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. 2017 Mar 8;37(10):2776–2794. doi: 10.1523/JNEUROSCI.2006-14.2016

Figure 1.

Figure 1.

Deregulation of NAD+/NADH ratio and PDH activity, E1alpha subunit expression, and phosphorylation in cells expressing mutant huntingtin (STHdhQ111/Q111). NAD+/NADH ratio (A) and PDH activity (B) were measured in total extracts of wild-type and mutant cells. Total PDH E1alpha protein levels (C) and phosphorylation at regulatory serines 293 (site 1), 300 (site 2), or 232 (site 3) (D) were assessed using the phospho-PDH in cell ELISA kit. Protein levels of PDK1 (E), PDK3 (F), and PDP1c (G) in wild-type and mutant cells were determined by Western blotting. Data are shown as the mean ± SEM of total NAD+/NADH ratio (in pmol/mg protein), total PDH activity (in slope/mg protein), PDH E1alpha/Janus green, phosphorylated form/total PDH E1alpha. Where mentioned, the average value obtained for wild-type was considered 100%. Absolute values for wild-type controls were 1178.00 ± 247.00 AU/min/mg protein (B), 3.60 ± 0.98 (C), 3.94 ± 1.16 for pSer293, 3.28 ± 0.62 for pSer300, and 4.32 ± 1.42 for pSer232 (D). Statistical analysis was performed by Student's t test; tp < 0.05, tttp < 0.001, and ttttp < 0.0001 compared with the respective wild-type/control cells.