FIG. 8.

Gag-Tsg101 interactions are detected by FRET. (A) Tsg101-YFP was expressed in Mel JuSo cells and imaged by confocal microscopy; the image shows YFP excitation-emission. (B) Tsg101-YFP coexpressed with Gag-CFP; the image shows YFP excitation-emission. (C) Gag-CFP and Tsg101-YFP were expressed in 293T cells, and the membrane and cytosolic fractions were isolated and analyzed by scanning cuvette fluorometry. Gag-CFP-Tsg101-YFP coexpression resulted in a FRET curve for the membrane fraction but not for the cytosolic fraction (closed and open diamonds, respectively). Tsg101-YFP alone displayed no FRET curve (closed and open squares). (D) Similar expression levels and membrane binding percentages were observed for Tsg101-YFP alone and in the presence of Gag-CFP from the experiment shown in panel C.