In vivo Perturb-Seq reveals neuronal and glial abnormalities associated with autism risk genes
Open accessPublished version of a preprint
Abstract
An in vivo analysis of autism risk genes: CRISPR targeting in vivo, especially in mammals, can be difficult and time consuming when attempting to determine the effects of a single gene. However, such studies may be required to identify pathological gene variants with effects in specific cells along a developmental trajectory. To study the function of genes implicated in autism spectrum disorders (ASDs), Jin et al. applied a gene-editing and single-cell–sequencing system, Perturb-Seq, to knock out 35 ASD candidate genes in multiple mice embryos (see the Perspective by Treutlein and Camp). This method identified networks of gene expression in neuronal and glial cells that suggest new functions in ASD-related genes. Science , this issue p. eaaz6063 ; see also p. 1038
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- Findingown data
Loss of Dyrk1a significantly reduced the proportion of microglia/macrophages.
“Only loss of Dyrk1a had a significant effect on cell type composition, increasing the proportion of oligodendrocytes and reducing the proportion of microglia/macrophages [FDR-corrected P<0.05 using Poisson regression (18)] (Figs. 1D and S5).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [18] Haber AL, A single-cell survey of the small intestinal epithelium. Nature 551, 333– 339 (2017). 29144463 PMC6022292 source
Not checked in the free run.
- Findingown data
ASD/ND risk gene perturbations had a very modest effect on the presence and proportions of the five major cell types relative to the negative control.
“ASD/ND risk gene perturbations had a very modest effect on the presence and proportions of these five major cell types relative to the negative control (targeting the GFP gene).”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- Findingown data
Lentiviral vector expression was detected in cortical microglia, indicated by BFP and perturbation barcode expression, across multiple individual experiments.
“we detected lentiviral vector expression in cortical microglia, indicated by the presence of BFP as well as perturbation barcode expression, across multiple individual experiments (fig. S4E–F).”
Quote in the paperNot a substantial claim - Findingown data
50% of cells in the five major cell classes had barcodes for a single gene.
“and 50% had barcodes for a single gene (fig. S4A–C, 18,044 cells), reflecting the low multiplicity of infection (fig. S4D).”
Quote in the paperSubstantial claim - Findingown data
Perturb-Seq vectors were expressed across a variety of neuronal and glial cell types in the cortex at P7.
“Both immunohistochemical analysis and scRNA-seq of BFP+ cells at P7 showed that the Perturb-Seq vectors were expressed across a variety of neuronal and glial cell types in the cortex (Figs. 1B–C and S3A–B).”
Quote in the paperSubstantial claim
- Findingcitation
Orthologs of the selected ASD/ND risk genes are expressed in diverse cell types in mouse cortex.
“Based on mouse cortical single-cell RNA sequencing (scRNA-seq) data, the orthologs of these ASD/ND risk genes are expressed in diverse cell types (fig. S2) (E18.5 data from the 10x Genomics public dataset (10); P7 data from this work).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [10] Data of 9k brain cells from an E18 mouse, 10x Genomics: https://support.10xgenomics.com/single-cell-gene-expression/datasets/2.1.0/neuron_9k.
Couldn’t be matched to a paper in Crossref, so it can’t be checked.
- Findingown data
92% of cells in the five major cell classes had at least one perturbation read assigned.
“From inspecting the perturbation barcodes from the lentiviral constructs, 92% (33,231 cells) of the cells in these five major cell classes had at least one perturbation read assigned to them”
Quote in the paperSubstantial claim - Findingown data
Loss of Dyrk1a significantly increased the proportion of oligodendrocytes.
“Only loss of Dyrk1a had a significant effect on cell type composition, increasing the proportion of oligodendrocytes and reducing the proportion of microglia/macrophages [FDR-corrected P<0.05 using Poisson regression (18)] (Figs. 1D and S5).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [18] Haber AL, A single-cell survey of the small intestinal epithelium. Nature 551, 333– 339 (2017). 29144463 PMC6022292 source
Not checked in the free run.
- Findingown data
35 ASD/ND risk gene perturbations were retained after excluding those with fewer than 70 perturbed cells.
“after excluding perturbations with <70 perturbed cells, we retained 35 ASD/ND risk gene perturbations.”
Quote in the paperSubstantial claim - Findingown data
Cells positive for MBP protein were increased in number in Chd8+/− mutant at P11 (FDR<0.05).
“cells positive for the MBP protein, a marker of myelinating oligodendrocytes, were increased in number and displayed elevated MBP levels in the Chd8+/− mutant (FDR corrected P<0.05, nonparametric ANOVA test) (Fig. 3F).”
