Publications by authors named "Samuel T Peters"

8 Publications

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DIAPH1 Variants in Non-East Asian Patients With Sporadic Moyamoya Disease.

JAMA Neurol 2021 Aug;78(8):993-1003

Yale Center for Genome Analysis, West Haven, Connecticut.

Importance: Moyamoya disease (MMD), a progressive vasculopathy leading to narrowing and ultimate occlusion of the intracranial internal carotid arteries, is a cause of childhood stroke. The cause of MMD is poorly understood, but genetic factors play a role. Several familial forms of MMD have been identified, but the cause of most cases remains elusive, especially among non-East Asian individuals.

Objective: To assess whether ultrarare de novo and rare, damaging transmitted variants with large effect sizes are associated with MMD risk.

Design, Setting, And Participants: A genetic association study was conducted using whole-exome sequencing case-parent MMD trios in a small discovery cohort collected over 3.5 years (2016-2019); data were analyzed in 2020. Medical records from US hospitals spanning a range of 1 month to 1.5 years were reviewed for phenotyping. Exomes from a larger validation cohort were analyzed to identify additional rare, large-effect variants in the top candidate gene. Participants included patients with MMD and, when available, their parents. All participants who met criteria and were presented with the option to join the study agreed to do so; none were excluded. Twenty-four probands (22 trios and 2 singletons) composed the discovery cohort, and 84 probands (29 trios and 55 singletons) composed the validation cohort.

Main Outcomes And Measures: Gene variants were identified and filtered using stringent criteria. Enrichment and case-control tests assessed gene-level variant burden. In silico modeling estimated the probability of variant association with protein structure. Integrative genomics assessed expression patterns of MMD risk genes derived from single-cell RNA sequencing data of human and mouse brain tissue.

Results: Of the 24 patients in the discovery cohort, 14 (58.3%) were men and 18 (75.0%) were of European ancestry. Three of 24 discovery cohort probands contained 2 do novo (1-tailed Poisson P = 1.1 × 10-6) and 1 rare, transmitted damaging variant (12.5% of cases) in DIAPH1 (mammalian diaphanous-1), a key regulator of actin remodeling in vascular cells and platelets. Four additional ultrarare damaging heterozygous DIAPH1 variants (3 unphased) were identified in 3 other patients in an 84-proband validation cohort (73.8% female, 77.4% European). All 6 patients were non-East Asian. Compound heterozygous variants were identified in ena/vasodilator-stimulated phosphoproteinlike protein EVL, a mammalian diaphanous-1 interactor that regulates actin polymerization. DIAPH1 and EVL mutant probands had severe, bilateral MMD associated with transfusion-dependent thrombocytopenia. DIAPH1 and other MMD risk genes are enriched in mural cells of midgestational human brain. The DIAPH1 coexpression network converges in vascular cell actin cytoskeleton regulatory pathways.

Conclusions And Relevance: These findings provide the largest collection to date of non-East Asian individuals with sporadic MMD harboring pathogenic variants in the same gene. The results suggest that DIAPH1 is a novel MMD risk gene and impaired vascular cell actin remodeling in MMD pathogenesis, with diagnostic and therapeutic ramifications.
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http://dx.doi.org/10.1001/jamaneurol.2021.1681DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8204259PMC
August 2021

Analysis workflow to assess genetic variants from human whole-exome sequencing.

STAR Protoc 2021 Mar 10;2(1):100383. Epub 2021 Mar 10.

Department of Genetics, Washington University School of Medicine, St. Louis, MO, USA.

Here, we present a protocol to analyze genetic variants derived from the whole-exome sequencing (WES) of proband-parent trios. We provide stepwise instructions for using existing pipelines to call mutations (DNMs) and determine whether the observed number of such mutations is enriched relative to the expected number. This protocol may be extended to any human disease trio-based cohort. Cohort size is a limiting determinant to the discovery of high-confidence pathogenic DNMs. For complete details on the use and execution of this protocol, please refer to Dong et al. (2020).
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http://dx.doi.org/10.1016/j.xpro.2021.100383DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7960548PMC
March 2021

Case Report: Early-Onset Behavioral Variant Frontotemporal Dementia in Patient With Retrotransposed Full-Length Transcript of Variant 5.

Front Neurol 2020 21;11:600468. Epub 2020 Dec 21.

Department of Neurology, N. Bud Grossman Center for Memory Research and Care, University of Minnesota, Minneapolis, MN, United States.

