Turnover and exchange of nucleosomal histones and their variants, a process long believed to be static in post-replicative cells, remains largely unexplored in brain. Here, we describe a novel mechanistic role for HIRA (histone cell cycle regulator) and proteasomal degradation-associated histone dynamics in the regulation of activity-dependent transcription, synaptic connectivity, and behavior. We uncover a dramatic developmental profile of nucleosome occupancy across the lifespan of both rodents and humans, with the histone variant H3.3 accumulating to near-saturating levels throughout the neuronal genome by mid-adolescence. Despite such accumulation, H3.3-containing nucleosomes remain highly dynamic-in a modification-independent manner-to control neuronal- and glial-specific gene expression patterns throughout life. Manipulating H3.3 dynamics in both embryonic and adult neurons confirmed its essential role in neuronal plasticity and cognition. Our findings establish histone turnover as a critical and previously undocumented regulator of cell type-specific transcription and plasticity in mammalian brain.

Adult women are twice as likely as men to suffer from affective and anxiety disorders, although the mechanisms underlying heightened female stress susceptibility are incompletely understood. Recent findings in mouse Nucleus Accumbens (NAc) suggest a role for DNA methylation-driven sex differences in genome-wide transcriptional profiles. However, the role of another epigenetic process-microRNA (miR) regulation-has yet to be explored. We exposed male and female mice to Subchronic Variable Stress (SCVS), a stress paradigm that produces depression-like behavior in female, but not male, mice, and performed next generation mRNA and miR sequencing on NAc tissue. We applied a combination of differential expression, miR-mRNA network and functional enrichment analyses to characterize the transcriptional and post-transcriptional landscape of sex differences in NAc stress response. We find that male and female mice exhibit largely non-overlapping miR and mRNA profiles following SCVS. The two sexes also show enrichment of different molecular pathways and functions. Collectively, our results suggest that males and females mount fundamentally different transcriptional and post-transcriptional responses to SCVS and engage sex-specific molecular processes following stress. These findings have implications for the pathophysiology and treatment of stress-related disorders in women.

Lasting changes in gene expression in brain reward regions, including nucleus accumbens (NAc), contribute to persistent functional changes in the addicted brain. We and others have demonstrated that altered expression of several candidate transcription factors in NAc regulates drug responses. A recent large-scale genome-wide study from our group predicted transcription factor E2F3 (E2F3) as a prominent upstream regulator of cocaine-induced changes in gene expression and alternative splicing.

Neuronal circuits within the prefrontal cortex (PFC) mediate higher cognitive functions and emotional regulation that are disrupted in psychiatric disorders. The PFC undergoes significant maturation during adolescence, a period when cannabis use in humans has been linked to subsequent vulnerability to psychiatric disorders such as addiction and schizophrenia. Here, we investigated in a rat model the effects of adolescent exposure to Δ9-tetrahydrocannabinol (THC), a psychoactive component of cannabis, on the morphological architecture and transcriptional profile of layer III pyramidal neurons-using cell type- and layer-specific high-resolution microscopy, laser capture microdissection and next-generation RNA-sequencing. The results confirmed known normal expansions in basal dendritic arborization and dendritic spine pruning during the transition from late adolescence to early adulthood that were accompanied by differential expression of gene networks associated with neurodevelopment in control animals. In contrast, THC exposure disrupted the normal developmental process by inducing premature pruning of dendritic spines and allostatic atrophy of dendritic arborization in early adulthood. Surprisingly, there was minimal overlap of the developmental transcriptomes between THC- and vehicle-exposed rats. THC altered functional gene networks related to cell morphogenesis, dendritic development, and cytoskeleton organization. Marked developmental network disturbances were evident for epigenetic regulators with enhanced co-expression of chromatin- and dendrite-related genes in THC-treated animals. Dysregulated PFC co-expression networks common to both the THC-treated animals and patients with schizophrenia were enriched for cytoskeletal and neurite development. Overall, adolescent THC exposure altered the morphological and transcriptional trajectory of PFC pyramidal neurons, which could enhance vulnerability to psychiatric disorders.

Depression is a complex, heterogeneous disorder and a leading contributor to the global burden of disease. Most previous research has focused on individual brain regions and genes contributing to depression. However, emerging evidence in humans and animal models suggests that dysregulated circuit function and gene expression across multiple brain regions drive depressive phenotypes. Here, we performed RNA sequencing on four brain regions from control animals and those susceptible or resilient to chronic social defeat stress at multiple time points. We employed an integrative network biology approach to identify transcriptional networks and key driver genes that regulate susceptibility to depressive-like symptoms. Further, we validated in vivo several key drivers and their associated transcriptional networks that regulate depression susceptibility and confirmed their functional significance at the levels of gene transcription, synaptic regulation, and behavior. Our study reveals novel transcriptional networks that control stress susceptibility and offers fundamentally new leads for antidepressant drug discovery.

