CRISPR interference-based specific and efficient gene inactivation in the brain

Zheng, Yi; Shen, Wei; Zhang, Jian; Yang, Bo; Yaonan, Liu; Qi, Huihui; Yu, Xia; Lu, Shaoping; Chen, Yun; Xu, Yuzhou; Li, Yun; Gage, Fred H.; Mi, Shuangli; Yao, Jun

doi:10.1038/s41593-018-0077-5

Cited by 139 publications

(103 citation statements)

References 51 publications

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“…Epigenetic editing in brain has seen major advances in its application to gene activation and silencing (Lee et al, 2018;Liu et al, 2018;Zheng et al, 2018). For example, targeted dCas9-Tet1 to CGG repeats in gene fragile X mental retardation 1 (FMR1) is sufficient to reverse the hypermethylation at this locus and sustain expression of fragile X mental retardation protein (FMRP) in edited neurons .…”

Section: Discussionmentioning

confidence: 99%

Chromatin-mediated alternative splicing regulates cocaine reward behavior

Lombroso

Carpenter

et al. 2019

Preprint

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Alternative splicing is a key mechanism for neuronal gene regulation, and is grossly altered in mouse brain reward regions following investigator-administered cocaine. It is well established that cocaine epigenetically regulates transcription, yet mechanism(s) by which cocaine-induced epigenetic modifications regulate alternative splicing is largely unexplored. Our group and others have previously identified the histone modification, H3K36me3, as a putative splicing regulator. However, it has not yet been possible to establish the direct causal relevance of this modification to alternative splicing in brain or any other context. We found that mouse cocaine self-administration caused widespread alternative splicing, concomitant with enrichment of H3K36me3 at splice junctions. Differentially spliced genes were enriched in the motif for splice factor, Srsf11, which was both differentially spliced and enriched in H3K36me3. Epigenetic editing led us to conclude that H3K36me3 functions directly in alternative splicing of Srsf11, and that Set2 mediated H3K36me3 bidirectionally regulates cocaine intake.

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Section: Discussionmentioning

confidence: 99%

Chromatin-mediated alternative splicing regulates cocaine reward behavior

Lombroso

Carpenter

et al. 2019

Preprint

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“…Cas9 has already been delivered with AAV in rodents and has been shown to induce a mild cellular response, but this has not been reported in the brain (Chew et al, 2016). CRISPRa is, in principle, less likely to have deleterious off-target effects than CRISPR gene editing because it does not cleave DNA (La Russa and Qi, 2015;Zheng et al, 2018), but further research is necessary.…”

Section: Discussionmentioning

confidence: 99%

In vivo CRISPRa decreases seizures and rescues cognitive deficits in a rodent model of epilepsy

Colasante

Qiu

Massimino

et al. 2018

Preprint

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Epilepsy is a major health burden, calling for new mechanistic and therapeutic insights.CRISPR-mediated gene editing shows promise to cure genetic pathologies, although hitherto it has mostly been applied ex-vivo. Its translational potential for treating non-genetic pathologies is still unexplored. Furthermore, neurological diseases represent an important challenge for the application of CRISPR, because of the need in many cases to manipulate gene function of neurons in situ. A variant of CRISPR, CRISPRa, offers the possibility to modulate the expression of endogenous genes by directly targeting their promoters. We asked if this strategy can effectively treat acquired focal epilepsy, focusing on ion channels because their manipulation is known be effective in changing network hyperactivity and hypersynchronisation. We applied a doxycycline-inducible CRISPRa technology to increase the expression of the potassium channel gene Kcna1 (encoding Kv1.1) in mouse hippocampal excitatory neurons. CRISPRa-mediated Kv1.1 upregulation led to a substantial decrease in neuronal excitability. Continuous video-EEG telemetry showed that AAV9-mediated delivery of CRISPRa, upon doxycycline administration, decreased spontaneous generalized tonicclonic seizures in a model of temporal lobe epilepsy, and rescued cognitive impairment and transcriptomic alterations associated with chronic epilepsy. The focal treatment minimizes concerns about off-target effects in other organs and brain areas. This study provides the proof of principle for a translational CRISPR-based approach to treat neurological diseases characterized by abnormal circuit excitability.

