Figure 3

Activation of UPR in carriers of GD mutations. A. RNA was isolated from skin fibroblasts that originated from carriers of GD mutations, and the corresponding cDNA was used for quantitative RT-PCR with primers specific for human BiP or CHOP. GAPDH was used as a normalizing control. B. cDNA, prepared as in (A), was subjected to RT-PCR, with primers specific for the spliced form of human Xbp1. GAPDH was used as a normalizing control. C. The results (three different experiments) were quantified as explained in the legend to Figure 2 and the values obtained for normal cells were considered 1. D. Protein lysates, prepared from the above-mentioned cells, were subjected to western blotting and interaction with anti phosphorylated eIF2α antibodies (p-eIF2α). As a loading control, the blots were interacted with anti eIF2α antibodies. E. The results (three different experiments) were quantified as explained in the legend to Figure 3B, and the values obtained for normal cells were considered 1. There are two blots, each with a normal control. This is due to the fact that cell lysates were prepared at different times, depending on the growth rate of the cell lines and, therefore, ran on different gels. Significance: * < 0.05; ** < 0.01; *** < 0.005. The genotypes for B and D are shown in C and E, respectively.