Figure 4.

USP48 downregulation stabilizes p65, mainly in the nuclear compartment. USP48 expression was downregulated in hTERT-RPE1 cells at both basal conditions (NT) and after treatment with TNFα at several different time points. Cells were harvested and processed for immunoblotting. (A) Densitometric quantification indicates that p65 levels significantly increased in NT cells. (B) No changes were detected in IκBα expression levels. (C) Nuclear and cytosolic extracts were prepared for immunoblotting with anti-USP48 and p65 antibodies. Anti-nucleolin (nuclear) and anti-tubulin (cytosolic) were used to detect both enrichment and contamination of fractionated cell samples. p65 levels increased specifically in the nuclear compartment when USP48 is downregulated. Images represent one of three (A,C) or four (B) independent experiments. Statistical analysis by Kruskal Wallis in (A,B): * p-value ≥ 0.05 and by Student’s t-test in (C): * p-value ≥ 0.05.