Quote in the paperSubstantial claim - Findingown data
Module Astro3 was affected by perturbations of Chd8, Pogz, and Upf3b.
“modules representing two distinct homeostatic signatures in astrocytes (Astro1 affected by perturbation of Scn2a1, and Astro3 affected by perturbations of Chd8, Pogz, and Upf3b)”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
The IN1 module was strongly correlated with a subcluster of inhibitory interneurons expressing Ndnf.
“This module was strongly correlated with a subcluster of inhibitory interneurons expressing Ndnf (fig. S10C, D)”
Quote in the paperNot a substantial claim - Findingown data
Module ODC1 was affected by Chd8 and Gatad2b.
“a module associated with oligodendrocyte progenitor cells (ODC1, Chd8 and Gatad2b)”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingcitation
Correlation increased with the age of human samples across brain regions of the BrainSpan dataset.
“Correlation also increased with the age of the human samples across brain regions of the BrainSpan dataset (9) (Figs. 4C–E and S13).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [9] Miller JA, Transcriptional landscape of the prenatal human brain. Nature 508, 199– 206 (2014). 24695229 PMC4105188
Not checked in the free run.
- Findingown data
In postmortem snRNA-seq data from 15 ASD donors and 16 controls, 14 genes were differentially expressed between patients and controls in at least one of three major cell types (inhibitory neurons, excitatory neurons, or oligodendrocytes) at FDR < 0.2 and had 1:1 mouse orthologs.
“We identified genes that were differentially expressed between patients and controls in at least one of three major cell types (inhibitory neurons, excitatory neurons, or oligodendrocytes) with FDR < 0.2, and selected those that have 1:1 orthologs in mice, resulting in 14 genes (Fig. 4F, Table S10).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [31] Velmeshev D, Single-cell genomics identifies cell type-specific molecular changes in autism. Science 364, 685– 689 (2019). 31097668 PMC7678724 source
Not checked in the free run.
- Findingown data
Ank2 perturbation led to increased expression of the interneuron module IN1 (FDR<0.05).
“In our multiplex in vivo Perturb-Seq results, Ank2 perturbation led to increased expression of an interneuron module (IN1) (FDR corrected P<0.05, fig. S10).”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
Module PN1 was affected by perturbations in Adnp, Ash1l, Scn2a1, and Stard9.
“a module associated with projection neurons of Layer 4 and 5 (PN1, affected by perturbations in Adnp, Ash1l, Scn2a1, and Stard9)”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
Within Ndnf+ interneurons, Ank2 perturbation led to upregulation of the IN1 module (FDR<0.05), confirming the Perturb-Seq result.
“Within the Ndnf+ interneurons, Ank2 perturbation led to upregulation of the IN1 module (FDR corrected P<0.05, fig. S10E), confirming the Perturb-Seq result.”
Quote in the paperSubstantial claim - Findingown data
Perturbations in 9 ASD/ND genes (Adnp, Ank2, Ash1l, Chd8, Gatad2b, Pogz, Scn2a1, Stard9, and Upf3b) had significant effects across 5 modules compared to GFP control (FDR<0.05).
“Perturbations in 9 ASD/ND genes (Adnp, Ank2, Ash1l, Chd8, Gatad2b, Pogz, Scn2a1, Stard9, and Upf3b) had significant effects across 5 modules (Fig. 2B, highlighted circles, compared to the GFP control, FDR corrected P<0.05, Table S4)”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
Module PN1 is associated with two defined subclusters of projection neurons of Layer 4 and Layer 5.
“For example, module PN1 is a module associated with two defined subclusters of projection neurons of Layer 4 and Layer 5 (fig. S6A).”
Quote in the paperNot a substantial claim - Findingown data
ODC1 variation across the oligodendrocyte cell cluster was significantly explained by perturbation state overall (van der Waerden test, FDR<0.05).
“Notably, the oligodendrocyte progenitor module (ODC1) also had a significant amount of its variation across the oligodendrocyte cell cluster explained by the perturbation state overall (van der Waerden test, a non-parametric alternative to ANOVA analysis, FDR corrected P<0.05) (fig. S5C)”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
Using Perturb-Seq to functionally test large gene sets in the developing embryo, the authors observed gene expression changes linked to ASD/ND genes in different cell types and processes.
“Using Perturb-Seq to functionally test large gene sets in the developing embryo, we observed gene expression changes linked to ASD/ND genes in different cell types and processes.”
Quote in the paperSubstantial claim- Rests on an assumption: The cellular and molecular substrates of ASD/ND are not yet defined.
- Rests on an assumption: A large number of highly penetrant de novo risk genes implicated through human genetic studies provides an entry point to identify the cell types, developmental events, and mechanisms underlying ASD/ND.
