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. 2014 May 2;9(5):e96092. doi: 10.1371/journal.pone.0096092

Figure 4. Increased levels of cytoplasmic CAR by proteasomal inhibition correlates with an increase in HSP70 levels.

Figure 4

Results shown are representative of three independent experiments. (A) HepG2 cells were transfected with V5-tagged mouse CAR (pcDNA3.1/V5-His-mCAR, 3 µg) and mouse CCRP (pcDNA3.1/V5-His-mCCRP, 0.3 µg), and 18 hours later transfected cells were treated for 24 hr with DMSO (0.1% v/v), TCPOBOP (250 nM in 0.1% DMSO, final concentrations), MG132 (5 µM in 0.1% DMSO, final concentrations), or TCPOBOP combined with MG132. Cells were then harvested and cytosolic extracts prepared and subjected to immunoblotting analysis with antibodies against the indicated proteins. (B) Extracts analyzed in (A) were subjected to immunoprecipitation analysis using anti-V5 or anti-IgG (as control) antibodies was performed, followed by immunoblotting analysis using anti-V5 (top panel), anti-HSP70 (middle panel), and anti-HSP90 (bottom panel) antibodies.