Document 0192
 DOCN  M9440192
 TI    Photoinactivation and kinetics of membrane fusion mediated by the human
       immunodeficiency virus type 1 envelope glycoprotein.
 DT    9404
 AU    Dimitrov DS; Blumenthal R; Section on Membrane Structure and Function,
       National Cancer; Institute, NIH, Bethesda, MD 20892.
 SO    J Virol. 1994 Mar;68(3):1956-61. Unique Identifier : AIDSLINE
       MED/94149891
 AB    The fusion kinetics of cells expressing the human immunodeficiency virus
       type 1 (HIV-1) envelope glycoprotein with CD4 target cells was
       continuously monitored by image-enhanced Nomarski differential
       interference contrast optics. The analysis of the videotape recordings
       showed that (i) cells made contact relatively rapidly (within minutes),
       in many cases by using microspikes to touch and adhere to adjoining
       cells; (ii) the adhered cells fused after a relatively long waiting
       period, which varied from 15 min to hours; (iii) the morphological
       changes after membrane fusion, which led to disappearance of the
       interface separating the two cells, were rapid (less than 1 min); and
       (iv) the process of syncytium formation involved subsequent fusion with
       other cells and not simultaneous fusion of many cells. To measure the
       kinetics of early stages of cell fusion, we used the recently developed
       very stable membrane-soluble dye, PKH26, which redistributes between
       labeled and unlabeled membranes after fusion but does not exchange
       spontaneously between membranes for prolonged periods. We found that
       photoactivation of this dye by illumination with green light inhibits
       fusion of cell membranes as indicated by the lack of dye transfer from
       the labeled HIV-1 envelope-expressing cells to unlabeled CD4 cells. The
       inhibitory effect was localized in space and time, which allowed us to
       develop a new assay for measuring the kinetics of membrane fusion by
       illuminating the cell mixture at different times after coculture. This
       assay has also been used to monitor the fusion kinetics of HIV-1 and
       recombinant vaccinia virus. The photoactivation of nonexchangeable
       membrane-soluble fluorescent dyes may be useful for development of new
       assays for measuring the kinetics of membrane fusion and could also be
       important in designing new antiviral approaches.
 DE    Clone Cells  Fluorescent Dyes/*PHARMACOLOGY  HIV Envelope Protein
       gp120/*RADIATION EFFECTS  HIV Envelope Protein gp41/*RADIATION EFFECTS
       HIV-1/*RADIATION EFFECTS  Kinetics  Light  Membrane Fusion/*RADIATION
       EFFECTS  Microscopy  Photosensitizing Agents/*PHARMACOLOGY  Support,
       U.S. Gov't, P.H.S.  Vaccinia Virus/RADIATION EFFECTS  Video Recording
       JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).