- Rests on an assumption: Scalable methods are required to define the function of risk-associated genes with cell-type specificity.
- Findingown data
8 out of 14 modules showed greater intra-module correlation than a comparable random gene set in the adult human brain dataset from Hodge et al.
“8 out of 14 modules showed greater intra-module correlation than a comparable random gene set in the adult human brain dataset from Hodge et al (30) (Fig. 4B).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [30] Hodge RD, Conserved cell types with divergent features in human versus mouse cortex. Nature 573, 61– 68 (2019). 31435019 PMC6919571 source
Not checked in the free run.
- Findingown data
Modularity was decreased in non-associated cell types, reflected by both the proportion of comparisons with significant correlation and the strength of significant correlations.
“We found that the modularity was decreased in non-associated cell types (fig. S12D–E), reflected by both the proportion of comparisons with significant correlation and by the strength of the significant correlations”
Quote in the paperSubstantial claim - Findingown data
Module Astro1 was affected by perturbation of Scn2a1.
“modules representing two distinct homeostatic signatures in astrocytes (Astro1 affected by perturbation of Scn2a1, and Astro3 affected by perturbations of Chd8, Pogz, and Upf3b)”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
Modules PN1, PN2, and PN5 displayed high levels of conservation of expression across all datasets, with at least 75% of their genes expressed by at least 5% of cells in the corresponding associated cell type.
“Some modules like PN1, PN2, and PN5 displayed high levels of conservation of expression (with at least 75% of the genes in these modules being expressed by at least 5% of cells in the corresponding associated cell type) across all datasets.”
Quote in the paperNot a substantial claim - Findingown data
The IN1 module contains genes such as Kcnq5 and Gabbr2.
“and contains genes such as Kcnq5 (a voltage-gated potassium channel) and Gabbr2 (GABA receptor subunit) (fig. S6B and table S2).”
Quote in the paperNot a substantial claim - Findingown data
MBP levels were elevated in Chd8+/− mutant at P11.
“cells positive for the MBP protein, a marker of myelinating oligodendrocytes, were increased in number and displayed elevated MBP levels in the Chd8+/− mutant (FDR corrected P<0.05, nonparametric ANOVA test) (Fig. 3F).”
Quote in the paperSubstantial claim - Findingown data
Cspg4 and Pdgfra were downregulated in P7 Chd8+/− cortex, consistent with in vivo Perturb-Seq results.
“Both were downregulated in P7 Chd8+/− cortex (Figs. 3E and S11B–D), consistent with our in vivo Perturb-Seq results.”
Quote in the paperSubstantial claim - Findingown data
The proportions of genes expressed in corresponding cell types in human tissues were generally lower than in mouse tissues.
“The proportions of the genes expressed in the corresponding cell types in human tissues were generally lower than in mouse tissues (fig. S12B).”
Quote in the paperNot a substantial claim - Findingown data
In addition to neurons, oligodendrocytes and astrocytes were affected by several perturbations.
“In addition to neurons, oligodendrocytes and astrocytes were also affected by several perturbations.”
Quote in the paperSubstantial claim - Findingown data
Module IN1 was affected by Ank2.
“a module associated with Ndnf+ interneurons (IN1, Ank2) (Fig. 2C, fig. S6).”
Quote in the paperNot a substantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
In fetal brain and 3-month brain organoid samples, glial cell types were sparsely represented due to early developmental stages.
“In the fetal brain and the 3-month brain organoid samples, glial cell types were sparsely represented due to the early developmental stages of the samples (fig. S12A).”
Quote in the paperNot a substantial claim - Findingown data
Six of the 14 gene modules previously reported as altered in ASD patients in the PsychEncode analysis were also significantly affected across 8 of the ASD/ND risk gene perturbations.
“6 of the 14 modules previously reported to be altered in the ASD patients in the PsychEncode analysis were also significantly affected across 8 of our ASD/ND risk gene perturbations (fig. S14).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [34] Gandal MJ, Shared molecular neuropathology across major psychiatric disorders parallels polygenic overlap. Science 359, 693– 697 (2018). 29439242 PMC5898828 source
Not checked in the free run.
- Findingown data
No GO terms were significantly altered by any perturbation after multiple hypothesis testing correction.
“After correcting for multiple hypothesis testing, no GO terms were significantly altered by any perturbation (Table S8).”
Quote in the paperSubstantial claim - Findingown data
Chd8 and Gatad2b perturbations significantly decreased expression of the ODC1 module in the oligodendrocyte cluster (FDR<0.05).
