Localization of PI ( 4 , 5)P
and phosphatidylinositol 3-phosphate in MDMs. MDMs were nucleofected to express PH-GFP, to localize (A,B) PI(4,5)P2 or (C) a double-FYVE domain construct fused to GFP (2xFYVE-GFP), a probe for phosphatidylinositol 3-phosphate. After 24 hours the cells were fixed and co-stained for CD81 or CD44 as indicated. (D) Seven-day-old MDMs were infected with HIV-1BaL and after a further 7 days were nucleofected to express PH-GFP, incubated for 24 hours, then fixed and stained for the viral matrix protein p17. Images show single confocal sections, and were acquired with a Leica SPE confocal microscope as in Figure 1. All experiments were repeated on cells from at least three different donors. Scale bars = 10 μm. PI(4,5)P2: Phosphatidylinositol 4,5-bisphosphate; PH-GFP: Phospholipase Cδ pleckstrin homology domain linked to GFP.