Biology subjects

Dougherty, J. D.

Publications and source records attributed to Dougherty, J. D..

At least 19 recordsLinked to original sources

bioRxiv2026

Protein expression is amplified by stabilizing mRNA using gene-specific antisense oligonucleotides

Effective and safe strategies to restore gene expression in haploinsufficiency disorders or to augment protective protein signaling have long been attractive but challenging therapeutic goals. Here, we demonstrate the potential of antisense oligonucleotides (ASOs) to upregulate gene targets through mRNA stabilization. The 3 untranslated regions (UTRs) of mRNAs regulate stability via activating and repressive elements. One prominent destabilizing cis-element found in ~30% of transcripts is the Pumilio RNA-binding protein recognition element (PRE), which can be targeted to enable gene-specific protein upregulation. A scan for PREs highlights hundreds of disease-relevant transcripts that may be targetable via ASO-mediated gene-specific blocking of Pumilio binding. We show that masking specific PREs with ASOs produces sustained protein upregulation through mRNA stabilization using multiple reporter and endogenous gene targets. Using PRE-masking ASOs, we neutralized PRE-mediated destabilization of TANK-binding kinase-1 (TBK1) mRNA and restored TBK1 protein expression in participant-derived fibroblasts with TBK1 haploinsufficiency, causal for amyotrophic lateral sclerosis. Additionally, by targeting PRE sites in the myoprotective gene follistatin (FST), we achieved effective FST mRNA upregulation in tested mouse tissues. These findings suggest that ASO strategies that neutralize destabilizing 3UTR sequences like PREs may provide a powerful approach to upregulate haploinsufficiency-related genes or enhance protective protein levels for therapeutic benefit.

molecular biology↗

bioRxiv2026

Sex differences in exploration-exploitation strategies during home-cage decision making

The exploration-exploitation trade-off refers to the conflict between using known strategies that reliably yield reward (exploitation) and sampling uncertain options that might yield better outcomes (exploration). Dysregulation of this balance is implicated in neuropsychiatric disease, and while sex differences in this balance have been described, the biological bases remain unclear. To quantify sex differences in this trade-off, we tested mice (n=74 male, 62 female) on four home-cage based foraging tasks with an operant pellet dispensing device, Feeding Experimentation Device 3 (FED3). Mice completed the tasks continuously over multiple days and the tasks were their only source of food. Across multiple tasks, males showed higher win-stay behaviour than females, indicating greater exploitation of previously rewarded actions, an effect that was modest in size but highly significant. Power analyses revealed that >30 mice per sex were needed to detect these modest but significant sex differences with 80% power. No consistent sex differences were observed in pellet intake, suggesting that differences in exploitation did not reflect differences in hunger drive or demand for pellets. Exploitation is a more efficient strategy when environmental parameters are fixed, while exploration can be more advantageous when parameters such as reward locations are changing and uncertain. We tested this idea by re-running our mice in a probabilistic foraging task, where actions led to uncertain probabilities of reward. While males continued to show higher levels of win-stay behaviour on this task, this no longer led to increases in accuracy. Behavioural modelling also supported this framework, demonstrating that stronger win-stay behaviour was most advantageous in deterministic models, and less advantageous in probabilistic models. Together, our findings demonstrate that male and female mice have small but significant differences in their exploration-exploitation balance, which leads to more accurate foraging in certain, but not uncertain, environments.

neuroscience↗

bioRxiv2025

Inducible Calling Cards: Developing Mouse Reagents for Experimentally Controlled Transposon Insertion in vivo.