“In our Perturb-Seq experiment, Chd8 and Gatad2b perturbations significantly decreased the expression of the ODC1 module in the oligodendrocyte cluster (Fig. 3A–D, FDR corrected P<0.05; see alternative measurement of effect size fig. S11A, estimated by log transcripts per million (TPM) gene expression differences).”
Quote in the paperSubstantial claim- Rests on an assumption: Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
- Findingown data
Within the power achievable with the number of cells that can be reasonably sequenced, some recurrent modules are affected across more than one ASD/ND risk gene perturbation.
“Within the power of the analysis that can be achieved with the number of cells that can be reasonably sequenced, we find that some recurrent modules are affected across more than one ASD/ND risk gene perturbation.”
Quote in the paperSubstantial claim - Findingown data
OPC cell number (PDGFRA+ cells) did not show significant differences between WT and Chd8+/− littermates at P11.
“OPC cell number (e.g., PDGFRA+ cells) did not show significant differences between the WT and Chd8+/− littermates, also consistent with in vivo Perturb-Seq”
Quote in the paperSubstantial claim - Findingown data
Only the PFC analysis yielded significant hits.
“Only analysis on the PFC yielded significant hits, which are presented in Fig 4F.”
Quote in the paperNot a substantial claim - Study design and conductown data
BFP from the lentiviral vector was robustly detected as one of the most highly expressed genes in all retained cells.
“BFP from the lentiviral vector was robustly detected as one of the most highly expressed genes in all retained cells (fig. S4E).”
Quote in the paperNot a substantial claim - Study design and conductown data
BFP detection rate in each cell type correlated with the average number of genes detected, supporting reliability of the readout.
“The BFP detection rate in each cell type correlated with the average number of genes detected (fig. S4F), further supporting the reliability of the readout.”
Quote in the paperNot a substantial claim - Premisecitation
The functions of the proteins encoded by these risk genes are substantially diverse, preventing a clear a priori prediction of the affected brain cell types, developmental processes, and molecular pathways during neurodevelopment.
“there is substantial diversity in the function of the gene products that these genes encode, precluding a clear a priori prediction of the underlying brain cell types, developmental processes, and molecular pathways affected during neurodevelopment (7).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [7] Mullins C, Fishell G, Tsien RW, Unifying Views of Autism Spectrum Disorders: A Consideration of Autoregulatory Feedback Loops. Neuron 89, 1131– 1156 (2016). 26985722 PMC5757244 source
Not checked in the free run.
- Interpretationown data
The decreased modularity in non-associated cell types suggests that the modules reflect cell type-specific effects.
“suggesting that our modules reflect cell type-specific effects.”
Quote in the paperSubstantial claim - Interpretationown data
The Perturb-Seq data suggest a role of Ank2 in the Ndnf+ interneuron subtype during cortical development, in addition to its known roles in excitatory neurons.
“Our Perturb-Seq data suggests a role of Ank2 in the Ndnf+ interneuron subtype during cortical development, in addition to its known roles in excitatory neurons.”
Quote in the paperSubstantial claim - Interpretationown data
The results suggest that expression and modularity of most gene modules in mouse are conserved in human brain tissue, pointing at potential shared functions.
“Altogether, our results suggest that expression and modularity of most gene modules in the mouse are conserved in human brain tissue, pointing at potential shared functions”
Quote in the paperSubstantial claim - Interpretationown data
These data demonstrate that in vivo Perturb-Seq has the power to identify cell type-specific molecular changes similar to those observed in a single-gene, germline-modified mouse model.
“These data further demonstrate that in vivo Perturb-Seq has the power to identify cell type-specific molecular changes similar to those observed in a single-gene, germline-modified mouse model.”
Quote in the paperSubstantial claim - Interpretationreasoning
The ODC1 module is linked to oligodendrocyte maturation.
“suggesting that this module is linked to oligodendrocyte maturation (Fig. 3A)”
Quote in the paperSubstantial claim - Premisecitation
The finding is consistent with recent reports that Chd8 loss-of-function potentiates an impaired OPC development phenotype caused by deletion of Chd7.
“This is consistent with recent reports that Chd8 loss-of-function potentiates an impaired OPC development phenotype caused by deletion of Chd7 (28).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [28] Marie C, Oligodendrocyte precursor survival and differentiation requires chromatin remodeling by Chd7 and Chd8. Proc Natl Acad Sci U S A, (2018). PMC6126750 30108144 source
Not checked in the free run.
- Interpretationown data
The ODC1 result suggests convergent effects across different perturbed genes.
“suggesting that this module represents convergent effects across different perturbed genes.”
Quote in the paperSubstantial claim - Interpretationown data
The Perturb-Seq data suggest an additional role for Ank2 in the Ndnf+ interneuron subtype, in addition to its known roles in excitatory neurons.
