Disassembly of viral membranes by complement independent of channel formation.

Abstract

We have compared the effects of the complement membrane attack complex (MAC), nystatin, and melittin on the envelope of murine leukemia viruses to determine if channel formation alone is sufficient to cause membranolysis. Nystatin is a channel former and mellitin is not, although both are hemolytic. Whereas MAC and melittin disintegrated the viral membrane, nystatin had no effect on mo hology, integ- (2), who measured the release of radiolabeled RNA from mouse leukemia viruses (retroviruses). Later experiments with retroviruses indicated that human complement alone, without specific antibody, could lyse retroviruses (3-5). The mechanism of lysis of enveloped viruses was assumed to be similar or identical to complement-mediated lysis of mammalian cells, a more extensively studied and better understood reaction.In complement-dependent cytolysis, five complement proteins-C5b, C6, C7, C8, and C9-become physically attached to the outer membrane of a cell. Without further enzymatic modification, these proteins form a multimolecular complex, the membrane attack complex (MAC), that lyses cells by purely physicochemical means (reviewed in ref. 6). Loss of the natural impermeability of the cell membrane to small solutes leads to colloid-osmotic swelling of the cell, rupture, and release of internal proteins. Mayer (7,8) theorized that the MAC forms hydrophilic protein channels; this action is reminiscent of the channel-forming (9) and hemolytic (10) properties of the antibiotics nystatin and amphotericin B.In contrast to Mayer's channel theory, we recently proposed (11, 12) that the MAC is membranolytic not because of channel formation, but because of its strong phospholipid binding capacity, through which it is enabled to reorient the lipid bilayer structure of membranes.Examining our hypothesis, we have compared the effects of nystatin, melittin, and the MAC on the envelope of murine leukemia virus (MuLV) to determine if channel formation alone is sufficient to cause membranolysis. Nystatin is a channel former and melittin is not, although it is membranolytic. Our results show that whereas the MAC and melittin are virolytic, nystatin has no effect on the morphology and infectivity of the virus.MATERIALS AND METHODS Materials. Nystatin, amphotericin B, and melittin were purchased from Sigma. Rauscher MuLV, prepared by Electro-Nucleonics (Bethesda, MD), was supplied by the Office of Program Resources and Logistics, Viral Oncology, National Cancer Institute. Freshly harvested Moloney, xenotropic (ATS-124) murine, and (BN-p 454-9) rat leukemia viruses were propagated as described (6). Fresh human serum (Community Blood and Plasma, San Diego, CA) frozen at -70'C was used as a source of complement.Uranyl formate was prepared according to published procedures (13); all other chemicals were of the best grade commercially available. Buffer osmolarities were measured with a freezing-point depression osmometer (Osmette, Precision Systems, Sudbury, MA).Virolysis Assays. Virus samples (100,ug or approximately 1010 virions ...

Proteolytic modification of human complement protein C9: loss of poly(C9) and circular lesion formation without impairment of function.

“…This hypothesis is supported by studies by Esser and co-workers, (9)(10)(11), who investigated the effect of the terminal proteins on lipid organization in the target membrane and the mechanism of virolysis by complement. In addition, Lauf (12) has reported that the functional characteristics of immune hemolysis are inconsistent with a 10-nm cylindrical pore as seen in the electron microscope, a view that is reinforced by the later data of Sims and Lauf (13)(14)(15).…”

Inhibitory Activity of Synthetic Peptide Antibiotics on Feline Immunodeficiency Virus Infectivity In Vitro

“…The membrane-targeted activity of peptide antibiotics was demonstrated in voltage-dependent channel formation in artificial membranes (1,18), lysis of liposomes, and direct lysis of enveloped virus (2,8,11,21,27,43). Despite general membrane disruption phenomena, the peptide antibiotics generally have strong selectivity against microbes but not normal host cells.…”

Late-Acting Components of Complement: Their Molecular Biochemistry, Role in the Host Defense, and Involvement in Pathology

“…Spin-label studies with ordered membranes indicated that upon C5b-8 and C5b-9 inser tion a reorientation of lipid occurs suggest ing a transition from a bilayer to a micellar packing [66], Also it has been reported that when the lytic complex embeds into mem branes, phospholipid vesicles are released [68]. The net effect can account for comple ment disassembly of viral envelope mem branes, as channel formation does not ap pear to be a virolytic mechanism [69]. Since cells arc much larger than virus and have a more involuted plastic membrane, this mechanism may not be of major importance for cytolysis.…”

“…While complement has been found to kill tumor and viral-infected cells, neutralize virus and destroy protozoan parasites [69,[77][78][79][80], the main benefit in the host defense of the com plement-terminal components is probably to kill gram-negative bacteria and mycoplasmas [81], The reason is that nucleated cells can vesiculate and shed cytolytic complexes. Consequently, lysis is inefficient and is char acterized by multiple-hit kinetics [82][83][84][85], By contrast the bactericidal action of serum is well established [82], and even serum di luted severalfold and depleted of lysozyme can kill effectively many strains of microor ganisms [81].…”