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Published on: November 6, 2017
Increased Susceptibility and Intrinsic Apoptotic Signaling in Neurons by Induced HDAC3 Expression
Heather M Schmitt1,2,3, Rachel L Fehrman1, Margaret E Maes4
1Department of Ophthalmology and Visual Sciences, University of Wisconsin-Madison, Madison WI, United States.
Purpose:
Inhibition or targeted deletion of histone deacetylase 3 (HDAC3) is neuroprotective in a variety neurodegenerative conditions, including retinal ganglion cells (RGCs) after acute optic nerve damage. Consistent with this, induced HDAC3 expression in cultured cells shows selective toxicity to neurons. Despite an established role for HDAC3 in neuronal pathology, little is known regarding the mechanism of this pathology.
Methods:
Induced expression of an HDAC3-mCherry fusion protein in mouse RGCs was accomplished by transduction with AAV2/2-Pgk-HDAC3-mCherry. Increased susceptibility to optic nerve damage in HDAC3-mCherry expressing RGCs was evaluated in transduced mice that received acute optic nerve crush surgery. Expression of HDAC3-FLAG or HDAC3-mCherry was induced by nucleofection or transfection of plasmids into differentiated or undifferentiated 661W tissue culture cells. Immunostaining for cleaved caspase 3, localization of a GFP-BAX fusion protein, and quantitative RT-PCR was used to evaluate HDAC3-induced damage.
Results:
Induced expression of exogenous HDAC3 in RGCs by viral-mediated gene transfer resulted in modest levels of cell death but significantly increased the sensitivity of these neurons to axonal damage. Undifferentiated 661W retinal precursor cells were resilient to induced HDAC3 expression, but after differentiation, HDAC3 induced GFP-BAX recruitment to the mitochondria and BAX/BAK dependent activation of caspase 3. This was accompanied by an increase in accumulation of transcripts for the JNK2/3 kinases and the p53-regulated BH3-only gene Bbc3/Puma. Cell cycle arrest of undifferentiated 661W cells did not increase their sensitivity to HDAC3 expression.
Conclusions:
Collectively, these results indicate that HDAC3-induced toxicity to neurons is mediated by the intrinsic apoptotic pathway.
Insights
Histone deacetylase 3 (HDAC3) expression increases neuronal susceptibility to axonal damage. This toxicity is mediated by the intrinsic apoptotic pathway, involving BAX/BAK activation and caspase 3.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Biology
Background:
- Histone deacetylase 3 (HDAC3) inhibition is neuroprotective in neurodegenerative diseases.
- HDAC3 expression in cultured cells demonstrates selective neuronal toxicity.
- The precise mechanism of HDAC3-induced neuronal pathology remains unclear.
Purpose of the Study:
- To investigate the mechanism underlying HDAC3-induced neuronal toxicity.
- To evaluate the role of HDAC3 in retinal ganglion cell (RGC) susceptibility to axonal damage.
Main Methods:
- Adeno-associated virus (AAV)-mediated gene transfer to induce HDAC3-mCherry expression in mouse RGCs.
- Assessment of RGCs' susceptibility to optic nerve crush injury.
- Evaluation of HDAC3-induced damage in 661W retinal precursor cells using immunostaining and qRT-PCR.
Main Results:
- Induced HDAC3 expression in RGCs increased sensitivity to axonal damage.
- Differentiated 661W cells expressing HDAC3 showed BAX recruitment to mitochondria and caspase 3 activation.
- HDAC3 expression upregulated transcripts for JNK2/3 kinases and the p53-regulated gene Bbc3/Puma.
Conclusions:
- HDAC3-induced neuronal toxicity is mediated by the intrinsic apoptotic pathway.
- This pathway involves mitochondrial outer membrane permeabilization and caspase activation.
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01:31The Intrinsic Apoptotic Pathway
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01:30Apoptosis
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