Publications by authors named "Carl Ernst"

64 Publications

Lesch-Nyhan disease causes impaired energy metabolism and reduced developmental potential in midbrain dopaminergic cells.

Stem Cell Reports 2021 Jul 1;16(7):1749-1762. Epub 2021 Jul 1.

Psychiatric Genetics Group, McGill University, Montreal, QC, Canada; Department of Psychiatry, McGill University and Douglas Hospital Research Institute, 6875 LaSalle Boulevard, Frank Common Building, Room 2101.2, Montreal, QC H4H 1R3, Canada. Electronic address:

Mutations in HPRT1, a gene encoding a rate-limiting enzyme for purine salvage, cause Lesch-Nyhan disease which is characterized by self-injury and motor impairments. We leveraged stem cell and genetic engineering technologies to model the disease in isogenic and patient-derived forebrain and midbrain cell types. Dopaminergic progenitor cells deficient in HPRT showed decreased intensity of all developmental cell-fate markers measured. Metabolic analyses revealed significant loss of all purine derivatives, except hypoxanthine, and impaired glycolysis and oxidative phosphorylation. real-time glucose tracing demonstrated increased shunting to the pentose phosphate pathway for de novo purine synthesis at the expense of ATP production. Purine depletion in dopaminergic progenitor cells resulted in loss of RHEB, impairing mTORC1 activation. These data demonstrate dopaminergic-specific effects of purine salvage deficiency and unexpectedly reveal that dopaminergic progenitor cells are programmed to a high-energy state prior to higher energy demands of terminally differentiated cells.
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http://dx.doi.org/10.1016/j.stemcr.2021.06.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8282463PMC
July 2021

Non-CG methylation and multiple histone profiles associate child abuse with immune and small GTPase dysregulation.

Nat Commun 2021 02 18;12(1):1132. Epub 2021 Feb 18.

McGill Group for Suicide Studies, Douglas Mental Health University Institute, McGill University, Montréal, Canada.

Early-life adversity (ELA) is a major predictor of psychopathology, and is thought to increase lifetime risk by epigenetically regulating the genome. Here, focusing on the lateral amygdala, a major brain site for emotional homeostasis, we describe molecular cross-talk among multiple mechanisms of genomic regulation, including 6 histone marks and DNA methylation, and the transcriptome, in subjects with a history of ELA and controls. In the healthy brain tissue, we first uncover interactions between different histone marks and non-CG methylation in the CAC context. Additionally, we find that ELA associates with methylomic changes that are as frequent in the CAC as in the canonical CG context, while these two forms of plasticity occur in sharply distinct genomic regions, features, and chromatin states. Combining these multiple data indicates that immune-related and small GTPase signaling pathways are most consistently impaired in the amygdala of ELA individuals. Overall, this work provides insights into genomic brain regulation as a function of early-life experience.
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http://dx.doi.org/10.1038/s41467-021-21365-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7892573PMC
February 2021

Cocaine-related DNA methylation in caudate neurons alters 3D chromatin structure of the IRXA gene cluster.

Mol Psychiatry 2021 Jul 12;26(7):3134-3151. Epub 2020 Oct 12.

McGill Group for Suicide Studies, Douglas Hospital Research Center, Montreal, QC, Canada.

Epigenetic mechanisms, like those involving DNA methylation, are thought to mediate the relationship between chronic cocaine dependence and molecular changes in addiction-related neurocircuitry, but have been understudied in human brain. We initially used reduced representation bisulfite sequencing (RRBS) to generate a methylome-wide profile of cocaine dependence in human post-mortem caudate tissue. We focused on the Iroquois Homeobox A (IRXA) gene cluster, where hypomethylation in exon 3 of IRX2 in neuronal nuclei was associated with cocaine dependence. We replicated this finding in an independent cohort and found similar results in the dorsal striatum from cocaine self-administering mice. Using epigenome editing and 3C assays, we demonstrated a causal relationship between methylation within the IRX2 gene body, CTCF protein binding, three-dimensional (3D) chromatin interaction, and gene expression. Together, these findings suggest that cocaine-related hypomethylation of IRX2 contributes to the development and maintenance of cocaine dependence through alterations in 3D chromatin structure in the caudate nucleus.
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http://dx.doi.org/10.1038/s41380-020-00909-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8039060PMC
July 2021

Stimulation of L-type calcium channels increases tyrosine hydroxylase and dopamine in ventral midbrain cells induced from somatic cells.

Stem Cells Transl Med 2020 Jun 10;9(6):697-712. Epub 2020 Mar 10.

Psychiatric Genetics Group, McGill University, Montreal, Quebec, Canada.

Making high-quality dopamine (DA)-producing cells for basic biological or small molecule screening studies is critical for the development of novel therapeutics for disorders of the ventral midbrain. Currently, many ventral midbrain assays have low signal-to-noise ratio due to low levels of cellular DA and the rate-limiting enzyme of DA synthesis, tyrosine hydroxylase (TH), hampering discovery efforts. Using intensively characterized ventral midbrain cells derived from human skin, which demonstrate calcium pacemaking activity and classical electrophysiological properties, we show that an L-type calcium agonist can significantly increase TH protein levels and DA content and release. Live calcium imaging suggests that it is the immediate influx of calcium occurring simultaneously in all cells that drives this effect. Genome-wide expression profiling suggests that L-type calcium channel stimulation has a significant effect on specific genes related to DA synthesis and affects expression of L-type calcium receptor subunits from the CACNA1 and CACNA2D families. Together, our findings provide an advance in the ability to increase DA and TH levels to improve the accuracy of disease modeling and small molecule screening for disorders of the ventral midbrain, including Parkinson's disease.
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http://dx.doi.org/10.1002/sctm.18-0180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7214648PMC
June 2020

A roadmap for neurodevelopmental disease modeling for non-stem cell biologists.