The piggyBac transposase enables robust forward genetic screens in mice. Further, fusions of piggyBac with specific transcription factors (TFs) enables recovery of recoverable transposons (Calling Cards) for recording location of DNA binding events in vitro and in vivo. Such applications would be enhanced by engineering inducible transposases, such that timing of recording could be precisely controlled in vivo, as has been previously developed in vitro. Here, we tested two approaches for applying inducible Calling Cards (iCC) in the murine brain. We engineered knock-ins of inducible versions of two TFs: Jun, an immediate early gene that serves as a proxy for neural activity, and Sp1, a promiscuous binder of CpG unmethylated regions that indicates active promoters. We fused hyperPiggyBac transposase and a tamoxifen-inducible domain (ERT2) to these TFs and tested the system for efficacy and temporal control of insertions, both in vitro and in vivo. Jun-iCC mice developed normally with no behavioral abnormalities, showed tamoxifen-dependent recording, and captured neural activity during pharmacologically induced seizures. Jun-iCC yields relatively low numbers of insertions, likely due to the transient expression of Jun. In contrast, Sp1-iCC provided substantially higher insertion numbers, but transgenic animals exhibited developmental abnormalities, including reduced viability, anophthalmia, and reduced body weight, suggesting that ERT2 domains may sequester Sp1 and thus significantly impact development. Nonetheless, these inducible Calling Cards mouse lines enable drug-inducible integration of transposon cargo into specific loci in the living mouse. SIGNIFICANCE STATEMENTTransposases, like the more widely used recombinases (Cre, Flp, etc.), enable the integration of a variety of DNA sequences into the genome, but without dependence on LoxP sites. However, few mouse reagents exist for transposases. Cre has been adapted to be drug-inducible, as well as activity-regulated (e.g., FOS-TRAP). Thus, additional applications would become available if transposases could also be drug-controlled and activity-dependent. Further, there may be benefits to targeting integration to specific genomic region types (e.g., promotors). Here, we report results testing mouse lines for two drug-inducible transposase fusions to transcription factors (TF) - the promoter binding SP1, and activity dependent Jun. Both allowed genomic integration, with viability, efficiency, inducibility and targeting varying by TF-fusion.

neuroscience↗

bioRxiv2025

A single binge ethanol exposure is apoptotic within hours across neurodevelopment and partially regulated by the Myt1l gene.

Ethanol rapidly produces widespread neuronal apoptosis during early development, but this susceptibility declines as the brain matures. In previous research, we found Myt1l (a proneuronal transcription factor) mutations can cause precocious differentiation, neuronal immaturity, and transcriptomic alterations, including many in apoptotic regulators. Therefore, we used a recently developed Myt1l haploinsufficient mouse model to examine this genes effects on ethanol-induced apoptosis across different developmental stages. We discovered that haploinsufficiency can moderately influence vulnerability to ethanol in a complex, age- and cell type-specific manner: apoptosis was reduced on P7, increased P21, but unaffected on P60. Remarkably, we also discovered the previously unrecognized ability of a single binge of ethanol to rapidly increase apoptosis within six hours in early adolescent and adult wild-type mice occurring in microglia and the newborn granule neurons in the hippocampus. This suggests apoptosis is an underappreciated contributor to ethanols neuropathology at older ages and, translated to human use, occurs far more frequently than previously recognized.

neuroscience↗

bioRxiv2025

Prior epigenetic status predicts future susceptibility to seizures in mice

Wide variation of responses to identical stimuli presented to genetically inbred mice suggests the hypothesis that stochastic epigenetic variation during neurodevelopment can mediate such phenotypic differences. However , this hypothesis is largely untested since capturing pre-existing molecular states requires non-destructive, longitudinal recording. Therefore, we tested the potential of Calling Cards (CC) to record transient neuronal enhancer activity during postnatal development, and thereby associate epigenetic variation with a subsequent phenotypic presentation - degree of seizure response to the pro-convulsant pentylenetetrazol. We show that recorded differences in epigenetics at 243 loci predict a severe vs. mild response, and that these are enriched near genes associated with human epilepsy. We also validated pharmacologically a seizure -modifying role for two novel genes, Htr1f and Let7c. This proof-of-principle supports using CC broadly to discover predisposition loci for other neuropsychiatric traits and behaviors. Finally, as, human disease is also influenced by non-inherited factors, similar epigenetic predispositions are possible in humans.

genomics↗

bioRxiv2025

Single-Cell Resolution of Individual Variation in Hypothalamic Neurons Allows Targeted Manipulation Affecting Social Motivation

