Characterization of a Novel Chromatin Sorting Tool Reveals Importance of Histone Variant H3.3 in Contextual Fear Memory and Motor Learning

The consolidation of short-term labile memories for long-term storage requires transcription and there is growing interest in defining the epigenetic mechanisms regulating these transcriptional events. In particular, it has been hypothesized that combinations of histone post-translational modifications (PTMs) have the potential to store memory by dynamically defining the transcriptional status of any given gene loci. Studying epigenetic phenomena during long-term memory consolidation, however, is complicated by the complex cellular heterogeneity of the brain, in which epigenetic signal from memory-relevant cells can be obscured or diluted by the surrounding milieu. To address this issue, we have developed a transgenic mouse line expressing a tetO-regulated, hemagglutinin (HA)-tagged histone H3.3 exclusively in excitatory neurons of the forebrain. Unlike canonical histones, histone H3.3 is incorporated at promoter regions of transcriptionally active genes in a DNA replication-independent manner, stably “barcoding” active regions of the genome in post-mitotic cells. Immunoprecipitating H3.3-HA containing nucleosomes from the hippocampus will therefore enrich for memory-relevant chromatin by isolating actively transcribed regions of the excitatory neuron genome. To evaluate the validity of using H3.3 “barcoding” to sort chromatin, we performed a molecular and behavioral characterization of the H3.3-HA transgenic mouse line. Expectedly, we find that H3.3-HA is incorporated preferentially at promoter regions of actively-transcribed neuronal genes and that expression can be effectively regulated by doxycycline. Additionally, H3.3-HA overexpression does not adversely affect exploratory or anxiety-related behaviors, nor does it affect spatial memory. Transgenic animals do, however, exhibit deficits in contextual memory and motor learning, revealing the importance of this histone isoform in the brain. Future studies in the H3.3-HA transgenic mouse line will define the combinatorial histone PTM landscape during spatial memory consolidation and will investigate the important contributions of histone H3.3 to the normal functioning of the brain.