Most people exposed to stress do not develop depression. Animal models have shown that stress resilience is an active state that requires broad transcriptional adaptations, but how this homeostatic process is regulated remains poorly understood. In this study, we analyze upstream regulators of genes differentially expressed after chronic social defeat stress. We identify estrogen receptor α (ERα) as the top regulator of pro-resilient transcriptional changes in the nucleus accumbens (NAc), a key brain reward region implicated in depression. In accordance with these findings, nuclear ERα protein levels are altered by stress in male and female mice. Further, overexpression of ERα in the NAc promotes stress resilience in both sexes. Subsequent RNA-sequencing reveals that ERα overexpression in NAc reproduces the transcriptional signature of resilience in male, but not female, mice. These results indicate that NAc ERα is an important regulator of pro-resilient transcriptional changes, but with sex-specific downstream targets.

Increasing evidence supports a role for altered gene expression in mediating the lasting effects of cocaine on the brain, and recent work has demonstrated the involvement of chromatin modifications in these alterations. However, all such studies to date have been restricted by their reliance on microarray technologies that have intrinsic limitations.

The rapid elimination of dying neurons and nonfunctional synapses in the brain is carried out by microglia, the resident myeloid cells of the brain. Here we show that microglia clearance activity in the adult brain is regionally regulated and depends on the rate of neuronal attrition. Cerebellar, but not striatal or cortical, microglia exhibited high levels of basal clearance activity, which correlated with an elevated degree of cerebellar neuronal attrition. Exposing forebrain microglia to apoptotic cells activated gene-expression programs supporting clearance activity. We provide evidence that the polycomb repressive complex 2 (PRC2) epigenetically restricts the expression of genes that support clearance activity in striatal and cortical microglia. Loss of PRC2 leads to aberrant activation of a microglia clearance phenotype, which triggers changes in neuronal morphology and behavior. Our data highlight a key role of epigenetic mechanisms in preventing microglia-induced neuronal alterations that are frequently associated with neurodegenerative and psychiatric diseases.

The role of somatostatin interneurons in nucleus accumbens (NAc), a key brain reward region, remains poorly understood due to the fact that these cells account for < 1% of NAc neurons. Here, we use optogenetics, electrophysiology, and RNA-sequencing to characterize the transcriptome and functioning of NAc somatostatin interneurons after repeated exposure to cocaine. We find that the activity of somatostatin interneurons regulates behavioral responses to cocaine, with repeated cocaine reducing the excitability of these neurons. Repeated cocaine also induces transcriptome-wide changes in gene expression within NAc somatostatin interneurons. We identify the JUND transcription factor as a key regulator of cocaine action and confirmed, by use of viral-mediated gene transfer, that JUND activity in somatostatin interneurons influences behavioral responses to cocaine. Our results identify alterations in NAc induced by cocaine in a sparse population of somatostatin interneurons, and illustrate the value of studying brain diseases using cell type-specific whole transcriptome RNA-sequencing.

Abuse, neglect, and other forms of early life stress (ELS) significantly increase risk for psychiatric disorders including depression. In this study, we show that ELS in a postnatal sensitive period increases sensitivity to adult stress in female mice, consistent with our earlier findings in male mice. We used RNA-sequencing in the ventral tegmental area, nucleus accumbens, and prefrontal cortex of male and female mice to show that adult stress is distinctly represented in the brain's transcriptome depending on ELS history. We identify: 1) biological pathways disrupted after ELS and associated with increased behavioral stress sensitivity, 2) putative transcriptional regulators of the effect of ELS on adult stress response, and 3) subsets of primed genes specifically associated with latent behavioral changes. We also provide transcriptomic evidence that ELS increases sensitivity to future stress through enhancement of known programs of cortical plasticity.

Depression is a common disorder that affects women at twice the rate of men. Here, we report that long non-coding RNAs (lncRNAs), a recently discovered class of regulatory transcripts, represent about one-third of the differentially expressed genes in the brains of depressed humans and display complex region- and sex-specific patterns of regulation. We identified the primate-specific, neuronal-enriched gene LINC00473 as downregulated in prefrontal cortex (PFC) of depressed females but not males. Using viral-mediated gene transfer to express LINC00473 in adult mouse PFC neurons, we mirrored the human sex-specific phenotype by inducing stress resilience solely in female mice. This sex-specific phenotype was accompanied by changes in synaptic function and gene expression selectively in female mice and, along with studies of human neuron-like cells in culture, implicates LINC00473 as a CREB effector. Together, our studies identify LINC00473 as a female-specific driver of stress resilience that is aberrant in female depression.