Nature 454, 236-240 (10 July 2008) | doi:10.1038/nature06998; Received 13 March 2008; Accepted 10 April 2008; Published online 25 May 2008

Imaging the biogenesis of individual HIV-1 virions in live cells

Nolwenn Jouvenet1,2, Paul D. Bieniasz1,2 & Sanford M. Simon3

  1. Aaron Diamond AIDS Research Center,
  2. Laboratory of Retrovirology, and,
  3. Laboratory of Cellular Biophysics, The Rockefeller University, New York, New York 10065, USA

Correspondence to: Paul D. Bieniasz1,2Sanford M. Simon3 Correspondence and requests for materials should be addressed to P.D.B. (Email: pbienias@adarc.org) or S.M.S. (Email: simon@rockefeller.edu).

Observations of individual virions in live cells have led to the characterization of their attachment, entry and intracellular transport1. However, the assembly of individual virions has never been observed in real time. Insights into this process have come primarily from biochemical analyses of populations of virions or from microscopic studies of fixed infected cells. Thus, some assembly properties, such as kinetics and location, are either unknown or controversial2, 3, 4, 5. Here we describe quantitatively the genesis of individual virions in real time, from initiation of assembly to budding and release. We studied fluorescently tagged derivatives of Gag, the major structural component of HIV-1—which is sufficient to drive the assembly of virus-like particles6—with the use of fluorescence resonance energy transfer, fluorescence recovery after photobleaching and total-internal-reflection fluorescent microscopy in living cells. Virions appeared individually at the plasma membrane, their assembly rate accelerated as Gag protein accumulated in cells, and typically 5–6 min was required to complete the assembly of a single virion. These approaches allow a previously unobserved view of the genesis of individual virions and the determination of parameters of viral assembly that are inaccessible with conventional techniques.