Authors:
Carl Ernst

Stem Cells Transl Med 2020 May 13;9(5):567-574. Epub 2020 Feb 13.

Department of Human Genetics, McGill University and Douglas Hospital Research Institute, Montreal, Quebec, Canada.

Stem and derivative cells induced from somatic tissues are a critical tool for disease modeling but significant technical hurdles hamper their use. The purpose of this review is to provide an overview of pitfalls and mitigation strategies for the nonstem cell biologist using induced pluripotent stem cells and investigating neurodevelopmental disorders. What sample sizes are reasonable? What derivation and purification protocols should be used to make human neurons? In what way should gene editing technologies be used to support discoveries? What kinds of preclinical studies are the most feasible? It is hoped that this roadmap will provide the necessary details for experimental planning and execution for those less familiar in the area of stem cell disease modeling. High-quality human preclinical models will allow for the discovery of molecular and cellular phenotypes specific to different neurodevelopmental disorders, and may provide the assays to advance translational medicine for unmet medical needs.
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http://dx.doi.org/10.1002/sctm.19-0344DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7180294PMC
May 2020

Human iPSC-derived Down syndrome astrocytes display genome-wide perturbations in gene expression, an altered adhesion profile, and increased cellular dynamics.

Hum Mol Genet 2020 03;29(5):785-802

Department of Neurology & Neurosurgery, Brain Repair and Integrative Neuroscience Program, Centre for Research in Neuroscience, Research Institute of the McGill University Health Centre, Montreal, QC H3G 1A4, Canada.

Down syndrome (DS), caused by the triplication of human chromosome 21, leads to significant alterations in brain development and is a major genetic cause of intellectual disability. While much is known about changes to neurons in DS, the effects of trisomy 21 on non-neuronal cells such as astrocytes are poorly understood. Astrocytes are critical for brain development and function, and their alteration may contribute to DS pathophysiology. To better understand the impact of trisomy 21 on astrocytes, we performed RNA-sequencing on astrocytes from newly produced DS human induced pluripotent stem cells (hiPSCs). While chromosome 21 genes were upregulated in DS astrocytes, we found consistent up- and down-regulation of genes across the genome with a strong dysregulation of neurodevelopmental, cell adhesion and extracellular matrix molecules. ATAC (assay for transposase-accessible chromatin)-seq also revealed a global alteration in chromatin state in DS astrocytes, showing modified chromatin accessibility at promoters of cell adhesion and extracellular matrix genes. Along with these transcriptomic and epigenomic changes, DS astrocytes displayed perturbations in cell size and cell spreading as well as modifications to cell-cell and cell-substrate recognition/adhesion, and increases in cellular motility and dynamics. Thus, triplication of chromosome 21 is associated with genome-wide transcriptional, epigenomic and functional alterations in astrocytes that may contribute to altered brain development and function in DS.
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http://dx.doi.org/10.1093/hmg/ddaa003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7104679PMC
March 2020

Dose in Brain Development.

Front Pediatr 2019 22;7:482. Epub 2019 Nov 22.

Department of Human Genetics, McGill University, Montreal, QC, Canada.

Brain development is a highly regulated process that involves the precise spatio-temporal activation of cell signaling cues. Transcription factors play an integral role in this process by relaying information from external signaling cues to the genome. The transcription factor Forkhead box G1 () is expressed in the developing nervous system with a critical role in forebrain development. Altered dosage of due to deletions, duplications, or functional gain- or loss-of-function mutations, leads to a complex array of cellular effects with important consequences for human disease including neurodevelopmental disorders. Here, we review studies in multiple species and cell models where dose is altered. We argue against a linear, symmetrical relationship between dosage states, although levels at the right time and place need to be carefully regulated. Neurodevelopmental disease states caused by mutations in may therefore be regulated through different mechanisms.
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http://dx.doi.org/10.3389/fped.2019.00482DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6882862PMC
November 2019

Mutations in ACTL6B Cause Neurodevelopmental Deficits and Epilepsy and Lead to Loss of Dendrites in Human Neurons.

Am J Hum Genet 2019 05 25;104(5):815-834. Epub 2019 Apr 25.

Department of Molecular Neuroscience, UCL Institute of Neurology, Queen Square, WC1N 3BG London, UK.