“Our Perturb-Seq data suggests an additional role of Ank2 in the Ndnf+ interneuron subtype, along with its known roles in excitatory neurons.”
Quote in the paperSubstantial claim - Interpretationreasoning
Processes identified as affected in the Perturb-Seq experiments are relevant to biological processes that may be developmentally regulated in the human brain.
“suggesting that processes identified as affected in our Perturb-Seq experiments are relevant to biological processes that may be developmentally regulated in the human brain.”
Quote in the paperSubstantial claim - Interpretationreasoning
Chd8 perturbation may result in acceleration of the progressive increase in MBP levels that occurs postnatally.
“this suggests that Chd8 perturbation may result in acceleration of the progressive increase in MBP levels that occurs postnatally.”
Quote in the paperSubstantial claim - Interpretationreasoning
Perturbation in Chd8 and Gatad2b might accelerate oligodendrocyte maturation.
“and therefore that perturbation in Chd8 and Gatad2b might accelerate oligodendrocyte maturation.”
Quote in the paperSubstantial claim - Interpretationown data
The individual perturbation experiment confirmed the results from the pooled Perturb-Seq screen.
“The individual perturbation experiment confirmed the results from the pooled Perturb-Seq screen.”
Quote in the paperSubstantial claim - Interpretationown data
Multiplexing perturbations in the pooled approach does not significantly distort the results observed for an individually perturbed gene.
“This finding indicates that multiplexing perturbations in the pooled approach does not significantly distort the results observed for an individually perturbed gene.”
Quote in the paperSubstantial claim - Interpretationown data
Despite differences in developmental stage, clinical heterogeneity of ASD, and patient genetic diversity, similar genes and cell types are affected in both the mouse Perturb-Seq analyses and human patient tissue studies.
“This indicates that despite the different developmental stages, high clinical heterogeneity in ASD, and patient genetic diversity, similar genes and cell types can be identified as affected in both our analyses and in studies of human patient tissue.”
Quote in the paperSubstantial claim - Interpretationown data
In vivo Perturb-Seq can serve as a scalable tool for systems genetic studies of large gene panels, revealing cell-intrinsic functions at single-cell resolution in complex tissues.
“In vivo Perturb-Seq can serve as a scalable tool for systems genetic studies of large gene panels to reveal their cell-intrinsic functions at single-cell resolution in complex tissues.”
Quote in the paperSubstantial claim- Rests on an assumption: The cellular and molecular substrates of ASD/ND are not yet defined.
- Rests on an assumption: A large number of highly penetrant de novo risk genes implicated through human genetic studies provides an entry point to identify the cell types, developmental events, and mechanisms underlying ASD/ND.
- Rests on an assumption: Scalable methods are required to define the function of risk-associated genes with cell-type specificity.
- Interpretationown data
The Perturb-Seq experiments can identify gene program abnormalities that are seen in human ASD patients.
“Although these analyses are limited by the relatively few available datasets of ASD patient brain samples, they suggest that our Perturb-Seq experiments can identify gene program abnormalities seen in human ASD patients.”
Quote in the paperSubstantial claim - Study design and conductown data
The result was further investigated and validated by examining oligodendrocyte development in a Chd8 germline heterozygous mutant model using several orthogonal methods.
“We further investigated and validated this result by examining oligodendrocyte development in a Chd8 germline heterozygous mutant model (as homozygous mutation is embryonic lethal (29)), using several orthogonal methods.”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [29] Nishiyama M, Early embryonic death in mice lacking the beta-catenin-binding protein Duplin. Mol Cell Biol 24, 8386– 8394 (2004). 15367660 PMC516734 source
Not checked in the free run.
- Study design and conductown data
The human snRNA-seq differential expression analysis used a statistically conservative pseudobulk-based approach with DESeq2, correcting for age, sex, and patient-to-patient variability.
“Using a dataset of snRNA-seq profiles from 15 ASD donors and 16 controls (31), we defined differentially expressed (DE) genes in each cell type using a statistically conservative pseudobulk-based analysis with DESeq2 (35, 36), correcting for age, sex, and patient-to-patient variability.”
Quote in the paperSubstantial claim0 of 3 sources checked- cites [31] Velmeshev D, Single-cell genomics identifies cell type-specific molecular changes in autism. Science 364, 685– 689 (2019). 31097668 PMC7678724 source
Not checked in the free run.
- cites [35] Love MI, Huber W, Anders S, Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol 15, 550 (2014). 25516281 PMC4302049 source
Not checked in the free run.
- cites [36] Lun ATL, Marioni JC, Overcoming confounding plate effects in differential expression analyses of single-cell RNA-seq data. Biostatistics 18, 451– 464 (2017). 28334062 PMC5862359 source
Not checked in the free run.
