The GFP-PLCδ1 PH domain accumulates on drug-induced endovesicles. (A-D) Fluorescence micrographs (60×/1.42 oil objective [Olympus]) of erythrocyte ghosts loaded with GFP- PLCδ1 PH domain (panel B; green) and treated with primaquine in the presence of 70-kDa rhodamine dextran (panel C; red). Rhodamine dextran accumulated inside endovesicles, and the GFP-PLCδ1 PH domain colocalized to the same region as rhodamine dextran. Merge shown in panels A (bar equals 10 μm) and D. Dashed box in panel A denotes cell magnified in panels B through D. Scale bar equals 3 μm (panels B-D). (E) Model of erythrocyte lipid uptake and exclusion in drug-induced vesicles (DIVs). Based on mass spectrometry and immunofluorescence results, DIVs appear to undergo complex but ordered lipid sorting during their primaquine-induced formation. In particular, whereas PE and PG are absent from isolated endovesicles, PS and especially PI phospholipids traffic to the endovesicle membranes. With respect to PIP₂, this is quantitatively apparent by mass spectrometry and fluorescence microscopy. DIVs may consist of smaller, dynamic raft microdomains containing PIP₂ and PS. The extent to which the ratio of cytoskeletal proteins such as spectrin and actin are changed in DIV as well as the presence of junctional complex proteins like 4.1 in DIV have not yet been established.