FIG. 3.

Glycosphingolipid synthesis inhibitors block BAAV transduction and cell entry, but not cell attachment. (A) Transduction. COS cells were pretreated with PPMP or PDMP before transduction with rAAV5-GFP and rBAAV-GFP, and transduced cells were counted 24 h after vector addition by flow cytometry. (B) Binding. COS cells were pretreated with 5 μM PPMP, chilled, and incubated with rAAV5-GFP and rBAAV-GFP at 4°C. After the cells were washed, bound virus was measured by quantitative PCR (QPCR). (C) Cell entry. COS cells were pretreated with 5 μM PPMP prior to transduction with rAAV5-GFP and rBAAV-GFP at 37°C. Four hours after transduction, cell-associated virus was removed by a combination of wash steps, neuraminidase treatment, and proteolytic digestion of intact cells, and internalized vector was quantitated by QPCR. The values are means from three experiments; the error bars represent standard deviations.