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. 2011 Oct;85(19):9834–9846. doi: 10.1128/JVI.02633-10

Fig. 11.

Fig. 11.

The YxY sequence in the cytoplasmic domain of BST-2 facilitates but is not essential for endocytosis. (A) Virion release. Cells (HEK 293T in wells of a 6-well plate) were transfected to express either wild-type (WT) BST-2 (75 ng of plasmid) or BST-2 in which tyrosines 6 and 8 in the cytoplasmic domain were replaced with alanines (Y6,8A) along with either wild-type or vpu-negative (ΔVpu) proviral plasmids (3.75 μg). The next day, the culture supernatants were removed and clarified by centrifugation at 300 × g. Virions were purified by centrifugation through a 20% sucrose cushion at 23,500 × g, suspended in the original supernatant volume, then analyzed in duplicate by p24 capsid antigen ELISA. (B) Constitutive endocytosis rate. Cells (HEK 293T in wells of a 12-well plate) were transfected to express either wild-type BST-2 (60 ng of plasmid) or BST-2 in which the tyrosines 6 and 8 in the cytoplasmic domain were replaced with alanines (Y6,8A), along with GFP (0.1 μg of plasmid) as a transfection marker. The next day, the fractional rate of endocytosis of BST-2 was measured, as shown in Fig. 6 and 8. Data are normalized to 100% at time zero in each case.