Opposing Effects of Sirtuins on Neuronal Survival: SIRT1-Mediated Neuroprotection Is Independent of Its Deacetylase Activity
Figure 1
Effect of SIRT 1–7 expression on neuronal survival.
(A) Sirtuin family expression and cerebellar granule neuron survival. Expression plasmids encoding Flag-tagged forms of SIRT 1–7 were transfected into CGNs and the cultures were switched to HK or LK medium. The proportion of transfected neurons that were apoptotic was quantified 24 hrs after the switch and compared with control cultures which were transfected with CMV-GFP. Viability for transfected cells was determined using DAPI staining of chromatin (*indicates p<.05 compared with GFP HK (control); # indicates p<.05 compared with GFP LK). (B) Expression of other SIRT1 constructs in neurons confirms neuroprotection. Neurons were transfected with Flag-SIRT1 (tag on the N-terminus), HA-SIRT1, or Trunc-SIRT1-Flag and switched to HK or LK medium. The proportion of transfected neurons that were apoptotic was quantified 24 hrs after the switch and compared with control cultures which were transfected with CMV-GFP. Apoptosis was determined for transfected cells using DAPI staining (* indicates p<.05 compared with GFP LK; control = GFP HK). (C) SIRT5 localization and CGNs survival. SIRT5 localization in CGNs was determined by immunocytochemistry following SIRT5-Flag transfection and viability assessed using DAPI staining 24 after HK and LK treatment. The localization of viable and apoptotic SIRT5-Flag expressing cells was scored as nuclear and cytoplasmic or mitochondrial (* indicates p<.05 compared with nuclear-cytoplasmic).