We identified individuals with variations in ACTL6B, a component of the chromatin remodeling machinery including the BAF complex. Ten individuals harbored bi-allelic mutations and presented with global developmental delay, epileptic encephalopathy, and spasticity, and ten individuals with de novo heterozygous mutations displayed intellectual disability, ambulation deficits, severe language impairment, hypotonia, Rett-like stereotypies, and minor facial dysmorphisms (wide mouth, diastema, bulbous nose). Nine of these ten unrelated individuals had the identical de novo c.1027G>A (p.Gly343Arg) mutation. Human-derived neurons were generated that recaptured ACTL6B expression patterns in development from progenitor cell to post-mitotic neuron, validating the use of this model. Engineered knock-out of ACTL6B in wild-type human neurons resulted in profound deficits in dendrite development, a result recapitulated in two individuals with different bi-allelic mutations, and reversed on clonal genetic repair or exogenous expression of ACTL6B. Whole-transcriptome analyses and whole-genomic profiling of the BAF complex in wild-type and bi-allelic mutant ACTL6B neural progenitor cells and neurons revealed increased genomic binding of the BAF complex in ACTL6B mutants, with corresponding transcriptional changes in several genes including TPPP and FSCN1, suggesting that altered regulation of some cytoskeletal genes contribute to altered dendrite development. Assessment of bi-alleic and heterozygous ACTL6B mutations on an ACTL6B knock-out human background demonstrated that bi-allelic mutations mimic engineered deletion deficits while heterozygous mutations do not, suggesting that the former are loss of function and the latter are gain of function. These results reveal a role for ACTL6B in neurodevelopment and implicate another component of chromatin remodeling machinery in brain disease.
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http://dx.doi.org/10.1016/j.ajhg.2019.03.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6507050PMC
May 2019

Differentiation of Human Induced Pluripotent Stem Cells (iPSCs) into an Effective Model of Forebrain Neural Progenitor Cells and Mature Neurons.

Bio Protoc 2019 Mar 5;9(5):e3188. Epub 2019 Mar 5.

Psychiatric Genetics Group, McGill University and Douglas Hospital Research Institute, Department of Psychiatry, Verdun, Montreal, QC H4H 1R3, Canada.

Induced Pluripotent Stem Cells (iPSCs) are pluripotent stem cells that can be generated from somatic cells, and provide a way to model the development of neural tissues . One particularly interesting application of iPSCs is the development of neurons analogous to those found in the human forebrain. Forebrain neurons play a central role in cognition and sensory processing, and deficits in forebrain neuronal activity contributes to a host of conditions, including epilepsy, Alzheimer's disease, and schizophrenia. Here, we present our protocol for differentiating iPSCs into forebrain neural progenitor cells (NPCs) and neurons, whereby neural rosettes are generated from stem cells without dissociation and NPCs purified from rosettes based on their adhesion, resulting in a more rapid generation of pure NPC cultures. Neural progenitor cells can be maintained as long-term cultures, or differentiated into forebrain neurons. This protocol provides a simplified and fast methodology of generating forebrain NPCs and neurons, and enables researchers to generate effective models to study forebrain disease and neurodevelopment. This protocol can also be easily adapted to generate other neural lineages.
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http://dx.doi.org/10.21769/BioProtoc.3188DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7854068PMC
March 2019

Correction: A new microdeletion syndrome involving TBC1D24, ATP6V0C, and PDPK1 causes epilepsy, microcephaly, and developmental delay.

Genet Med 2019 Sep;21(9):2159-2160

Department of Pediatrics, Université de Montréal, Montreal, QC, Canada.

The original version of this Article contained an error in the spelling of the author Siddharth Banka, which was incorrectly given as Siddhart Banka. This has now been corrected in both the PDF and HTML versions of the Article.
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http://dx.doi.org/10.1038/s41436-018-0413-xDOI Listing
September 2019

Identification of a de novo case of -related Ehlers-Danlos syndrome in an infant in the West Indies leading to improved targeted clinical care.

Clin Case Rep 2018 Nov 15;6(11):2256-2261. Epub 2018 Oct 15.

Department of Biochemistry St. George's University St. George's Grenada.

A 1-year-old girl from an underserved community presented with irritability, pain, and delayed motor skills. Our genetics outreach program facilitated the diagnosis of Ehlers-Danlos syndrome masquerading as developmental delay after noting hyperextensible skin. Diagnosis for this family allows for state-of-the-art cardiac monitoring and appropriate symptomatic treatment for this rare disease.
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http://dx.doi.org/10.1002/ccr3.1873DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6230631PMC
November 2018

A Dual Noradrenergic Mechanism for the Relief of Neuropathic Allodynia by the Antidepressant Drugs Duloxetine and Amitriptyline.

J Neurosci 2018 11 24;38(46):9934-9954. Epub 2018 Sep 24.

Centre National de la Recherche Scientifique, Université de Strasbourg, Institut des Neurosciences Cellulaires et Intégratives, 67000 Strasbourg, France,

