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. Author manuscript; available in PMC: 2017 Sep 2.
Published in final edited form as: Virus Res. 2016 Jul 25;223:161–169. doi: 10.1016/j.virusres.2016.07.009

Figure 5.

Figure 5

Ubiquitylation of viral and cellular proteins. (A) Ubiquitylation of USP15 and Nef. Ub-expressing plasmid (pUb) was transfected with USP15- (Top) or with nef-expressing plasmid (Bottom), and ubiquitylation of these proteins was determined by immunoprecipitation with anti-HA Ab for Ub followed by Western blot analysis with anti-USP15 (Top) or anti-Myc antibody for Nef (lane 2). Lane 1 indicates WB analysis of USP15 (Top) and Nef (Bottom). Arabic numbers on the left indicate positions of pre-stained protein size marker (kDa) (Fisher Scientific). (B and C) Ubiquitylation of cellular proteins. Total cell lysates generated from 293T cells transfected with pUb and 1 or 4 μg of USP15- or Nef-expressors were analyzed by WB with anti-HA Ab for ubiquitylated cellular proteins, followed with anti-Myc Ab for USP15 and Nef, and anti-β-actin Ab for β-actin protein, and (C) the relative amount of ubiquitylated proteins was determined by scanning the entire area of each lane, using ChemiDoc Touch Imaging System (Bio-Rad, Hercules, CA) and depicted as mean +/− SD of triplicates, shown in (B). The data were representative of three independent experiments.