Biology subjects

West, A. E.

Publications and source records attributed to West, A. E..

8 recordsLinked to original sources

bioRxiv2026

The linker histone H1.4 condenses chromatin in maturing postmitotic neurons

H1 histones are abundant nuclear proteins that bind to linker DNA at the entry and exit points of nucleosomes. Although individual H1 family members play partially redundant roles in chromatin organization, the discovery of disease-associated mutations in H1 genes has raised interest in their cell-type specific functions. Heterozygous, de novo frameshift mutations in H1-4 cause the neurodevelopmental disorder Rahman Syndrome, which is characterized by mild to severe intellectual disability along with other neurological and morphological features. H1.4 has mostly been studied in dividing cells, and its expression in the brain was poorly understood. Here we characterized the expression of H1f4 mRNA and H1.4 protein in the brains of male and female mice across postnatal development. Using an epitope-tagged H1f4 knockin mouse, we show that this linker histone is robustly expressed throughout the brain including in mature, post-mitotic neurons of adult mice. By chromatin immunoprecipitation, we observe that H1.4 binds broadly across the genome in neural progenitors and accumulates in heterochromatin over the course of neuronal maturation. Finally we show that developmental maturation of chromatin compaction in cerebellar granule neurons is disrupted in H1f2/H1f4 double knockout mice. These data raise the possibility that the neurological changes in Rahman Syndrome may arise from disrupted functions of histone H1.4 in neurons.

cell biology↗

bioRxiv2026

Transcriptional plasticity of ventral tegmental area neurons induced after cessation of chronic cocaine exposure is required for incubation of cocaine seeking

Chronic cocaine triggers changes in brain function that persist long after drug taking has ceased. Relapse to substance use during abstinence can be triggered by drug-associated cues, implicating the drug-free period after chronic cocaine as a time when the probability of relapse is modulated by plasticity mechanisms. Here, we studied the regulation and function of transcriptional plasticity activated in a time-dependent manner in dopaminergic and glutamatergic neurons of the mouse ventral tegmental area (VTA) over a drug-free period after chronic cocaine. We used in vivo dCas9/CRISPR inhibition to demonstrate a causal role for Brain-Derived Neurotrophic Factor transcription in the incubation of cocaine seeking during abstinence, and we used single-nucleus sequencing to identify a program of gene regulation induced in VTA neurons selectively by the prolonged absence of cocaine. These data advance the understanding of plasticity mechanisms that modulate reward functions of VTA neurons, which may be a main factor propagating relapse in substance use disorders.

neuroscience↗

bioRxiv2026

CRISPR inhibition of activity-dependent Arc expression in the adult mouse brain has limited effects on plasticity in visual cortex and nucleus accumbens

Neuronal activity drives long-lasting change in circuit function by inducing the expression of gene products that modulate the function of synapses. Arc is one of the most robustly activity-regulated neuronal genes, being induced broadly in neurons by a wide range of physiologically-relevant stimuli. The ability of Arc to promote internalization of AMPA-type glutamate receptors plays an important role in activity-dependent refinement of synaptic connectivity during development. However, the consequences of Arc induction for circuit plasticity in the adult brain are less well understood. We reasoned that we could test the requirement for experience-induced Arc expression in downstream plasticities by disrupting regulatory elements that mediate the inducibility of Arc transcription. To achieve this goal, we developed and validated a CRISPR-based inhibition strategy to conditionally block stimulus-induced expression of Arc in specific regions of the brains of male and female adult mice in vivo. We show that recruiting transgenic dCas9-KRAB to decrease transcriptional activity of either the promoter or the synaptic activity-regulated enhancer of the Arc gene is sufficient to block Arc protein expression in visual cortex (V1) following light exposure or in the nucleus accumbens (NAc) following cocaine administration. However, loss of Arc in V1 failed to alter plasticity of orientation selectivity and loss of Arc in NAc did not block cocaine conditioned place preference or novel object recognition memory. These data show that the relationship between Arc induction and plasticity is not universal and suggest that additional contextual factors determine the functional consequences of Arc induction for plasticity. Significance StatementArc is a neuronal activity-regulated gene whose expression is robustly induced in the adult brain by a wide range of stimuli that drive plasticity. However, the functional requirements of Arc induction for brain plasticity are incompletely understood. Here, we show that we can use transgenic mice expressing the CRISPR-based transcriptional repressor dCas9-KRAB to block stimulus-induced expression of Arc in specific regions of the adult mouse brain. Despite highly effective inhibition of Arc induction, we found that many forms of plasticity remained intact. These data deepen understanding of the contextual importance of activity-induced Arc expression in the adult brain.

