Monitoring HIV-1 Protein Oligomerization by FLIM FRET Microscopy

  • Ludovic Richert
  • Pascal Didier
  • Hugues de Rocquigny
  • Yves MélyEmail author
Part of the Springer Series in Chemical Physics book series (CHEMICAL, volume 111)


The majority of the human immunodeficiency virus type 1 (HIV-1) proteins are able to self assemble into oligomers. Since these oligomers generally exhibit functions that differ from those of their monomeric counterpart, the regulation of the monomer-oligomer equilibria plays a central role in the viral cycle. To characterize the oligomerization of these proteins in live cells, the combination of fluorescence lifetime imaging microscopy (FLIM) with Förster resonance energy transfer (FRET) has proven to be very powerful. In this review, we illustrate the application of FRET-FLIM on the characterization of the oligomerization of the Vpr, Vif and Pr55Gag proteins of HIV-1 in fusion with eGFP and mCherry. For Vpr and Pr55Gag proteins, very high levels of FRET leading to strong decreases in eGFP fluorescence lifetime are obtained, as a consequence of the rather small size of the viral proteins, the strong packing of the protomers and the presence of multiple acceptors for one donor. Analyzing the time-resolved decays by a two-component analysis further provides the possibility to discriminate monomers from oligomers and to monitor the spatiotemporal evolution of both populations in the cells. Though FRET-FLIM unambiguously reveals the oligomerization of a given protein, it hardly discloses the oligomer stoichiometry (number of protomers per oligomers). This parameter can be obtained by fluorescence correlation spectroscopy, which allows further interpreting the FRET-FLIM data. FRET-FLIM is also highly useful to identify the determinants of the oligomerization process and to investigate its regulation by other HIV-1 proteins and host proteins.


Fluorescence Lifetime Fluorescence Correlation Spectroscopy Fluorescence Lifetime Imaging Microscopy Fluorescence Correlation Spectroscopy Measurement Fast Decay Component 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



We thank Salah Edin El Meshri for his technical help. This work was supported by the European Project THINPAD “Targeting the HIV-1 Nucleocapsid Protein to fight Antiretroviral Drug Resistance” (FP7—grant agreement 601969), Agence National Recherche sur le SIDA (ANRS) (2012-14. CSS2), SIDACTION (AI22-1-01963), Centre National Recherche Scientifique (CNRS), Université de Strasbourg and Institut National sur la Santé Et la Recherche Médicale (INSERM).


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Copyright information

© Springer International Publishing Switzerland 2015

Authors and Affiliations

  • Ludovic Richert
    • 1
  • Pascal Didier
    • 1
  • Hugues de Rocquigny
    • 1
  • Yves Mély
    • 1
    Email author
  1. 1.UMR 7213 CNRS, Laboratoire de Biophotonique et PharmacologieFaculté de PharmacieIllkirchFrance

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