Filovirus vectors and noninfectious filovirus-based particles

Abstract

Cloned filovirus genomic cDNA and methods of using the cDNA are provided. Further provided are noninfectious lipid encapsulated filovirus-based particles.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is a divisional of U.S. patent application Ser. No. 10 / 353,856, filed Jan. 29, 2003, which claims the benefit of the filing date of U.S. application Ser. No. 60 / 353,972, filed on Jan. 31, 2002, under 35 U.S.C. §119(e). Which applications are incorporated by reference herein.STATEMENT OF GOVERNMENT RIGHTS[0002]This invention was made with government support awarded by the National Institutes of Health, Grant Nos. AI42774 and AI44386. The Government has certain rights in this invention.BACKGROUND OF THE INVENTION[0003]Ebola virus, a member of the family Filoviridae and the order Mononegavirales, is an enveloped, nonsegmented negative-strand RNA virus and is one of the most lethal human and nonhuman primate pathogens recognized to date (Feldmann et al., 1998; Vanderzanden et al., 1998). Four subtypes of Ebola virus have been identified, including Zaire, Sudan, Ivory Coast, and Reston (Sanchez et al., 1993). Human infection w...

AI Technical Summary

Benefits of technology

[0014]As also described herein, VP40, when expressed apart from other viral proteins in mammalian cells, induced particle formation, which differed in length but with uniform diameters of approximately 65 nm. Efficient particle formation may rely on a conserved N-terminal PPXY motif, as mutation or loss of this motif resulted in markedly reduced particle formation. These findings demonstrate that VP40 alone possesses the information necessary to induce particle formation, and this process most likely requires cellular WW-domain-containing proteins that interact with the PPXY motif of VP40. Flotation gradient analysis indicated that VP40 binds to membranes in a hydrophobic manner, as NaCl at 1 M did not release the protein from the lipid bilayer. Triton X-114 phase-partitioning analysis suggested that VP40 possesses only minor features of an integral membrane protein. Truncation of the C-terminal 50 amino acids of VP40 resulted in decreased association with cellular membranes, and demonstrated that this deletion disrupts hydrophobic interactions of VP40 with the lipid bilayer, as well as abolishing particle formation. Truncation of the C-terminal 150 amino acids or N-terminal 100 amino acids of VP40 enhanced the protein's hydrophobic association with cellular membranes. These mutants may be useful as dominant negatives, to determine targets for antivirals.

Problems solved by technology

However, identification of major determinants of Ebola virus pathogenicity has been hampered by the lack of effective strategies for experimental mutagenesis.

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