In addition to treating depression, antidepressant drugs are also a first-line treatment for neuropathic pain, which is pain secondary to lesion or pathology of the nervous system. Despite the widespread use of these drugs, the mechanism underlying their therapeutic action in this pain context remains partly elusive. The present study combined data collected in male and female mice from a model of neuropathic pain and data from the clinical setting to understand how antidepressant drugs act. We show two distinct mechanisms by which the selective inhibitor of serotonin and noradrenaline reuptake duloxetine and the tricyclic antidepressant amitriptyline relieve neuropathic allodynia. One of these mechanisms is acute, central, and requires descending noradrenergic inhibitory controls and α adrenoceptors, as well as the mu and delta opioid receptors. The second mechanism is delayed, peripheral, and requires noradrenaline from peripheral sympathetic endings and β adrenoceptors, as well as the delta opioid receptors. We then conducted a transcriptomic analysis in dorsal root ganglia, which suggested that the peripheral component of duloxetine action involves the inhibition of neuroimmune mechanisms accompanying nerve injury, including the downregulation of the TNF-α-NF-κB signaling pathway. Accordingly, immunotherapies against either TNF-α or Toll-like receptor 2 (TLR2) provided allodynia relief. We also compared duloxetine plasma levels in the animal model and in patients and we observed that patients' drug concentrations were compatible with those measured in animals under chronic treatment involving the peripheral mechanism. Our study highlights a peripheral neuroimmune component of antidepressant drugs that is relevant to their delayed therapeutic action against neuropathic pain. In addition to treating depression, antidepressant drugs are also a first-line treatment for neuropathic pain, which is pain secondary to lesion or pathology of the nervous system. However, the mechanism by which antidepressant drugs can relieve neuropathic pain remained in part elusive. Indeed, preclinical studies led to contradictions concerning the anatomical and molecular substrates of this action. In the present work, we overcame these apparent contradictions by highlighting the existence of two independent mechanisms. One is rapid and centrally mediated by descending controls from the brain to the spinal cord and the other is delayed, peripheral, and relies on the anti-neuroimmune action of chronic antidepressant treatment.
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http://dx.doi.org/10.1523/JNEUROSCI.1004-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6596240PMC
November 2018

A new microdeletion syndrome involving TBC1D24, ATP6V0C, and PDPK1 causes epilepsy, microcephaly, and developmental delay.

Genet Med 2019 05 24;21(5):1058-1064. Epub 2018 Sep 24.

Department of Pediatrics, Université de Montréal, Montreal, QC, Canada.

Purpose: Contiguous gene deletions are known to cause several neurodevelopmental syndromes, many of which are caused by recurrent events on chromosome 16. However, chromosomal microarray studies (CMA) still yield copy-number variants (CNVs) of unknown clinical significance. We sought to characterize eight individuals with overlapping 205-kb to 504-kb 16p13.3 microdeletions that are distinct from previously published deletion syndromes.

Methods: Clinical information on the patients and bioinformatic scores for the deleted genes were analyzed.

Results: All individuals in our cohort displayed developmental delay, intellectual disability, and various forms of seizures. Six individuals were microcephalic and two had strabismus. The deletion was absent in all 13 parents who were available for testing. The area of overlap encompasses seven genes including TBC1D24, ATP6V0C, and PDPK1 (also known as PDK1). Bi-allelic TBC1D24 pathogenic variants are known to cause nonsyndromic deafness, epileptic disorders, or DOORS syndrome (deafness, onychodystrophy, osteodystrophy, mental retardation, seizures). Sanger sequencing of the nondeleted TBC1D24 allele did not yield any additional pathogenic variants.

Conclusions: We propose that 16p13.3 microdeletions resulting in simultaneous haploinsufficiencies of TBC1D24, ATP6V0C, and PDPK1 cause a novel rare contiguous gene deletion syndrome of microcephaly, developmental delay, intellectual disability, and epilepsy.
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http://dx.doi.org/10.1038/s41436-018-0290-3DOI Listing
May 2019

Umap and Bismap: quantifying genome and methylome mappability.

Nucleic Acids Res 2018 11;46(20):e120

Princess Margaret Cancer Centre, M5G 1L7, Toronto, ON, Canada.

Short-read sequencing enables assessment of genetic and biochemical traits of individual genomic regions, such as the location of genetic variation, protein binding and chemical modifications. Every region in a genome assembly has a property called 'mappability', which measures the extent to which it can be uniquely mapped by sequence reads. In regions of lower mappability, estimates of genomic and epigenomic characteristics from sequencing assays are less reliable. These regions have increased susceptibility to spurious mapping from reads from other regions of the genome with sequencing errors or unexpected genetic variation. Bisulfite sequencing approaches used to identify DNA methylation exacerbate these problems by introducing large numbers of reads that map to multiple regions. Both to correct assumptions of uniformity in downstream analysis and to identify regions where the analysis is less reliable, it is necessary to know the mappability of both ordinary and bisulfite-converted genomes. We introduce the Umap software for identifying uniquely mappable regions of any genome. Its Bismap extension identifies mappability of the bisulfite-converted genome. A Umap and Bismap track hub for human genome assemblies GRCh37/hg19 and GRCh38/hg38, and mouse assemblies GRCm37/mm9 and GRCm38/mm10 is available at https://bismap.hoffmanlab.org for use with genome browsers.
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http://dx.doi.org/10.1093/nar/gky677DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6237805PMC
November 2018

Disruption of GRIN2B Impairs Differentiation in Human Neurons.

Stem Cell Reports 2018 07 21;11(1):183-196. Epub 2018 Jun 21.