Despite decades of research, connecting molecular and cellular phenotypes to complex behavioral traits remains an elusive goal1. Social motivation exhibits individual trait variation2, which we hypothesize is mediated by molecular and cellular variability across hypothalamic neurons. To test this, we generated single-nucleus RNA-sequencing profiles3,4 of >120,000 neurons from tuberal hypothalamus and adjacent thalamus in 36 mice, balanced across sex and autism-associated mutation5, with all mice assessed for social motivation2. First, we show that molecular activation patterns predict behavior across individuals: specifically, activation of paraventricular Agtr1a+ (angiotensin receptor 1a) neurons predicted reduced social behavior. Subsequent inhibition of AGTR1A with telmisartan--an FDA-approved antihypertensive6--improved social orienting. Second, we show natural variation in neuronal proportions--likely arising from stochastic developmental events7--is sufficient to shape adult behavior even among genetically-identical individuals: we identified multiple neuronal populations whose relative abundance predicted social reward-seeking behavior. Chemogenetic inhibition of one such population, Nxph4+ neurons of the postero-lateral hypothalamus8, suppressed multiple aspects of social motivation. This work establishes proof-of-principle for an approach where single-cell genomics precisely maps neural substrates governing behavior. This approach revealed that stochastic variations in neuronal architecture deterministically influence social motivation, and enabled identification of therapeutically-actionable targets with immediate translational potential for disorders with social deficits.

genetics↗

bioRxiv2024

A survey of hypothalamic phenotypes identifies molecular and behavioral consequences of MYT1L haploinsufficiency in male and female mice.

The transcription factor MYT1L supports proper neuronal differentiation and maturation during brain development. MYT1L haploinsufficiency results in a neurodevelopmental disorder characterized by intellectual disability, developmental delay, autism, behavioral disruptions, aggression, obesity and epilepsy. While MYT1L is expressed throughout the brain, how it supports proper neuronal function in distinct regions has not been assessed. Some features of MYT1L Neurodevelopmental Syndrome suggest disruption of hypothalamic function, such as obesity and endocrine issues, and previous research showed changes in hypothalamic neuropeptide expression following knockdown in zebrafish. Here, we leveraged our heterozygous Myt1l mutant, previously shown to recapitulate aspects of the human syndrome such as hyperactivity, social challenges, and obesity, to examine the impact of MYT1L loss on hypothalamic function. Examining the molecular profile of the MYT1L haploinsufficient hypothalamus revealed a similar scale of disruption to previously studied brain regions, yet with region-specific roles for MYT1L, including regulation of neuropeptide systems. Alterations in oxytocin and arginine vasopressin cell numbers were also found. Behaviors studied included maternal care, social group hierarchies, and aggression, all of which were unchanged. Feeding and metabolic markers were also largely unchanged in MYT1L haploinsufficient mice, yet an interaction was observed between diet and MYT1L genotype on weight gain. Our findings here suggest that gross endocrine function was not altered by MYT1L haploinsufficiency, and that key sex-specific behaviors related to proper hypothalamic function remain intact. Further study is needed to understand the functional impact of the altered hypothalamic molecular profile and changes in neuropeptide cell numbers that result from MYT1L haploinsufficiency.

animal behavior and cognition↗

bioRxiv2024

Lifespan in rodents with MYT1L heterozygous mutation

MYT1L syndrome is a newly recognized disorder characterized by intellectual disability, speech and motor delay, neuroendocrine disruptions, ADHD, and autism. In order to study this gene and its association with these phenotypes, our lab recently created a Myt1l heterozygous mutant mouse inspired by a clinically relevant mutation. This model recapitulates several of the physical and neurologic abnormalities seen in humans with MYT1L syndrome, such as weight gain, microcephaly, and behavioral disruptions. The majority of patients with this syndrome are young, and little is known about the impact of age on health and mortality in these patients. Using a Myt1l mutant mouse, we examined the impact of Myt1l mutation on body weights, lifespan, and histopathology findings of mice at the end of life. This cohort of heterozygous mice demonstrated increased body weight across the lifespan, however there was no significant difference in lifespan, apparent cause of death, or end of life histopathological findings between Myt1l heterozygous and wildtype mice. These findings suggest while Myt1l heterozygous mutation may influence overall brain development, it does not strongly impact other organ systems in the body over time.