neuroscience↗

bioRxiv2024

Genome binding properties of Zic transcription factors underlie their changing functions during neuronal maturation

BackgroundThe Zic family of transcription factors (TFs) promote both proliferation and maturation of cerebellar granule neurons (CGNs), raising the question of how a single, constitutively expressed TF family can support distinct developmental processes. Here we use an integrative experimental and bioinformatic approach to discover the regulatory relationship between Zic TF binding and changing programs of gene transcription during CGN differentiation. ResultsWe first established a bioinformatic pipeline to integrate Zic ChIP-seq data from the developing mouse cerebellum with other genomic datasets from the same tissue. In newborn CGNs, Zic TF binding predominates at active enhancers that are co-bound by developmentally-regulated TFs including Atoh1, whereas in mature CGNs, Zic TF binding consolidates toward promoters where it co-localizes with activity-regulated TFs. We then performed CUT&RUN-seq in differentiating CGNs to define both the time course of developmental shifts in Zic TF binding and their relationship to gene expression. Mapping Zic TF binding sites to genes using chromatin looping, we identified the set of Zic target genes that have altered expression in RNA-seq from Zic1 or Zic2 knockdown CGNs. ConclusionOur data show that Zic TFs are required for both induction and repression of distinct, developmentally regulated target genes through a mechanism that is largely independent of changes in Zic TF binding. We suggest that the differential collaboration of Zic TFs with other TF families underlies the shift in their biological functions across CGN development.

neuroscience↗

bioRxiv2023

Single-nucleus transcriptional profiling of GAD2-positive neurons from mouse lateral habenula reveals distinct expression of neurotransmission- and depression-related genes

Glutamatergic projection neurons of the lateral habenula (LHb) drive behavioral state modulation by regulating the activity of midbrain monoaminergic neurons. Identifying circuit mechanisms that modulate LHb output is of interest for understanding control of motivated behaviors. A small population of neurons within the medial subnucleus of the mouse LHb express the GABAergic synthesizing enzyme GAD2, and they can inhibit nearby LHb projection neurons; however, these neurons lack markers of classic inhibitory interneurons and they co-express the vesicular glutamate transporter VGLUT2. To determine the molecular phenotype of these neurons, we genetically tagged the nuclei of GAD2-positive cells and used fluorescence-activated nuclear sorting to isolate and enrich these nuclei for single nuclear RNA sequencing (FANS-snRNAseq). Our data confirm that GAD2+/VGLUT2+ neurons intrinsic to the LHb co-express markers of both glutamatergic and GABAergic transmission and that they are transcriptionally distinct from either GABAergic interneurons or habenular glutamatergic neurons. We identify gene expression programs within these cells that show sex-specific differences in expression and that are implicated in major depressive disorder (MDD), which has been linked to LHb hyperactivity. Finally, we identify the Ntng2 gene encoding the cell adhesion protein Netrin-G2 as a marker of LHb GAD2+/VGLUT+ neurons and a gene product that may contribute to their target projections. These data show the value of using genetic enrichment of rare cell types for transcriptome studies, and they advance understanding of the molecular composition of a functionally important class of GAD2+ neurons in the LHb.

neuroscience↗

bioRxiv2022

Bidirectional changes in postmitotic H3K27me3 distributions underlie cerebellar granule neuron maturation dynamics

