Figure 2. Effect of HDAC4 overexpression on neuronal apoptosis.

(A and B) 5 day old culture of CGNs were infected with Ad-GFP or Ad-HDAC4 (Flag tag) and switched to HK or LK medium for 24 hrs. Infected neurons were identified by GFP fluorescence or Flag immunoreactivity and the status of the nuclei were assessed by Syto 13 staining. (A) Subcellular localization of HDAC4 in HK and LK medium. HDAC4 (red) is primarily in the cytoplasm in HK but translocates to the nucleus (green) in LK. Panel B quantifies survival of HDAC4 and GFP infected cells in HK and LK medium as percent of control (viability of GFP-infected neurons in HK). P < 0.006 for comparison with HK-GFP. (C) HT22 cells were infected with HDAC4 and GFP adenovirus and then treated with 2 mM HCA. Because most cells in the culture were infected viability was measured using the MTT assay. Cultures overexpressing HDAC4 proliferate slower than control cultures precluding a direct comparison with GFP infected cultures. Hence, viability (after 24 hrs of treatment) in the presence of HCA is compared with the viability of cultures infected with the same construct (either GFP or HDAC4) in the absence of HCA. P < 0.02 for comparison with control--GFP.