Frontotemporal dementia (FTD) rarely occurs in individuals under the age of 30, and genetic causes of early-onset FTD are largely unknown. The current report follows a 27 year-old patient with no significant past medical history presenting with two years of progressive changes in behavior, rushed speech, verbal aggression, and social withdrawal. MRI and FDG-PET imaging of the brain revealed changes maximally in the frontal and temporal lobes, which along with the clinical features, are consistent with behavioral variant FTD. Next generation sequencing of a panel of 28 genes associated with dementia and amyotrophic lateral sclerosis (ALS) initially revealed a duplication of exon 15 in (). Whole genome sequencing determined that this genetic anomaly was, in fact, a sequence corresponding with full-length variant 5 inserted into chromosome 12, indicating retrotransposition from a messenger RNA intermediate. To our knowledge, this is a novel mutation of , as the majority of mutations in linked to FTD-ALS are point mutations. Genomic DNA analysis revealed that this mutation is also present in one unaffected first-degree relative and one unaffected second-degree relative. This suggests that the mutation is either a disease-causing mutation with incomplete penetrance, which has been observed in heritable FTD, or a benign variant. Retrotransposons are not often implicated in neurodegenerative diseases; thus, it is crucial to clarify the potential role of this variant 5 retrotransposition in early-onset FTD.
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http://dx.doi.org/10.3389/fneur.2020.600468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7779795PMC
December 2020

In Vivo CRISPR/Cas9-Based Targeted Disruption and Knockin of a Long Noncoding RNA.

Methods Mol Biol 2021 ;2254:305-321

Department of Physiology and Pharmacology, University of Toledo College of Medicine and Life Sciences, Toledo, OH, USA.

The CRISPR/Cas9 system has been widely used as an efficient genome-editing tool for studying physiological functions of long noncoding RNAs (lncRNAs). In this chapter, we describe the experimental procedures for using the CRISPR/Cas9 system to genetically modify a long noncoding RNA in vivo through the targeted disruption and knockin approaches.
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http://dx.doi.org/10.1007/978-1-0716-1158-6_19DOI Listing
March 2021

Ablating Tau Reduces Hyperexcitability and Moderates Electroencephalographic Slowing in Transgenic Mice Expressing A53T Human α-Synuclein.

Front Neurol 2020 19;11:563. Epub 2020 Jun 19.

Department of Neurology, N. Bud Grossman Center for Memory Research and Care, University of Minnesota, Minneapolis, MN, United States.

Abnormal intraneuronal accumulation of the presynaptic protein α-synuclein (α-syn) is implicated in the etiology of dementia with Lewy bodies (DLB) and Parkinson's disease with dementia (PDD). Recent work revealed that mice expressing human α-syn with the alanine-53-threonine (A53T) mutation have a similar phenotype to the human condition, exhibiting long-term potentiation deficits, learning and memory deficits, and inhibitory hippocampal remodeling, all of which were reversed by genetic ablation of microtubule-associated protein tau. Significantly, memory deficits were associated with histological signs of network hyperactivity/seizures. Electrophysiological abnormalities are often seen in parkinsonian dementias. Baseline electroencephalogram (EEG) slowing is used as a supportive diagnostic feature in DLB and PDD, and patients with these diseases may exhibit indicators of broad network dysfunction such as sleep dysregulation, myoclonus, and seizures. Given the translational significance, we examined whether human A53T α-syn expressing mice exhibit endogenous-tau-dependent EEG abnormalities, as measured with epidural electrodes over the frontal and parietal cortices. Using template-based waveform sorting, we determined that A53T mice have significantly high numbers of epileptiform events as early as 3-4 months of age and throughout life, and this effect is markedly attenuated in the absence of tau. Epileptic myoclonus occurred in half of A53T mice and was markedly reduced by tau ablation. In spectral analysis, tau ablation partially reduced EEG slowing in 6-7 month transgenic mice. We found abnormal sleeping patterns in transgenic mice that were more pronounced in older groups, but did not find evidence that this was influenced by tau genotype. Together, these data support the notion that tau facilitates A53T α-syn-induced hyperexcitability that both precedes and coincides with associated synaptic, cognitive, and behavioral effects. Tau also contributes to some aspects of EEG slowing in A53T mice. Importantly, our work supports tau-based approaches as an effective early intervention in α-synucleinopathies to treat aberrant network activity.
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http://dx.doi.org/10.3389/fneur.2020.00563DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7316964PMC
June 2020

Genome editing of HBG1 and HBG2 to induce fetal hemoglobin.

Blood Adv 2019 11;3(21):3379-3392

Department of Hematology, St. Jude Children's Research Hospital, Memphis, TN.