McGill University and Douglas Hospital Research Institute, Department of Psychiatry, 6875 LaSalle Boulevard, Frank Common Building, Room 2101.2, Verdun, Montreal, QC H4H 1R3, Canada. Electronic address:

Heterozygous loss-of-function mutations in GRIN2B, a subunit of the NMDA receptor, cause intellectual disability and language impairment. We developed clonal models of GRIN2B deletion and loss-of-function mutations in a region coding for the glutamate binding domain in human cells and generated neurons from a patient harboring a missense mutation in the same domain. Transcriptome analysis revealed extensive increases in genes associated with cell proliferation and decreases in genes associated with neuron differentiation, a result supported by extensive protein analyses. Using electrophysiology and calcium imaging, we demonstrate that NMDA receptors are present on neural progenitor cells and that human mutations in GRIN2B can impair calcium influx and membrane depolarization even in a presumed undifferentiated cell state, highlighting an important role for non-synaptic NMDA receptors. It may be this function, in part, which underlies the neurological disease observed in patients with GRIN2B mutations.
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http://dx.doi.org/10.1016/j.stemcr.2018.05.018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6067152PMC
July 2018

Strategies to Advance Drug Discovery in Rare Monogenic Intellectual Disability Syndromes.

Int J Neuropsychopharmacol 2018 03;21(3):201-206

Department of Human Genetics, McGill University, Montreal, QC, Canada.

Some intellectual disability syndromes are caused by a mutation in a single gene and have been the focus of therapeutic intervention attempts, such as Fragile X and Rett Syndrome, albeit with limited success. The rate at which new drugs are discovered and tested in humans for intellectual disability is progressing at a relatively slow pace. This is particularly true for rare diseases where so few patients make high-quality clinical trials challenging. We discuss how new advances in human stem cell reprogramming and gene editing can facilitate preclinical study design and we propose new workflows for how the preclinical to clinical trajectory might proceed given the small number of subjects available in rare monogenic intellectual disability syndromes.
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http://dx.doi.org/10.1093/ijnp/pyx090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836272PMC
March 2018

Strategies to Advance Drug Discovery in Rare Monogenic Intellectual Disability Syndromes.

Int J Neuropsychopharmacol 2018 03;21(3):201-206

Department of Human Genetics, McGill University, Montreal, QC, Canada.

Some intellectual disability syndromes are caused by a mutation in a single gene and have been the focus of therapeutic intervention attempts, such as Fragile X and Rett Syndrome, albeit with limited success. The rate at which new drugs are discovered and tested in humans for intellectual disability is progressing at a relatively slow pace. This is particularly true for rare diseases where so few patients make high-quality clinical trials challenging. We discuss how new advances in human stem cell reprogramming and gene editing can facilitate preclinical study design and we propose new workflows for how the preclinical to clinical trajectory might proceed given the small number of subjects available in rare monogenic intellectual disability syndromes.
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http://dx.doi.org/10.1093/ijnp/pyx090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836272PMC
March 2018

DNA Methylation Dynamics and Cocaine in the Brain: Progress and Prospects.

Genes (Basel) 2017 May 12;8(5). Epub 2017 May 12.

Department of Psychiatry, McGill Group for Suicide Studies, Douglas Mental Health University Institute, McGill University, Verdun, QC H4H 1R3, Canada.

Cytosine modifications, including DNA methylation, are stable epigenetic marks that may translate environmental change into transcriptional regulation. Research has begun to investigate DNA methylation dynamics in relation to cocaine use disorders. Specifically, DNA methylation machinery, including methyltransferases and binding proteins, are dysregulated in brain reward pathways after chronic cocaine exposure. In addition, numerous methylome-wide and candidate promoter studies have identified differential methylation, at the nucleotide level, in rodent models of cocaine abuse and drug seeking behavior. This review highlights the current progress in the field of cocaine-related methylation, and offers considerations for future research.
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http://dx.doi.org/10.3390/genes8050138DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5448012PMC
May 2017

A Rapid Pipeline to Model Rare Neurodevelopmental Disorders with Simultaneous CRISPR/Cas9 Gene Editing.

Stem Cells Transl Med 2017 03 1;6(3):886-896. Epub 2016 Dec 1.

McGill Group for Suicide Studies, Douglas Hospital Research Institute, Montreal, Quebec, H4H 1R3, Canada.

The development of targeted therapeutics for rare neurodevelopmental disorders (NDDs) faces significant challenges due to the scarcity of subjects and the difficulty of obtaining human neural cells. Here, we illustrate a rapid, simple protocol by which patient derived cells can be reprogrammed to induced pluripotent stem cells (iPSCs) using an episomal vector and differentiated into neurons. Using this platform enables patient somatic cells to be converted to physiologically active neurons in less than two months with minimal labor. This platform includes a method to combine somatic cell reprogramming with CRISPR/Cas9 gene editing at single cell resolution, which enables the concurrent development of clonal knockout or knock-in models that can be used as isogenic control lines. This platform reduces the logistical barrier for using iPSC technology, allows for the development of appropriate control lines for use in rare neurodevelopmental disease research, and establishes a fundamental component to targeted therapeutics and precision medicine. Stem Cells Translational Medicine 2017;6:886-896.
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http://dx.doi.org/10.1002/sctm.16-0158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5442775PMC
March 2017

Potential molecular consequences of transgene integration: The R6/2 mouse example.

Sci Rep 2017 01 25;7:41120. Epub 2017 Jan 25.

Molecular Neurogenetics Unit, Center for Human Genetic Research, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.

