Bcl-2 and Bax interactions in mitochondria probed with green fluorescent protein and fluorescence resonance energy transfer

Nat Biotechnol. 1998 Jun;16(6):547-52. doi: 10.1038/nbt0698-547.

Abstract

It has been hypothesized that interaction of Bcl-2 and Bax may regulate apoptosis. The spatial and temporal interaction of Bcl-2 and Bax at the single cell level has not, however, been demonstrated. To achieve this goal, we have developed two-fusion FRET (fluorescence resonance energy transfer). Using green fluorescent protein (GFP)-Bax and blue fluorescent protein (BFP)-Bcl-2 fusion proteins coexpressed in the same cell, we demonstrate a direct interaction between Bcl-2 and Bax in individual mitochondria. Mitochondrially localized cytochrome c-GFP and BFP-Bcl-2 showed little or no FRET, while nuclear-localized GFP-human papillomavirus E6 and BFP-Bcl-2 did not interact when coexpressed in the same cell. These findings indicate that two-fusion FRET provides an opportunity to examine the interaction between two different proteins coexpressed in single intact mammalian cells.

MeSH terms

  • 3T3 Cells
  • Animals
  • Fluorescent Dyes / metabolism
  • Green Fluorescent Proteins
  • Indicators and Reagents
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism*
  • Mice
  • Microscopy, Fluorescence
  • Mitochondria / genetics
  • Mitochondria / metabolism*
  • Molecular Probes / metabolism
  • Proto-Oncogene Proteins / genetics
  • Proto-Oncogene Proteins / metabolism*
  • Proto-Oncogene Proteins c-bcl-2 / genetics
  • Proto-Oncogene Proteins c-bcl-2 / metabolism*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Spectrometry, Fluorescence
  • bcl-2-Associated X Protein

Substances

  • Bax protein, mouse
  • Fluorescent Dyes
  • Indicators and Reagents
  • Luminescent Proteins
  • Molecular Probes
  • Proto-Oncogene Proteins
  • Proto-Oncogene Proteins c-bcl-2
  • Recombinant Fusion Proteins
  • bcl-2-Associated X Protein
  • Green Fluorescent Proteins