Induction of fetal hemoglobin (HbF) via clustered regularly interspaced short palindromic repeats/Cas9-mediated disruption of DNA regulatory elements that repress γ-globin gene (HBG1 and HBG2) expression is a promising therapeutic strategy for sickle cell disease (SCD) and β-thalassemia, although the optimal technical approaches and limiting toxicities are not yet fully defined. We disrupted an HBG1/HBG2 gene promoter motif that is bound by the transcriptional repressor BCL11A. Electroporation of Cas9 single guide RNA ribonucleoprotein complex into normal and SCD donor CD34+ hematopoietic stem and progenitor cells resulted in high frequencies of on-target mutations and the induction of HbF to potentially therapeutic levels in erythroid progeny generated in vitro and in vivo after transplantation of hematopoietic stem and progenitor cells into nonobese diabetic/severe combined immunodeficiency/Il2rγ-/-/KitW41/W41 immunodeficient mice. On-target editing did not impair CD34+ cell regeneration or differentiation into erythroid, T, B, or myeloid cell lineages at 16 to 17 weeks after xenotransplantation. No off-target mutations were detected by targeted sequencing of candidate sites identified by circularization for in vitro reporting of cleavage effects by sequencing (CIRCLE-seq), an in vitro genome-scale method for detecting Cas9 activity. Engineered Cas9 containing 3 nuclear localization sequences edited human hematopoietic stem and progenitor cells more efficiently and consistently than conventional Cas9 with 2 nuclear localization sequences. Our studies provide novel and essential preclinical evidence supporting the safety, feasibility, and efficacy of a mechanism-based approach to induce HbF for treating hemoglobinopathies.
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http://dx.doi.org/10.1182/bloodadvances.2019000820DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6855127PMC
November 2019

The COPII cargo adapter SEC24C is essential for neuronal homeostasis.

J Clin Invest 2018 08 25;128(8):3319-3332. Epub 2018 Jun 25.

Department of Pathology.

SEC24 family members are components of the coat protein complex II (COPII) machinery that interact directly with cargo or with other adapters to ensure proper sorting of secretory cargo into COPII vesicles. SEC24C is 1 of 4 mammalian SEC24 paralogs (SEC24A-D), which segregate into 2 subfamilies on the basis of sequence homology (SEC24A/SEC24B and SEC24C/SEC24D). Here, we demonstrate that postmitotic neurons, unlike professional secretory cells in other tissues, are exquisitely sensitive to loss of SEC24C. Conditional KO of Sec24c in neural progenitors during embryogenesis caused perinatal mortality and microcephaly, with activation of the unfolded protein response and apoptotic cell death of postmitotic neurons in the murine cerebral cortex. The cell-autonomous function of SEC24C in postmitotic neurons was further highlighted by the loss of cell viability caused by disrupting Sec24c expression in forebrain neurons of mice postnatally and in differentiated neurons derived from human induced pluripotent stem cells. The neuronal cell death associated with Sec24c deficiency was rescued in knockin mice expressing Sec24d in place of Sec24c. These data suggest that SEC24C is a major cargo adapter for COPII-dependent transport in postmitotic neurons in developing and adult brains and that its functions overlap at least partially with those of SEC24D in mammals.
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http://dx.doi.org/10.1172/JCI98194DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6063503PMC
August 2018

A Survey of Validation Strategies for CRISPR-Cas9 Editing.

Sci Rep 2018 01 17;8(1):888. Epub 2018 Jan 17.

St. Jude Children's Research Hospital, Department of Cell & Molecular Biology, Memphis, 38105, USA.

The T7 endonuclease 1 (T7E1) mismatch detection assay is a widely used method for evaluating the activity of site-specific nucleases, such as the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system. To determine the accuracy and sensitivity of this assay, we compared the editing estimates derived by the T7E1 assay with that of targeted next-generation sequencing (NGS) in pools of edited mammalian cells. Here, we report that estimates of nuclease activity determined by T7E1 most often do not accurately reflect the activity observed in edited cells. Editing efficiencies of CRISPR-Cas9 complexes with similar activity by T7E1 can prove dramatically different by NGS. Additionally, we compared editing efficiencies predicted by the Tracking of Indels by Decomposition (TIDE) assay and the Indel Detection by Amplicon Analysis (IDAA) assay to that observed by targeted NGS for both cellular pools and single-cell derived clones. We show that targeted NGS, TIDE, and IDAA assays predict similar editing efficiencies for pools of cells but that TIDE and IDAA can miscall alleles in edited clones.
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http://dx.doi.org/10.1038/s41598-018-19441-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5772360PMC
January 2018
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