title: Probing HIV-1 membrane liquid order by Laurdan staining reveals producer cell-dependent differences.
abstract: Viruses acquire their envelope by budding from a host cell membrane, but viral lipid composition may differ from that of the budding membrane. We have previously reported that the HIV-1 membrane is highly enriched in cholesterol, sphingolipids, and other raft lipids, suggesting that the virus may bud from pre-existing or virus-induced lipid rafts. Here, we employed the environmentally sensitive fluorescent dye Laurdan to study the membrane lateral structure of HIV-1 derived from different cell lines. Differences in viral membrane order detected by Laurdan staining were shown by mass spectrometry to be due to differences in lipid composition. Isogenic viruses from two different cell lines were both strongly enriched in raft lipids and displayed a liquid-ordered membrane, but these effects were significantly more pronounced for HIV-1 from the T-cell line MT-4 compared with virus from 293T cells. Host-dependent differences in the lipidomes predominantly affected the ratio of sphingomyelins (including dihydrosphingomyelin) to phosphatidylcholine, whereas cholesterol contents were similar. Accordingly, treatment of infectious HIV-1 with the sphingomyelin-binding toxins Equinatoxin-II or lysenin showed differential inhibition of infectivity. Liposomes consisting of lipids that had been extracted from viral particles exhibited slightly less liquid order than the respective viral membranes, which is likely to be due to absence of membrane proteins and to loss of lipid asymmetry. Synthetic liposomes consisting of a quaternary lipid mixture emulating the viral lipids showed a liquid order similar to liposomes derived from virion lipids. Thus, Laurdan staining represents a rapid and quantitative method to probe viral membrane liquid order and may prove useful in the search for lipid active drugs.
authors: Department of Virology, Universitätsklinikum Heidelberg, Heidelberg D-69120, Germany.
authors: Maier
authors: M
authors: Lorizate
authors: Heidelberg University Biochemistry Center, Heidelberg D-69120, Germany.
authors: Britta
authors: B
authors: Brügger
authors: Department of Virology, Universitätsklinikum Heidelberg, Heidelberg D-69120, Germany.
authors: Hisashi
authors: H
authors: Akiyama
authors: Department of Virology, Universitätsklinikum Heidelberg, Heidelberg D-69120, Germany.
authors: Bärbel
authors: B
authors: Glass
authors: Department of Virology, Universitätsklinikum Heidelberg, Heidelberg D-69120, Germany.
authors: Barbara
authors: B
authors: Müller
authors: Department of Biology, Biotechnical Faculty, University of Ljubljana, Večna pot 111, Ljubljana 1000, Slovenia.
authors: Gregor
authors: G
authors: Anderluh
authors: Heidelberg University Biochemistry Center, Heidelberg D-69120, Germany.
authors: Felix T
authors: FT
authors: Wieland
authors: Department of Virology, Universitätsklinikum Heidelberg, Heidelberg D-69120, Germany.
authors: Hans-Georg
authors: HG
authors: Kräusslich
mesh_terms: D015081:2-Naphthylamine / Q000031:analogs & derivatives / Q000494:pharmacology; D002460:Cell Line; D003064:Cnidarian Venoms / Q000737:chemistry; D015658:HIV Infections / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D007848:Laurates / Q000494:pharmacology; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D021962:Membrane Microdomains / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000379:methods; D013696:Temperature; D014118:Toxins, Biological / Q000737:chemistry
chemical_list: D003064:Cnidarian Venoms; D007848:Laurates; D008055:Lipids; D008081:Liposomes; D014118:Toxins, Biological; C107040:lysenin; C012307:equinatoxin; D015081:2-Naphthylamine; C065580:laurdan