genetics↗

bioRxiv2024

Chromosomal and gonadal sex have differing effects on social motivation in mice

Plain English SummaryAs our brain develops, many factors influence how we behave later in life. The brain forms differently in males and females, potentially leading to sex variation seen in many behaviors including sociability. In addition, conditions defined by differences in social behaviors, such as autism, are diagnosed more in males than females. However, researchers dont know exactly how distinct sex factors, such as hormones and sex chromosome genes, lead to different behaviors in males and females. In this study, we used mouse models and tests of mouse behavior to explore these differences. Results show that sex hormones primarily contributed to differences in social motivation between sexes. Yet when we repeated these same assays in a mouse model of genetic liability for a human neurodevelopmental syndrome, we found that sex chromosome genes rather than sex hormones played a larger role in the behavioral consequences of impaired neurodevelopment. These insights can inform future research on the biological mechanisms of social behavior in the context of genetic liability for neurodevelopmental disorders. HighlightsO_LIFour-core genotype mouse model crossed with MYT1L heterozygous mouse revealed independent effects of chromosomal and gonadal sex on social motivation. C_LIO_LIMyt1l haploinsufficiency was associated with increased activity in both males and females. C_LIO_LIWhile females are more active, contributions of chromosomes and gonadal hormones to this sex effect are environment dependent. C_LIO_LIPresence of ovaries was associated with increased measures of social seeking and orienting regardless of genotype. C_LIO_LIChromosomal sex interacted with MYT1L genotype, with increased social orienting and seeking specifically in XX MYT1L heterozygous mice. C_LI BackgroundSex differences in brain development are thought to lead to sex variation in social behavior. Sex differences are fundamentally driven by both gonadal (i.e., hormonal) and chromosomal sex, yet little is known about the independent effects of each on social behavior. Further, mouse models of the genetic liability for the neurodevelopmental disorder MYT1L Syndrome have shown sex specific deficits in social motivation. In this study, we aimed to determine if hormonal or chromosomal sex primarily mediate the sex differences seen in mouse social behavior, both at baseline and in the context of Myt1l haploinsufficiency. MethodsFour-core genotype (FCG) mice, which uncouple gonadal and chromosomal sex, were crossed with MYT1L heterozygous mice to create eight different groups with unique combinations of sex factors and MYT1L genotype. A total of 131 mice from all eight groups were assayed for activity and social behavior via the open field and social operant paradigms. Measures of social seeking and orienting were analyzed for main effects of chromosome, gonads, and their interactions with Myt1l mutation. ResultsThe FCGxMYT1L cross revealed independent effects of both gonadal and chromosomal sex on activity and social behavior. Specifically, the presence of ovaries, and by extension the presence of ovarian hormones, increased overall activity, social seeking, and social orienting regardless of genotype. In contrast, sex chromosomes affected social behavior mainly in the MYT1L heterozygous group, with XX sex karyotype when combined with MYT1L genotype contributing to increased social orienting and seeking. ConclusionsGonadal and chromosomal sex have independent mechanisms of driving increased social motivation in females. Additionally, sex chromosomes may interact with neurodevelopmental mutations to influence sex variation in atypical social behavior.

animal behavior and cognition↗

bioRxiv2024

A glial circadian gene expression atlas reveals cell type and disease-specific reprogramming in response to amyloid pathology or aging.

While circadian rhythm disruption may promote neurodegenerative disease, how aging and neurodegenerative pathology impact circadian gene expression patterns in different brain cell types is unknown. Here, we used translating ribosome affinity purification methods to define the circadian translatomes of astrocytes, microglia, and bulk cerebral cortex, in healthy mouse brain and in the settings of amyloid-beta plaque pathology or aging. Our data reveal that glial circadian translatomes are highly cell type-specific and exhibit profound, context-dependent reprogramming of rhythmic transcripts in response to amyloid pathology or aging. Transcripts involved in glial activation, immunometabolism, and proteostasis, as well as nearly half of all Alzheimer Disease (AD)-associated risk genes, displayed circadian oscillations, many of which were altered by pathology. Amyloid-related differential gene expression was also dependent on time of day. Thus, circadian rhythms in gene expression are cell- and context dependent and provide important insights into glial gene regulation in health, AD, and aging.

neuroscience↗

bioRxiv2024

MYT1L deficiency impairs excitatory neuron trajectory during cortical development

Mutations that reduce the function of MYT1L, a neuron-specific transcription factor, are associated with a syndromic neurodevelopmental disorder. Furthermore, MYT1L is routinely used as a proneural factor in fibroblast-to-neuron transdifferentiation. MYT1L has been hypothesized to play a role in the trajectory of neuronal specification and subtype specific maturation, but this hypothesis has not been directly tested, nor is it clear which neuron types are most impacted by MYT1L loss. In this study, we profiled 313,335 nuclei from the forebrains of wild-type and MYT1L-deficient mice at two developmental stages: E14 at the peak of neurogenesis and P21, when neurogenesis is complete, to examine the role of MYT1L levels in the trajectory of neuronal development. We found that MYT1L deficiency significantly disrupted the relative proportion of cortical excitatory neurons at E14 and P21. Significant changes in gene expression were largely concentrated in excitatory neurons, suggesting that transcriptional effects of MYT1L deficiency are largely due to disruption of neuronal maturation programs. Most effects on gene expression were cell autonomous and persistent through development. In addition, while MYT1L can both activate and repress gene expression, the repressive effects were most sensitive to haploinsufficiency, and thus more likely mediate MYT1L syndrome. These findings illuminate the intricate role of MYT1L in orchestrating gene expression dynamics during neuronal development, providing insights into the molecular underpinnings of MYT1L syndrome.

