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Fig. 4 | Molecular Brain

Fig. 4

From: Down-regulation of p21-activated serine/threonine kinase 1 is involved in loss of mesencephalic dopamine neurons

Fig. 4

PAK1-DN expression reduces the Bcl-2 level. a PAK1-DN expression reduces level of anti-apoptotic Bcl-2 family proteins. Lysates of SH-SY5Y cells expressing GFP, PAK1-CA, or PAK1-DN were subjected to Western Blot analysis using antibodies against anti-apoptotic or pro-apoptotic Bcl-2 family proteins. b Reduction in Bcl-2 induced by PAK1-DN expression is mediated by ubiquitin-dependent protein degradation. HEK cells were transfected with 1, 1.5, 2, or 3 μg of PAK1-DN vector and incubated for 36 h and treated with 10 μM lactacystin or without for 12 h. The cell lysates were subjected to Western blot analyses using the indicated antibodies. c Quantification of Western blot analysis. To obtain relative values of Bcl-2 or Bcl-xL level, band intensities from 1 and 2 μg expression of PAK1-DN was normalized by that from 3 μg expression of PAK1-DN in each blot. Reduction in Bcl-2 induced by dose-dependent expression of PAK1-DN reversed by lactacystin treatment [−Lact: 362.68 % (1 μg) → 182.6 % (2 μg) → 100.0 % (3 μg), +Lact: 118.5 ± 19.26 %, n = 3 (1 μg) → 109.05 ± 15.10 %, n = 3 (2 μg) → 100.0 %, n = 3 (3 μg)]. Bcl-xL level was not altered [−Lact: 127.7 % (1 μg) → 148.4 % (2 μg:) → 100.0 % (3 μg), +Lact: 97.0 ± 13.89 %, n = 3 (1 μg) → 100.94 ± 12.59 %, n = 3 (2 μg) → 100.0 %, n = 3 (3 μg)]. d PAK1-DN expression increased the ubiquitination of Bcl-2 protein, but PAK1-CA expression did not increase the ubiquitination of Bcl-2 protein

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