- May combine two claims. 29
- Classified as probably not a substantial claim. 22
- The quote only partly states this claim. 1
May combine two claims.
FindingcitationThe selected ASD/ND risk genes are expressed in human brain tissue, with some highly expressed at embryonic stages and others from early postnatal to adult stages.
“These ASD/ND risk genes are expressed in human brain tissue, as assessed by the BrainSpan bulk RNA-seq dataset (9); some are highly expressed at embryonic stages, and others highly expressed from early postnatal to adult stages (fig. S1B).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [9] Miller JA, Transcriptional landscape of the prenatal human brain. Nature 508, 199– 206 (2014). 24695229 PMC4105188
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
Findingown dataA median of 338 cells per perturbation was found.
“We found a median of 338 cells per perturbation: after excluding perturbations with <70 perturbed cells, we retained 35 ASD/ND risk gene perturbations.”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataSST in interneurons showed decreased expression in ASD patients and was significantly decreased across the panel of 35 ASD risk gene perturbations at FDR < 0.1.
“From this analysis, we identified two genes, SST in interneurons and NRN1 in excitatory neurons, both of which showed decreased expression in ASD patients and were likewise significantly decreased in expression across our panel of perturbations (FDR < 0.1), albeit with different effect sizes (Fig. 4F, Table S10).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [31] Velmeshev D, Single-cell genomics identifies cell type-specific molecular changes in autism. Science 364, 685– 689 (2019). 31097668 PMC7678724 source
Not checked in the free run.
May combine two claims.
Findingown dataNRN1 in excitatory neurons showed decreased expression in ASD patients and was significantly decreased across the panel of 35 ASD risk gene perturbations at FDR < 0.1.
“From this analysis, we identified two genes, SST in interneurons and NRN1 in excitatory neurons, both of which showed decreased expression in ASD patients and were likewise significantly decreased in expression across our panel of perturbations (FDR < 0.1), albeit with different effect sizes (Fig. 4F, Table S10).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [31] Velmeshev D, Single-cell genomics identifies cell type-specific molecular changes in autism. Science 364, 685– 689 (2019). 31097668 PMC7678724 source
Not checked in the free run.
May combine two claims.
Findingown dataAnk2-perturbed cells were present across all cell types and overall proportions of cells were not significantly changed.
“Ank2-perturbed cells were present across all cell types and overall proportions of cells were not significantly changed (fig. S10B).”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataThe expression of each module was largely conserved in all human datasets, with different modules showing distinct levels of conservation of expression in each dataset.
“The expression of each module was largely conserved in all human datasets, with different modules showing distinct levels of conservation of expression in each dataset (Fig. 4A).”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataModule PN2 is associated with genes involved in neurite development and varied across cells in multiple projection neuron subclusters.
“For example, module PN2 is associated with genes involved in neurite development and varied across cells in multiple projection neuron subclusters (fig. S6A).”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataThe ODC1 module is highly expressed in cycling cells and oligodendrocyte precursor cells (OPC), and lowly expressed in committed oligodendrocyte progenitor cells (COP) and newly formed oligodendrocytes (NFOL).
“The ODC1 module is highly expressed in cycling cells and oligodendrocyte precursor cells (OPC), and lowly expressed in committed oligodendrocyte progenitor cells (COP) and newly formed oligodendrocytes (NFOL), suggesting that this module is linked to oligodendrocyte maturation (Fig. 3A)”
Quote in the paperSubstantial claimMay combine two claims.
FindingcitationThe results showed that Chd8 modulates gene modules for oligodendrocyte differentiation and maturation, consistent with prior ChIP-Seq findings that CHD8 interacts directly with OPC maturation genes at perinatal stages.
“Our results showed that Chd8 modulates gene modules for oligodendrocyte differentiation and maturation, consistent with previously reported ChIP-Seq results showing that CHD8 interacts directly with OPC maturation genes at perinatal stages of development (28, 43).”
Quote in the paperNot a substantial claim0 of 2 sources checked- cites [28] Marie C, Oligodendrocyte precursor survival and differentiation requires chromatin remodeling by Chd7 and Chd8. Proc Natl Acad Sci U S A, (2018). PMC6126750 30108144 source
Not checked in the free run.
- cites [43] Zhao C, Dual Requirement of CHD8 for Chromatin Landscape Establishment and Histone Methyltransferase Recruitment to Promote CNS Myelination and Repair. Dev Cell 45, 753– 768 e758 (2018). 29920279 PMC6063525 source
Not checked in the free run.
May combine two claims. The quote only partly states this claim.
