Figure 4
From: Blockade of Smad signaling by 3′-deoxyadenosine: a mechanism for its anti-fibrotic potential
Lack of eIF2α-mediated translational suppression for the effect of 3′-deoxyadenosine (3′-DA). (a, b) NRK-52E cells were treated with 3′-DA for 9 h with or without 10 μM MG132 (a) or 20 μM chloroquine (b) and subjected to western blot analysis of Smad proteins. Quantification of individual signals is shown in graphs. (c) Cells were treated with 3′-DA for indicated time periods and subjected to western blot analysis of phosphorylated eIF2α. The protein level of total eIF2α is shown as a loading control. Quantification of phosphorylated eIF2α normalized by the level of total eIF2α protein is shown. (d) Cells were treated with 50 μM salubrinal for indicated time periods and subjected to western blot analysis of Smad proteins. Quantification of Smad proteins normalized by the level of β-actin is shown. (e, f) NRK-52E cells were transfected with p(CAGA)12-MLP-Luc (e) or pBRE-Luc (f) together with empty vector (Vector), eIF2α-DN or GADD34, treated with TGF-β1 (e) or BMP-4 (f) in the absence or presence of 3′-DA for 24 h and subjected to luciferase assay. NS, not statistically significant.
