FIG. 1.
The G protein partitions into membrane microdomains independently of other viral components. (A) Comparison of G protein cell surface expression in VSV-infected cells to that in cells transfected with pCAGGS-G DNA. BHK cells were infected with wt VSV for 4.5 h or were transfected with pCAGGS-G DNA for 12, 24, or 36 h. Following infection or transfection, cells were labeled for immunofluorescence with an anti-G protein antibody and a secondary antibody conjugated to fluorescein. G protein cell surface fluorescence was quantitated by flow cytometry. The data are presented as the mean levels of fluorescence of the G protein-positive population ± SD for three experiments. (B) Representative micrograph of immunogold labeling of the G protein in transfected cells. a to c, groups of gold particles that might represent G protein-containing microdomains. Bar = 100 nm. (C) Analysis of the G protein distribution in the plasma membranes of cells transfected with pCAGGS-G. A series of 50 electron micrographs of arbitrarily chosen areas of plasma membrane similar to those shown in panel B were digitized and analyzed by tracing the plasma membrane from the left side of the image to the right while registering each gold particle that was encountered in the trace. The data were converted to pairwise distance measurements between all the gold particles in a micrograph and were plotted as a histogram with the number of pairs of gold particles on the y axis and the distance between gold particles on the x axis displayed as successive 20-nm windows. The y values were normalized by dividing by the number of gold particles analyzed to give average densities. Horizontal dashed line, average concentration of the G protein in the plasma membrane; solid black line through the data points, best-fit line for the data; arrow, point at which slope changes, which gives the diameter of the microdomains.