Nucleoredoxin-Dependent Targets and Processes in Neuronal Cells

Figure 4

Redox state of cytosolic peroxiredoxins, cofilin, and GAPDH in Nrx-depleted HeLa cells. The redox state of Prx1 and Prx2 was analyzed in Nrx-depleted HeLa cells by 2-Cys Prx redox blot (a). Prior to lysis, free thiols were alkylated with NEM, and the monomer and dimer levels of the two proteins were analyzed via the 2-Cys Prx redox blot using specific antibodies against Prx1 and Prx2, respectively (a). The quantification of the Western blot signals using ImageJ (b) shows that more dimeric, i.e., oxidized Prx1, is present in the absence of Nrx ( value = 0.1). (c-f) Analysis of the redox state of potential Nrx targets by 2-dimensional diagonal gel electrophoresis and Western blotting. Both GAPDH and cofilin showed an increased staining of spots that fell below the diagonal in the second dimension in the presence of Nrx (c, e), compared to HeLa cells lacking Nrx (d, f). In the overlayed pictures, the total protein content in the diagonal is depicted in blue, the specific protein of interest in black.

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