neuroscience↗

bioRxiv2024

Mice lacking Astn2 have ASD-like behaviors and altered cerebellar circuit properties

Astrotactin 2 (ASTN2) is a transmembrane neuronal protein highly expressed in the cerebellum that functions in receptor trafficking and modulates cerebellar Purkinje cell (PC) synaptic activity. We recently reported a family with a paternally inherited intragenic ASTN2 duplication with a range of neurodevelopmental disorders, including autism spectrum disorder (ASD), learning difficulties, and speech and language delay. To provide a genetic model for the role of the cerebellum in ASD-related behaviors and study the role of ASTN2 in cerebellar circuit function, we generated global and PC-specific conditional Astn2 knockout (KO and cKO, respectively) mouse lines. Astn2 KO mice exhibit strong ASD-related behavioral phenotypes, including a marked decrease in separation-induced pup ultrasonic vocalization calls, hyperactivity and repetitive behaviors, altered social behaviors, and impaired cerebellar-dependent eyeblink conditioning. Hyperactivity and repetitive behaviors were also prominent in Astn2 cKO animals. By Golgi staining, Astn2 KO PCs have region-specific changes in dendritic spine density and filopodia numbers. Proteomic analysis of Astn2 KO cerebellum reveals a marked upregulation of ASTN2 family member, ASTN1, a neuron-glial adhesion protein. Immunohistochemistry and electron microscopy demonstrates a significant increase in Bergmann glia volume in the molecular layer of Astn2 KO animals. Electrophysiological experiments indicate a reduced frequency of spontaneous excitatory postsynaptic currents (EPSCs), as well as increased amplitudes of both spontaneous EPSCs and inhibitory postsynaptic currents (IPSCs) in the Astn2 KO animals, suggesting that pre- and postsynaptic components of synaptic transmission are altered. Thus, ASTN2 regulates ASD-like behaviors and cerebellar circuit properties.

neuroscience↗

bioRxiv2023

Evaluation of gliovascular functions of Aqp4 readthrough isoforms

Aquaporin-4 (AQP4) is a water channel protein that links astrocytic endfeet to the blood-brain barrier (BBB) and regulates water and potassium homeostasis in the brain, as well as the glymphatic clearance of waste products that would otherwise potentiate neurological diseases. Recently, translational readthrough was shown to generate a C-terminally extended variant of AQP4, known as AQP4x, that preferentially localizes around the BBB through interaction with the scaffolding protein -syntrophin, and loss of AQP4x disrupts waste clearance from the brain. To investigate the function of AQP4x, we generated a novel mouse AQP4 line (AllX) to increase relative levels of the readthrough variant above the [~]15% of AQP4 in the brain of wildtype (WT) mice. We validated the line and assessed characteristics that are affected by the presence of AQP4x, including AQP4 and -syntrophin localization, integrity of the BBB, and neurovascular coupling. We compared AllXHom and AllXHet mice to wildtype, and to previously characterized AQP4 NoXHet and NoXHom mice, which cannot produce AQP4x. Increased dose of AQP4x enhanced perivascular localization of - syntrophin and AQP4, while total protein expression of the two were unchanged. However, at 100% readthrough, AQP4x localization and formation of higher-order complexes was disrupted. Electron microscopy showed that overall blood vessel morphology was unchanged except for increased endothelial cell vesicles in NoXHom mice, which may correspond to a leakier BBB or altered efflux that was identified in NoX mice using MRI. These data demonstrate that AQP4x plays a small but measurable role in maintaining BBB integrity as well as recruiting structural and functional support proteins to the blood vessel. This also establishes a new set of genetic tools for quantitatively modulating AQP4x levels. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/549379v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@18f8f32org.highwire.dtl.DTLVardef@25afeorg.highwire.dtl.DTLVardef@a3e0e1org.highwire.dtl.DTLVardef@100f2e1_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2023