The functional maturation of neurons is a prolonged process that extends well beyond mitotic exit and is mediated by the chromatin-dependent orchestration of gene transcription programs. We find that the postnatal maturation of cerebellar granule neurons (CGNs) requires dynamic changes in the genomic distribution of histone H3 lysine 27 trimethylation (H3K27me3), demonstrating a function for this chromatin modification beyond its role in cell fate specification. The developmental loss of H3K27me3 at promoters of genes that turn on as CGNs mature is facilitated by the lysine demethylase, and ASD-risk gene, Kdm6b. Interestingly, inhibition of the H3K27 methyltransferase EZH2 in newborn CGNs not only blocks the repression of progenitor genes but also impairs the induction of mature CGN genes, showing the importance of bidirectional H3K27me3 regulation across the genome. These data demonstrate that H3K27me3 turnover in developing postmitotic neurons regulates the temporal coordination of gene expression programs that underlie functional neuronal maturation.

molecular biology↗

bioRxiv2021

Cell-type specific transcriptional adaptations of nucleus accumbens interneurons to amphetamine

Parvalbumin-expressing (PV+) interneurons of the nucleus accumbens (NAc) play an essential role in the addictive-like behaviors induced by psychostimulant exposure. To identify molecular mechanisms of PV+ neuron plasticity, we isolated interneuron nuclei from the NAc of male and female mice following acute or repeated exposure to amphetamine (AMPH) and sequenced for cell type-specific RNA expression and chromatin accessibility. AMPH regulated the transcription of hundreds of genes in PV+ interneurons, and this program was largely distinct from that regulated in other NAc GABAergic neurons. Chromatin accessibility at enhancers predicted cell-type specific gene regulation, identifying transcriptional mechanisms of differential AMPH responses. Finally, we observed dysregulation of multiple PV-specific, AMPH-regulated genes in an Mecp2 mutant mouse strain that shows heightened behavioral sensitivity to psychostimulants, suggesting the functional importance of this transcriptional program. Together these data provide novel insight into the cell-type specific programs of transcriptional plasticity in NAc neurons that underlie addictive-like behaviors.

neuroscience↗

bioRxiv2017

Bayesian inference of transcription dynamics from population snapshots of single-molecule RNA FISH in single cells

Single-molecule RNA fluorescence in situ hybridization (smFISH) provides unparalleled resolution on the abundance and localization of nascent and mature transcripts in single cells. Gene expression dynamics are typically inferred by measuring mRNA abundance in small numbers of fixed cells sampled from a population at multiple time-points after induction. The sparse data that arise from the small number of cells obtained using smFISH present a challenge for inferring transcription dynamics. Here, we developed a computational pipeline (BayFish) to infer kinetic parameters of gene expression from smFISH data at multiple time points after induction. Given an underlying model of gene expression, BayFish uses a Monte Carlo method to estimate the Bayesian posterior probability of the model parameters and quantify the parameter uncertainty given the observed smFISH data. We tested BayFish on smFISH measurements of the neuronal activity inducible gene Npas4 in primary neurons. We showed that a 2-state promoter model can recapitulate Npas4 dynamics after induction and we inferred that the transition rate from the promoter OFF state to the ON state is increased by the stimulus.\n\nAuthor SummaryGene expression can exhibit cell-to-cell variability due to the stochastic nature of biochemical reactions. Single cell assays (e.g. smFISH) directly quantify stochastic gene expression by measuring the number of active promoters and transcripts per cell in a population of cells. The data are distributions and their shape and time-evolution contain critical information on the underlying process of gene expression. Recent work has combined models of stochastic gene expression with maximum likelihood methods to infer kinetic parameters from smFISH distributions. However, these approaches do not provide a probability distribution or likelihood of model parameters inferred from the smFISH data. This information is useful because it indicates which parameters are loosely constrained by the data and suggests follow up experiments. We developed a suite of MATLAB programs (BayFish) that estimate the Bayesian posterior probability of model parameters from smFISH data. The user specifies an underlying model of stochastic gene expression with unknown parameters ({theta}) and provides smFISH data (Y). BayFish uses a Monte Carlo algorithm to estimate the Bayesian posterior probability P({theta}|Y) of model parameters. BayFish is easily modified and can be applied to other models of stochastic gene expression and smFISH data sets.

systems biology↗