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Histone variants H2A.Z and H3.3 are epigenetic regulators of memory, but roles of other variants are not well characterized. macroH2A (mH2A) is a structurally unique histone that contains a globular macrodomain connected to the histone region by an unstructured linker. Here we assessed if mH2A regulates memory and if this role varies for the two mH2A-encoding genes, H2afy (mH2A1) and H2afy2 (mH2A2). We show that fear memory is impaired in mH2A1, but not in mH2A2-deficient mice, whereas both groups were impaired in a non-aversive spatial memory task. However, impairment was larger for mH2A1- deficient mice, indicating a preferential role for mH2A1 over mH2A2 in memory. Accordingly, mH2A1 depletion in the mouse hippocampus resulted in more extensive transcriptional de-repression compared to mH2A2 depletion. mH2A1-depleted mice failed to induce a normal transcriptional response to fear conditioning, suggesting that mH2A1 depletion impairs memory by altering transcription. Using chromatin immunoprecipitation (ChIP) sequencing, we found that both mH2A proteins are enriched on transcriptionally repressed genes, but only mH2A1 occupancy was dynamically modified during learning, displaying reduced occupancy on upregulated genes after training. These data identify mH2A as a regulator of memory and suggest that mH2A1 supports memory by repressing spurious transcription and promoting learning-induced transcriptional activation.

SUMMARY Rapid removal of histone H2A.Z from neuronal chromatin is a key step in learning-induced gene expression and memory formation, but mechanisms underlying learning-induced H2A.Z removal are unclear. Anp32e was recently identified as an H2A.Z-specific histone chaperone that removes H2A.Z from nucleosomes in dividing cells, but its role in non-dividing neurons is unclear. Moreover, prior studies investigated Anp32e function under steady-state rather than stimulus-induced conditions. Here, we show that Anp32e regulates H2A.Z binding in neurons under steady-state conditions, with lesser impact on stimulus-induced H2A.Z removal. Functionally, Anp32e depletion leads to H2A.Z-dependent impairment in transcription and dendritic arborization in cultured hippocampal neurons, as well as impaired recall of contextual fear memory and transcriptional regulation. Together, these data indicate that Anp32e regulates behavioral and morphological outcomes by preventing H2A.Z accumulation in chromatin rather than by regulating activity-mediated H2A.Z dynamics., Graphical abstract, In brief Histone variants are highly dynamic regulators of neural plasticity and memory formation. Stefanelli et al. show that histone H2A.Z in neurons is regulated by Anp32e, whereby Anp32e keeps H2A.Z binding in check. In cases of Anp32e deficiency, neurons accumulate H2A.Z, their development is impaired, and mice exhibit impaired memory formation.

Emerging evidence suggests that histone variants are novel epigenetic regulators of memory, whereby histone H2A.Z suppresses fear memory. However, it is not clear if altered fear memory can also modify risk for PTSD, and whether these effects differ in males and females. Using conditional-inducible H2A.Z knockout (cKO) mice, we showed that H2A.Z binding is higher in females and that H2A.Z cKO enhanced fear memory only in males. However, H2A.Z cKO improved memory on the non-aversive object-in-place task in both sexes, suggesting that H2A.Z suppresses non-stressful memory irrespective of sex. Given that risk for fear-related disorders, such as PTSD, is biased toward females, we examined whether H2A.Z cKO also has sex-specific effects on fear sensitization in the stress-enhanced fear learning (SEFL) model of PTSD, as well as associated changes in pain sensitivity. We found that H2A.Z cKO reduced stress-induced sensitization of fear learning and pain responses preferentially in female mice, indicating that the effects of H2A.Z depend on sex and the type of task, and are influenced by history of stress. These data suggest that H2A.Z may be a sex-specific epigenetic risk factor for PTSD susceptibility, with implications for developing sex-specific therapeutic interventions.