Calling Cards: a customizable platform to longitudinally record protein-DNA interactions over time in cells and tissues

Calling Cards is a platform technology to record a cumulative history of transient protein-DNA interactions in the genome of genetically targeted cell types. The record of these interactions is recovered by next generation sequencing. Compared to other genomic assays, whose readout provides a snapshot at the time of harvest, Calling Cards enables correlation of historical molecular states to eventual outcomes or phenotypes. To achieve this, Calling Cards uses the piggyBac transposase to insert self-reporting transposon (SRT) "Calling Cards" into the genome, leaving permanent marks at interaction sites. Calling Cards can be deployed in a variety of in vitro and in vivo biological systems to study gene regulatory networks involved in development, aging, and disease. Out of the box, it assesses enhancer usage but can be adapted to profile specific transcription factor binding with custom transcription factor (TF)-piggyBac fusion proteins. The Calling Cards workflow has five main stages: delivery of Calling Card reagents, sample preparation, library preparation, sequencing, and data analysis. Here, we first present a comprehensive guide for experimental design, reagent selection, and optional customization of the platform to study additional TFs. Then, we provide an updated protocol for the five steps, using reagents that improve throughput and decrease costs, including an overview of a newly deployed computational pipeline. This protocol is designed for users with basic molecular biology experience to process samples into sequencing libraries in 1-2 days. Familiarity with bioinformatic analysis and command line tools is required to set up the pipeline in a high-performance computing environment and to conduct downstream analyses. Basic Protocol 1: Preparation and delivery of Calling Cards reagents Basic Protocol 2: Sample preparation Basic Protocol 3: Sequencing library preparation Basic Protocol 4: Library pooling and sequencing Basic Protocol 5: Data analysis

genomics↗

bioRxiv2023

Visual deprivation during mouse critical period reorganizes network-level functional connectivity

A classic example of experience-dependent plasticity is ocular dominance (OD) shift, in which the responsiveness of neurons in the visual cortex is profoundly altered following monocular deprivation (MD). It has been postulated that OD shifts also modify global neural networks, but such effects have never been demonstrated. Here, we used longitudinal wide-field optical calcium imaging to measure resting-state functional connectivity during acute (3-day) MD in mice. First, delta GCaMP6 power in the deprived visual cortex decreased, suggesting that excitatory activity was reduced in the region. In parallel, interhemispheric visual homotopic functional connectivity was rapidly reduced by the disruption of visual drive through MD and was sustained significantly below baseline state. This reduction of visual homotopic connectivity was accompanied by a reduction in parietal and motor homotopic connectivity. Finally, we observed enhanced internetwork connectivity between visual and parietal cortex that peaked at MD2. Together, these findings support the hypothesis that early MD induces dynamic reorganization of disparate functional networks including association cortices. Significance StatementMonocular deprivation during the visual critical period triggers several plasticity mechanisms that collaborate to shift the excitability of neurons in the visual cortex. However, little is known about the impacts of MD on cortex-wide functional networks. Here, we measured cortical functional connectivity during short-term critical period MD. We demonstrate that critical period MD has immediate effects on functional networks beyond the visual cortex, and identify regions of substantial functional connectivity reorganization in response to MD.

neuroscience↗

bioRxiv2023

Tyrosinase-induced neuromelanin accumulation triggers rapid dysregulation and degeneration of the mouse locus coeruleus