Integration of exogenous DNA into a host genome represents an important route to generate animal and cellular models for exploration into human disease and therapeutic development. In most models, little is known concerning structural integrity of the transgene, precise site of integration, or its impact on the host genome. We previously used whole-genome and targeted sequencing approaches to reconstruct transgene structure and integration sites in models of Huntington's disease, revealing complex structural rearrangements that can result from transgenesis. Here, we demonstrate in the R6/2 mouse, a widely used Huntington's disease model, that integration of a rearranged transgene with coincident deletion of 5,444 bp of host genome within the gene Gm12695 has striking molecular consequences. Gm12695, the function of which is unknown, is normally expressed at negligible levels in mouse brain, but transgene integration has resulted in cortical expression of a partial fragment (exons 8-11) 3' to the transgene integration site in R6/2. This transcript shows significant expression among the extensive network of differentially expressed genes associated with this model, including synaptic transmission, cell signalling and transcription. These data illustrate the value of sequence-level resolution of transgene insertions and transcription analysis to inform phenotypic characterization of transgenic models utilized in therapeutic research.
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http://dx.doi.org/10.1038/srep41120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5264158PMC
January 2017

Medium throughput bisulfite sequencing for accurate detection of 5-methylcytosine and 5-hydroxymethylcytosine.

BMC Genomics 2017 01 18;18(1):96. Epub 2017 Jan 18.

Department of Psychiatry, McGill Group for Suicide Studies, Douglas Mental Health University Institute, McGill University, Montreal, 6875 boul. LaSalle, Verdun, QC, H4H 1R3, Canada.

Background: Epigenetic modifications of DNA, such as 5-methylcytosine and 5-hydroxymethycytosine, play important roles in development and disease. Here, we present a cost-effective and versatile methodology for the analysis of DNA methylation in targeted genomic regions, which comprises multiplexed, PCR-based preparation of bisulfite DNA libraries followed by customized MiSeq sequencing.

Results: Using bisulfite and oxidative bisulfite conversion of DNA, we have performed multiplexed targeted sequencing to analyse several kilobases of genomic DNA in up to 478 samples, and achieved high coverage data of 5-methylcytosine and 5-hydroxymethycytosine at single-base resolution. Our results demonstrate the ability of this methodology to detect all levels of cytosine modifications at greater than 100× coverage in large sample sets at low cost compared to other targeted methods.

Conclusions: This approach can be applied to multiple settings, from candidate gene to clinical studies, and is especially useful for validation of differentially methylated or hydroxymethylated regions following whole-genome analyses.
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http://dx.doi.org/10.1186/s12864-017-3489-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5242011PMC
January 2017

Lesch-Nyhan Syndrome: Models, Theories, and Therapies.

Mol Syndromol 2016 Nov 24;7(6):302-311. Epub 2016 Sep 24.

Department of Psychiatry, Douglas Hospital Research Institute, McGill University, Montreal, Que., Canada.

Lesch-Nyhan syndrome (LNS) is a rare X-linked disorder caused by mutations in HPRT1, an important enzyme in the purine salvage pathway. Symptoms of LNS include dystonia, gout, intellectual disability, and self-mutilation. Despite having been characterized over 50 years ago, it remains unclear precisely how deficits in hypoxanthine and guanine recycling can lead to such a profound neurological phenotype. Several studies have proposed different hypotheses regarding the etiology of this disease, and several treatments have been tried in patients, though none have led to a satisfactory explanation of the disease. New technologies such as next-generation sequencing, optogenetics, genome editing, and induced pluripotent stem cells provide a unique opportunity to map the precise sequential pathways leading from genotype to phenotype.
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http://dx.doi.org/10.1159/000449296DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5131334PMC
November 2016

Implication of LRRC4C and DPP6 in neurodevelopmental disorders.

Am J Med Genet A 2017 Feb 19;173(2):395-406. Epub 2016 Oct 19.

The Centre for Applied Genomics and Genetics and Genome Biology, The Hospital for Sick Children, Toronto, Canada.

We performed whole-genome sequencing on an individual from a family with variable psychiatric phenotypes that had a sensory processing disorder, apraxia, and autism. The proband harbored a maternally inherited balanced translocation (46,XY,t(11;14)(p12;p12)mat) that disrupted LRRC4C, a member of the highly specialized netrin G family of axon guidance molecules. The proband also inherited a paternally derived chromosomal inversion that disrupted DPP6, a potassium channel interacting protein. Copy Number (CN) analysis in 14,077 cases with neurodevelopmental disorders and 8,960 control subjects revealed that 60% of cases with exonic deletions in LRRC4C had a second clinically recognizable syndrome associated with variable clinical phenotypes, including 16p11.2, 1q44, and 2q33.1 CN syndromes, suggesting LRRC4C deletion variants may be modifiers of neurodevelopmental disorders. In vitro, functional assessments modeling patient deletions in LRRC4C suggest a negative regulatory role of these exons found in the untranslated region of LRRC4C, which has a single, terminal coding exon. These data suggest that the proband's autism may be due to the inheritance of disruptions in both DPP6 and LRRC4C, and may highlight the importance of the netrin G family and potassium channel interacting molecules in neurodevelopmental disorders. © 2016 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/ajmg.a.38021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5833302PMC
February 2017

Proliferation and Differentiation Deficits are a Major Convergence Point for Neurodevelopmental Disorders.

