Conditional Deletion of Histone Deacetylase-4 in the Central Nervous System Has No Major Effect on Brain Architecture or Neuronal Viability

Histone deacetylases (HDACs) are enzymes that catalyze the removal of acetyl groups from histones as well as a long and growing list of nonhistone nuclear, cytoplasmic, and mitochondrial proteins (Yang and Seto, 2008; Haberland et al., 2009). Mammals express 18 different histone deacetylases that are divided into four groups. Class I HDACs (HDAC1, –2, –3, and –8) are ubiquitously expressed and localize to the nucleus, with the exception of HDAC3, which is both nuclear and cytoplasmic. Class II HDACs (HDAC4, –5, –6, –7, –9, and –10) are specifically expressed in a variety of tissues, with highest expression in the brain and skeletal muscle. Some of the class II HDACs have been shown to translocate between the nucleus and cytoplasm in a phosphorylation-dependent manner. Class III HDACs (SIRT1–7) are structurally distinct from the other three classes of classical HDACs (I, II, and IV) and are referred to as the sirtuins. While still encompassing histone deacetylase activity, sirtuins are considered nonclassical HDACs because of their requirement of NAD+ for their activity, as opposed to the zinc-dependence of classical HDACs (Yang and Seto, 2008; Haberland et al., 2009). Finally, class IV HDACs consists of only one member, HDAC11, which shares sequence similarity with both class I and class II HDACs. A solid body of evidence suggests that HDACs regulate neuronal viability (Morrison et al., 2007; Kazantsev and Thompson, 2008; Majdzadeh et al., 2008a; Sleiman et al., 2009; D’Mello, 2009). Whereas HDAC3 has been shown to promote neuronal death (Bardai and D’Mello, 2011), other HDACs maintain neuronal survival and protect neurons from apoptotic stimuli (Iwata et al., 2005; Morrison et al., 2006; Pandey et al., 2007; Boyault et al., 2007; Pfister et al., 2008; Ma and D’Mello, 2011); among these is HDAC4. We previously reported that elevated expression of HDAC4 protects cultured cerebellar granule neurons (CGNs) and cortical neurons from death (Majdzadeh et al., 2008b). Although HDAC4 normally resides in the cytoplasm, we found that, in neurons primed to die, HDAC4 translocates to the nucleus, where its protection was mediated (Majdzadeh et al., 2008b). This was confirmed by the inability of an HDAC4 mutant, lacking a 72-amino-acid region containing the nuclear loacalization signal, to protect neurons primed to die. A neuroprotective role for HDAC4 was also described by Chen and Cepko (2009) for the mouse retina. These authors found that RNAi-mediated knockdown of HDAC4 expression during normal retinal development led to apoptosis of rod photoreceptors and bipolar interneurons. On the other hand, retrovirus-mediated overexpression of HDAC4 reduced naturally occurring cell death of retinal neurons (Chen and Cepko, 2009). Furthermore, HDAC4 over-expression prolonged photoreceptor survival in a mouse model of retinal degeneration (Chen and Cepko, 2009). Surprisingly, and in contradiction to what was found in cultured CGNs and cortical neurons, neuroprotection by HDAC4 in the intact retina was mediated in the cytoplasm (Chen and Cepko, 2009). More recently, interaction between HDAC4 and specific HSP40 proteins was shown to be necessary for the ability of these HSP40 proteins to suppress polyglutamine protein aggregation and toxicity (Hageman et al., 2010). Together these studies strongly suggest that HDAC4 promotes neuronal survival both in vivo and in vitro. HDAC4 null mice have severe bone malformations resulting from chondrocyte hypertrophy and die perinatally (Vega et al., 2004). An examination of the brains of these mice revealed Purkinje cell degeneration starting within a few days of birth (Majdzadeh et al., 2008b), consistent with the conclusion that HDAC4 is required for neuronal survival. However, the early death of these mice precludes a more detailed analysis of the consequences of HDAC4 deficiency to neuronal well-being in the postnatal and adult brain. We therefore generated two separate lines of conditional knockout (cKO) mice in which HDAC4 is deleted selectively in the cortex and hippocampus or in neural progenitor cells of the CNS by breeding HDAC4-flox mice with mice expressing Cre through a Thy1 or nestin promoter, respectively. Surprisingly, both lines of HDAC4 cKO mice are normal in appearance, breeding, and performance in locomotor tests. Histological analysis of two mouse lines in which HDAC4 was deleted selectively in the cortex and hippocampus as well as the entire CNS shows no increase in cell death in the brain and no discernible abnormality in brain cytoarchitecture.