Study design and conductown dataThe approach examined effects of each perturbation across a wide range of cell types from distinct brain regions under sparse labeling conditions where less than 0.1% of cortical cells were perturbed, making it highly unlikely that development of individual perturbed cells was affected by perturbed neighbors.
“under sparse labeling conditions where less than 0.1% of cells in the cortex were perturbed, and thus development of individual perturbed cells is highly unlikely to be affected by perturbed neighbors (fig. S3A–C).”
Quote in the paperSubstantial claimMay combine two claims.
PremisecitationFocusing on gene modules instead of individual genes provides more statistical power to detect biologically meaningful perturbation effects with fewer cells and captures diversity within and across cell types.
“As previous work has shown (11–13, 19), focusing on gene modules instead of individual genes provides more statistical power to detect biologically-meaningful perturbation effects using fewer cells than would be required for single gene-level analysis, and can capture diversity both within and across cell types.”
Quote in the paperSubstantial claim0 of 4 sources checked- cites [11] Adamson B, A Multiplexed Single-Cell CRISPR Screening Platform Enables Systematic Dissection of the Unfolded Protein Response. Cell 167, 1867– 1882 e1821 (2016). 27984733 PMC5315571 source
Not checked in the free run.
- cites [12] Dixit A, Perturb-Seq: Dissecting Molecular Circuits with Scalable Single-Cell RNA Profiling of Pooled Genetic Screens. Cell 167, 1853– 1866 e1817 (2016). 27984732 PMC5181115 source
Not checked in the free run.
- cites [13] Jaitin DA, Dissecting Immune Circuits by Linking CRISPR-Pooled Screens with Single-Cell RNA-Seq. Cell 167, 1883– 1896 e1815 (2016). 27984734 source
Not checked in the free run.
- cites [19] Duan B, Model-based understanding of single-cell CRISPR screening. Nat Commun 10, 2233 (2019). 31110232 PMC6527552 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductown dataThe control construct was included in the same pool as perturbation vectors to minimize batch-dependent variation.
“To minimize batch-dependent variation, the control construct was included in the same pool as the perturbation vectors (fig. S3F).”
Quote in the paperNot a substantial claimClassified as probably not a substantial claim.
PremisecitationTrio-based whole-exome sequencing analyses have linked many de novo loss-of-function variants to risk of neurodevelopmental conditions, including autism spectrum disorder and neurodevelopmental delay.
“analysis of trio-based whole-exome sequencing (WES) has implicated a large number of de novo loss-of-function variants contributing to risk of neurodevelopmental pathologies, including autism spectrum disorders and neurodevelopmental delay (ASD/ND) (4, 5).”
Quote in the paperSubstantial claim0 of 2 sources checked- cites [4] Satterstrom FK, Novel genes for autism implicate both excitatory and inhibitory cell lineages in risk. bioRxiv, 484113 (2018). source
Not checked in the free run.
- cites [5] Sanders SJ, De novo mutations revealed by whole-exome sequencing are strongly associated with autism. Nature 485, 237– 241 (2012). 22495306 PMC3667984 source
Not checked in the free run.
May combine two claims.
Study design and conductreasoningThe multiplexed design tested cell-autonomous effects of all perturbations against a negative control construct targeting endogenous GFP in Rosa26, controlling for viral infection and other confounders.
“This multiplexed experimental design allowed us to test the cell-autonomous effect of all perturbations against the effect of a negative control construct targeting the endogenous GFP in the Rosa26 locus, thus controlling for effects related to viral infection, among other confounders.”
Quote in the paperSubstantial claimMay combine two claims.
Study design and conductown dataCell survival after FACS was 78%, and frameshift insertion/deletion was confirmed at 40-70% for each gRNA target among infected cells.
“The cell survival rate after FACS was 78%, and we confirmed a 40–70% frameshift insertion/deletion for each gRNA target among the infected cells (fig. S3D–E).”
Quote in the paperNot a substantial claimMay combine two claims.
InterpretationreasoningThese ASD/ND genes could act in many different cell types and temporal frames, requiring scalable methods to test gene function across cell types and developmental events.
“Thus, these ASD/ND genes could, in principle, act in many different cell types and temporal frames, requiring scalable methods to test gene function across a range of cell types and developmental events.”
Quote in the paperSubstantial claim- Rests on an assumption: Few ASD/ND risk genes have been studied in animal or cellular models, leaving their developmental brain function poorly defined.
Classified as probably not a substantial claim. May combine two claims.
PremisecitationASD/ND is a broad group of neurodevelopmental disorders with highly heterogeneous genetic contributions, including hundreds of highly penetrant de novo risk variant genes.