Authors:
Carl Ernst

Trends Neurosci 2016 05 28;39(5):290-299. Epub 2016 Mar 28.

Department of Psychiatry, McGill University, Montreal, QC, Canada; Department of Human Genetics, McGill University, Montreal, QC, Canada; Integrated Program in Neuroscience, McGill University, Montreal, QC, Canada; McGill Group for Suicide Studies, Douglas Hospital Research Institute, Montreal, QC, Canada. Electronic address:

Several lines of evidence suggest that proliferation and differentiation in neural stem cells (NSCs) are a major convergence point of neurodevelopmental disorders (NDDs). Most genes with truncating mutations are implicated in NSC proliferation and differentiation (e.g., MBD5, CDKL5, and MECP2). Similarly, reciprocal deletion/duplication copy-number variants (CNVs), such as 1q21.1 and 16p11.2, are inversely correlated with head size. In addition, pathways such as MAPK, mTOR, and RAS, which are important in cancer, a disease of uncontrolled cell proliferation, are implicated in NDDs. These deficits are a measurable output of patient-derived induced neural progenitor cells, and may represent a diagnostic tool and a possible clinical intervention point for molecular therapies, irrespective of genotype.
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http://dx.doi.org/10.1016/j.tins.2016.03.001DOI Listing
May 2016

Neurons diversify astrocytes in the adult brain through sonic hedgehog signaling.

Science 2016 Feb;351(6275):849-54

Centre for Research in Neuroscience, Department of Neurology and Neurosurgery, Brain Repair and Integrative Neuroscience Program, The Research Institute of the McGill University Health Centre, Montreal General Hospital, Montreal, Quebec, Canada.

Astrocytes are specialized and heterogeneous cells that contribute to central nervous system function and homeostasis. However, the mechanisms that create and maintain differences among astrocytes and allow them to fulfill particular physiological roles remain poorly defined. We reveal that neurons actively determine the features of astrocytes in the healthy adult brain and define a role for neuron-derived sonic hedgehog (Shh) in regulating the molecular and functional profile of astrocytes. Thus, the molecular and physiological program of astrocytes is not hardwired during development but, rather, depends on cues from neurons that drive and sustain their specialized properties.
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http://dx.doi.org/10.1126/science.aab3103DOI Listing
February 2016

A de novo frameshift mutation in chromodomain helicase DNA-binding domain 8 (CHD8): A case report and literature review.

Am J Med Genet A 2016 May 20;170A(5):1225-35. Epub 2016 Jan 20.

Department of Neurology and Neurosurgery, McGill University, Montreal, Quebec, Canada.

Mutations in chromodomain helicase DNA-binding domain 8 (CHD8) have been identified in independent genotyping studies of autism spectrum disorder. To better understand the phenotype associated with CHD8 mutations, we genotyped all CHD8 exons in carefully assessed cohorts of autism (n = 142), schizophrenia (SCZ; n = 143), and intellectual disability (ID; n = 94). We identified one frameshift mutation, seven non-synonymous variants, and six synonymous variants. The frameshift mutation, p.Asn2092Lysfs*2, which creates a premature stop codon leading to the loss of 212 amino acids of the protein, was from an autism case on whom we present multiple clinical assessments and pharmacological treatments spanning more than 10 years. RNA and protein analysis support a model where the transcript generated from the mutant allele results in haploinsufficiency of CHD8. This case report supports the association of CHD8 mutations with classical autism, macrocephaly, infantile hypotonia, speech delay, lack of major ID, and psychopathology in late adolescence caused by insufficient dosage of CHD8. Review of 16 other CHD8 mutation cases suggests that clinical features and their severity vary considerably across individuals; however, these data support a CHD8 mutation syndrome, further highlighting the importance of genomic medicine to guide clinical assessment and treatment.
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http://dx.doi.org/10.1002/ajmg.a.37566DOI Listing
May 2016

Characterizing 5-hydroxymethylcytosine in human prefrontal cortex at single base resolution.

BMC Genomics 2015 Sep 3;16:672. Epub 2015 Sep 3.

McGill Group for Suicide Studies, Douglas Mental Health University Institute, 6875 boul. Lasalle, Montreal, Quebec, Canada.

Background: The recent discovery that methylated cytosines are converted to 5-hydroxymethylated cytosines (5hmC) by the family of ten-eleven translocation enzymes has sparked significant interest on the genomic location, the abundance in different tissues, the putative functions, and the stability of this epigenetic mark. 5hmC plays a key role in the brain, where it is particularly abundant and dynamic during development.

Results: Here, we comprehensively characterize 5hmC in the prefrontal cortices of 24 subjects. We show that, although there is inter-individual variability in 5hmC content among unrelated individuals, approximately 8 % of all CpGs on autosomal chromosomes contain 5hmC, while sex chromosomes contain far less. Our data also provide evidence suggesting that 5hmC has transcriptional regulatory properties, as the density of 5hmC was highest in enhancer regions and within exons. Furthermore, we link increased 5hmC density to histone modification binding sites, to the gene bodies of actively transcribed genes, and to exon-intron boundaries. Finally, we provide several genomic regions of interest that contain gender-specific 5hmC.

