (26). Various influenza A-specific CTL were tested against both types of target cells.…”