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. 2004 Dec 6;167(5):903–913. doi: 10.1083/jcb.200407031

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Copyright © 2004, The Rockefeller University Press

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Figure 2. — HIV-1 Nef targets MHC-I into an AP-1–dependent pathway. (A and B) Western blot analysis of adaptin and Nef expression in siRNA-treated cells. Astrocytic cells (A) or CEM T cells (B) were transfected with the indicated siRNA and transduced with control or Nef-expressing adenoviruses as described in Materials and methods. Lysates were harvested and immunoblotted for the indicated protein. (C and D) Flow cytometric analysis of siRNA-treated cells. Astrocytic cells (C) or CEM T cells (D) were stained for surface HLA-A2 expression (left column) or surface CD4 expression (right column); shaded curve, control adenovirus; black line, Nef-expressing adenovirus. (E) Depletion of AP-1 inhibits Nef-dependent HLA-A2 degradation. CEM T cells were transduced and treated with siRNAs as described above except that an additional siRNA transfection was included before transduction. Lysates were prepared and analyzed by immunoblot to examine the expression levels of the indicated proteins.