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“…Egr3 Egr2 log2(Fold change) Target/LacZ log2(Fold change) Dopaplex/LacZ enabled rapid and multiplexable transcriptional control in the mammalian central nervous system, providing an avenue to investigate how gene programs regulate normal and maladaptive brain states (35,(42)(43)(44). To simultaneously activate multiple genes in this DA-induced gene expression profile, we harnessed a neuron-optimized CRISPR activation (CRISPRa) system ( Fig.…”

Section: Controlmentioning

confidence: 99%

A dopamine-induced gene expression signature regulates neuronal function and cocaine response

Savell

Zipperly

Tuscher

et al. 2019

Preprint

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Drug addiction is a worldwide health problem, with overdose rates of both psychostimulants and opioids currently on the rise in many developed countries. Drugs of abuse elevate dopamine levels in the nucleus accumbens (NAc) and alter transcriptional programs believed to promote long-lasting synaptic and behavioral adaptations. However, even with well-studied drugs such as cocaine, druginduced transcriptional responses remain poorly understood due to the cellular heterogeneity of the NAc and complex drug actions via multiple neurotransmitter systems. Here, we leveraged highthroughput single-nucleus RNA-sequencing to create a comprehensive molecular atlas of cell subtypes in the NAc, defining both sex-specific and cell type-specific responses to acute cocaine experience in a rat model system. Using this transcriptional map, we identified specific neuronal subpopulations that are activated by cocaine, and defined an immediate early gene expression program that is upregulated following cocaine experience in vivo and dopamine (DA) receptor activation in vitro. To characterize the neuronal response to this DA-mediated gene expression signature, we engineered a large-scale CRISPR/dCas9 activation strategy to recreate this program. Multiplexed induction of this gene program initiated a secondary synapse-centric transcriptional profile, altered striatal physiology in vitro, and enhanced cocaine sensitization in vivo. Taken together, these results define the genome-wide transcriptional response to cocaine with cellular precision, and demonstrate that drug-responsive gene programs are sufficient to initiate both physiological and behavioral adaptations to drugs of abuse.NEARLY 5 MILLION Americans reported cocaine use in 2017, and recent increases in cocaine-related drug overdoses present significant public health challenges (1). A hallmark trait of drugs of abuse is the acute elevation of dopamine (DA) in the nucleus accumbens (NAc), a central integrator of the reward circuit (2-4). Abused drugs produce increases in DA that are greater in both concentration and duration than natural rewards (2,5,6), and this signaling is hypothesized to underlie maladaptive reinforcement after repeated drug use (7). Exposure to drugs of abuse results in significant transcriptional and epigenetic reorganization in the NAc (8-13), initiating synaptic and behavioral plasticity associated with the transition to drug addiction (7,14,15). However, even with well-studied drugs such as cocaine, drug-induced transcriptional responses remain poorly understood. This is in part due to the cellular heterogeneity of the NAc, which is a diverse structure containing multiple neuronal and non-neuronal subpopulations and complex neuronal circuitry. Additionally, many drugs of abuse engage multiple neurotransmitter systems in the NAc (16-22), and the specific contributions of DA-dependent transcriptional programs to neuronal physiology and behavior is not clear. Further, although drug experience leads to large scale transcriptional changes in the NAc, previou...

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CRISPR interference-based specific and efficient gene inactivation in the brain

Cited by 139 publications

References 51 publications

Chromatin-mediated alternative splicing regulates cocaine reward behavior

Chromatin-mediated alternative splicing regulates cocaine reward behavior

In vivo CRISPRa decreases seizures and rescues cognitive deficits in a rodent model of epilepsy

A dopamine-induced gene expression signature regulates neuronal function and cocaine response

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