The locus coeruleus (LC), the major source of norepinephrine (NE) in the brain, is among the first sites of pathology in both Alzheimers disease (AD) and Parkinsons disease (PD), and it undergoes catastrophic degeneration later in both disorders. Dysregulation of the LC is thought to contribute to early behavioral symptoms of AD and PD such as anxiety and sleep disturbances, while frank LC loss promotes cognitive decline. However, the mechanisms responsible for this selective vulnerability are unknown. It has been suggested that neuromelanin (NM) pigment contributes to LC susceptibility, but a causal relationship has been difficult to test because rodents do not naturally produce NM. Here, we report that viral-mediated expression of human tyrosinase-induced pigmentation in male and female mouse LC neurons recapitulated key macroscopic and ultrastructural features of natural primate NM. One week of NM accumulation resulted in LC neuron hyperactivity, reduced tissue NE levels, transcriptional changes, and anxiety-like behavior. By 6 weeks, NM accumulation was associated with severe cell-autonomous LC neuron degeneration, neuroinflammation, and microglial engulfment of the pigment granules, while the anxiety-like behavior abated. These phenotypes are reminiscent of LC dysfunction and cell death in AD and PD, validating this model for studying the consequences of NM accumulation in the LC as it relates to neurodegenerative diseases. Significance StatementAlzheimers disease (AD) and Parkinsons disease (PD) are the most common neurodegenerative diseases worldwide. Because therapies that cure or prevent their progression are lacking, research is focused on the identifying the earliest signs of disease as targets for diagnosis and treatment. The locus coeruleus (LC), the main source of norepinephrine (NE) in the brain, is one of the first brain regions affected in both AD and PD. Early on, LC dysregulation promotes behavioral symptoms of AD and PD, while its subsequent degeneration accelerates disease progression. Here we identify neuromelanin (NM) pigment as an LC vulnerability factor that induces neuronal hyperactivity followed by cell death. Approaches that mitigate NM accumulation and toxicity may target the earliest phases of neurodegenerative disease.

neuroscience↗

bioRxiv2023

Distinct disease mutations in DNMT3A result in a spectrum of behavioral, epigenetic, and transcriptional deficits

Phenotypic heterogeneity is a common feature of monogenic neurodevelopmental disorders that can arise from differential severity of missense variants underlying disease, but how distinct alleles impact molecular mechanisms to drive variable disease presentation is not well understood. Here, we investigate missense mutations in the DNA methyltransferase DNMT3A associated with variable overgrowth, intellectual disability, and autism, to uncover molecular correlates of phenotypic heterogeneity in neurodevelopmental disease. We generate a DNMT3A P900L/+ mouse model mimicking a disease mutation with mild-to-moderate severity and compare phenotypic and epigenomic effects with a severe R878H mutation. We show that the P900L mutation leads to disease-relevant overgrowth, obesity, and social deficits shared across DNMT3A disorder models, while the R878H mutation causes more extensive epigenomic disruption leading to differential dysregulation of enhancers elements. We identify distinct gene sets disrupted in each mutant which may contribute to mild or severe disease, and detect shared transcriptomic disruption that likely drives common phenotypes across affected individuals. Finally, we demonstrate that core gene dysregulation detected in DNMT3A mutant mice overlaps effects in other developmental disorder models, highlighting the importance of DNMT3A-deposited methylation in neurodevelopment. Together, these findings define central drivers of DNMT3A disorders and illustrate how variable disruption of transcriptional mechanisms can drive the spectrum of phenotypes in neurodevelopmental disease.

neuroscience↗

bioRxiv2023

Extensive characterization of a Williams Syndrome murine model shows Gtf2ird1-mediated rescue of select sensorimotor tasks, but no effect on enhanced social behavior

Williams Syndrome is a rare neurodevelopmental disorder exhibiting cognitive and behavioral abnormalities, including increased social motivation, risk of anxiety and specific phobias along with perturbed motor function. Williams Syndrome is caused by a microdeletion of 26-28 genes on chromosome 7, including GTF2IRD1, which encodes a transcription factor suggested to play a role in the behavioral profile of Williams Syndrome. Duplications of the full region also lead to frequent autism diagnosis, social phobias, and language delay. Thus, genes in the region appear to regulate social motivation in a dose-sensitive manner. A Complete Deletion mouse, heterozygously eliminating the syntenic Williams Syndrome region, has been deeply characterized for cardiac phenotypes, but direct measures of social motivation have not been assessed. Furthermore, the role of Gtf2ird1 in these behaviors has not been addressed in a relevant genetic context. Here, we have generated a mouse overexpressing Gtf2ird1, which can be used both to model duplication of this gene alone and to rescue Gtf2ird1 expression in the Complete Deletion mice. Using a comprehensive behavioral pipeline and direct measures of social motivation, we provide evidence that the Williams Syndrome Critical Region regulates social motivation along with motor and anxiety phenotypes, but that Gtf2ird1 complementation is not sufficient to rescue most of these traits, and duplication does not decrease social motivation. However, Gtf2ird1 complementation does rescue light-aversive behavior and performance on select sensorimotor tasks, perhaps indicating a role for this gene in sensory processing or integration.

animal behavior and cognition↗