Conclusions: Collectively, these results present an important reference for the growing number of studies that are interested in the investigation of the role of 5hmC in brain and mental disorders.
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http://dx.doi.org/10.1186/s12864-015-1875-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4559220PMC
September 2015

Investigation of genes important in neurodevelopment disorders in adult human brain.

Hum Genet 2015 Oct 21;134(10):1037-53. Epub 2015 Jul 21.

Department of Psychiatry, McGill University, Montreal, QC, H4H 1R3, Canada.

Several neurodevelopmental disorders (NDDs) are caused by mutations in genes expressed in fetal brain, but little is known about these same genes in adult human brain. Here, we test the hypothesis that genes associated with NDDs continue to have a role in adult human brain to explore the idea that NDD symptoms may be partially a result of their adult function rather than just their neurodevelopmental function. To demonstrate adult brain function, we performed expression analyses and ChIPseq in human neural stem cell(NSC) lines at different developmental stages and adult human brain, targeting two genes associated with NDDs, SATB2 and EHMT1, and the WNT signaling gene TCF7L2, which has not been associated with NDDs. Analysis of DNA interaction sites in neural stem cells reveals high (40-50 %) overlap between proliferating and differentiating cells for each gene in temporal space. Studies in adult brain demonstrate that consensus sites are similar to NSCs but occur at different genomic locations. We also performed expression analyses using BrainSpan data for NDD-associated genes SATB2, EHMT1, FMR1, MECP2, MBD5, CTNND2, RAI1, CHD8, GRIN2A, GRIN2B, TCF4, SCN2A, and DYRK1A and find high expression of these genes in adult brain, at least comparable to developing human brain, confirming that genes associated with NDDs likely have a role in adult tissue. Adult function of genes associated with NDDs might be important in clinical disease presentation and may be suitable targets for therapeutic intervention.
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http://dx.doi.org/10.1007/s00439-015-1584-zDOI Listing
October 2015

Biomarker discovery: quantification of microRNAs and other small non-coding RNAs using next generation sequencing.

BMC Med Genomics 2015 Jul 1;8:35. Epub 2015 Jul 1.

McGill Group for Suicide Studies (MGSS), Douglas Mental Health University Institute, McGill University, Frank B Common Pavilion, Room F-2101.2, 6875 LaSalle Boulevard, Montreal, QC, H4H 1R3, Canada.

Background: Small ncRNAs (sncRNAs) offer great hope as biomarkers of disease and response to treatment. This has been highlighted in the context of several medical conditions such as cancer, liver disease, cardiovascular disease, and central nervous system disorders, among many others. Here we assessed several steps involved in the development of an ncRNA biomarker discovery pipeline, ranging from sample preparation to bioinformatic processing of small RNA sequencing data.

Methods: A total of 45 biological samples were included in the present study. All libraries were prepared using the Illumina TruSeq Small RNA protocol and sequenced using the HiSeq2500 or MiSeq Illumina sequencers. Small RNA sequencing data was validated using qRT-PCR. At each stage, we evaluated the pros and cons of different techniques that may be suitable for different experimental designs. Evaluation methods included quality of data output in relation to hands-on laboratory time, cost, and efficiency of processing.

Results: Our results show that good quality sequencing libraries can be prepared from small amounts of total RNA and that varying degradation levels in the samples do not have a significant effect on the overall quantification of sncRNAs via NGS. In addition, we describe the strengths and limitations of three commercially available library preparation methods: (1) Novex TBE PAGE gel; (2) Pippin Prep automated gel system; and (3) AMPure XP beads. We describe our bioinformatics pipeline, provide recommendations for sequencing coverage, and describe in detail the expression and distribution of all sncRNAs in four human tissues: whole-blood, brain, heart and liver.

Conclusions: Ultimately this study provides tools and outcome metrics that will aid researchers and clinicians in choosing an appropriate and effective high-throughput sequencing quantification method for various study designs, and overall generating valuable information that can contribute to our understanding of small ncRNAs as potential biomarkers and mediators of biological functions and disease.
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http://dx.doi.org/10.1186/s12920-015-0109-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4487992PMC
July 2015

Comparative analysis of self-injury in people with psychopathology or neurodevelopmental disorders.

Pediatr Clin North Am 2015 Jun 7;62(3):619-31. Epub 2015 Apr 7.

Integrated Program in Neuroscience, McGill University, Montreal, Quebec H3A 0G4, Canada; McGill Group for Suicide Studies, Douglas Mental Health University Institute, Montreal, Quebec H4H 1R3, Canada; Department of Psychiatry, McGill University, Montreal, Quebec H3A 0G4, Canada; Department of Human Genetics, McGill University, Montreal, Quebec H3A 0G4, Canada. Electronic address:

Self-injury is a complex and poorly understood behavior observed in people with psychopathology or neurodevelopmental disorders (NDD). Despite the differences in etiology and progression of these distinct disease domains, it is possible that overlapping molecular pathways underlie the expression of self-injurious behaviors (SIBs). This review outlines the similarities and differences at the behavioural and molecular level, where SIBs in both conditions may involve opioid, nucleoside, and dopamine signalling. These points of convergence have important implications for treatment and research of SIB in both populations.
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http://dx.doi.org/10.1016/j.pcl.2015.03.001DOI Listing
June 2015
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