“ASD/ND comprises a broad collection of neurodevelopmental disorders with highly heterogeneous genetic contributions, including hundreds of highly penetrant de novo risk variant genes (6).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [6] Chen JA, Penagarikano O, Belgard TG, Swarup V, Geschwind DH, The emerging picture of autism spectrum disorder: genetics and pathology. Annu Rev Pathol 10, 111– 144 (2015). 25621659 source
Not checked in the free run.
May combine two claims.
Interpretationown dataThe GO-term approach is limited by the large number of tests performed and the limited number of GO terms relevant to developing cortex.
“However, this approach is limited by the large number of tests performed (one test per GO term per cell type per perturbation, for a total of 510,265 tests), as well as the limited number of GO terms relevant to the developing cortex.”
Quote in the paperSubstantial claimMay combine two claims.
InterpretationreasoningThe observed number of convergent modules across perturbations is likely an underestimation, and larger-scale experiments with more cells might reveal more.
“It is likely that this represents an underestimation of the number of convergent modules across perturbations which might be revealed by larger-scale experiments using greater numbers of cells.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationAnk2 encodes an ankyrin protein and is broadly expressed in excitatory and inhibitory neurons and glial cells in the brain.
“Ank2 encodes an ankyrin protein and is expressed broadly in excitatory and inhibitory neurons as well as glial cells in the brain (23).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [23] Saunders A, Molecular Diversity and Specializations among the Cells of the Adult Mouse Brain. Cell 174, 1015– 1030 e1016 (2018). 30096299 PMC6447408 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationStudies of Ank2 loss-of-function suggest it is involved in axonal morphology, connectivity, and calcium signaling in excitatory neurons.
“Studies examining Ank2 loss-of-function suggest that it is involved in axonal morphology, connectivity, and calcium signaling in excitatory neurons (24–27).”
Quote in the paperNot a substantial claim0 of 4 sources checked- cites [24] Scotland P, Zhou D, Benveniste H, Bennett V, Nervous system defects of AnkyrinB (−/−) mice suggest functional overlap between the cell adhesion molecule L1 and 440-kD AnkyrinB in premyelinated axons. J Cell Biol 143, 1305– 1315 (1998). 9832558 PMC2133070 source
Not checked in the free run.
- cites [25] Tuvia S, Buhusi M, Davis L, Reedy M, Bennett V, Ankyrin-B is required for intracellular sorting of structurally diverse Ca2+ homeostasis proteins. J Cell Biol 147, 995– 1008 (1999). 10579720 PMC2169334 source
Not checked in the free run.
- cites [26] Kline CF, Scott J, Curran J, Hund TJ, Mohler PJ, Ankyrin-B regulates Cav2.1 and Cav2.2 channel expression and targeting. J Biol Chem 289, 5285– 5295 (2014). 24394417 PMC3931084 source
Not checked in the free run.
- cites [27] Yang R, ANK2 autism mutation targeting giant ankyrin-B promotes axon branching and ectopic connectivity. Proc Natl Acad Sci U S A 116, 15262– 15271 (2019). 31285321 PMC6660793 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationAnk2 encodes an ankyrin protein and is expressed broadly in excitatory and inhibitory neurons as well as glial cells in the brain.
“Ank2 encodes an ankyrin protein and is expressed broadly in excitatory and inhibitory neurons as well as glial cells in the brain (23).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [23] Saunders A, Molecular Diversity and Specializations among the Cells of the Adult Mouse Brain. Cell 174, 1015– 1030 e1016 (2018). 30096299 PMC6447408 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationAbnormal maturation of oligodendrocytes may be linked to long-lasting changes in neural wiring and brain function.
“Oligodendrocytes modulate and consolidate neural circuit refinement, and abnormal maturation of oligodendrocytes may be linked to long-lasting changes in neural wiring and brain function (38).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [38] Bercury KK, Macklin WB, Dynamics and mechanisms of CNS myelination. Dev Cell 32, 447– 458 (2015). 25710531 PMC6715306 source
Not checked in the free run.
May combine two claims.
PremisecitationAnk2 mutants show misregulation of intracellular calcium homeostasis and calcium channel expression in excitatory neurons.
“Ank2 mutants showed misregulation of intracellular calcium homeostasis and calcium channel expression in excitatory neurons (25, 26), as well as increased axonal branching and ectopic connectivity (27).”
Quote in the paperNot a substantial claim0 of 2 sources checked- cites [25] Tuvia S, Buhusi M, Davis L, Reedy M, Bennett V, Ankyrin-B is required for intracellular sorting of structurally diverse Ca2+ homeostasis proteins. J Cell Biol 147, 995– 1008 (1999). 10579720 PMC2169334 source
Not checked in the free run.
- cites [26] Kline CF, Scott J, Curran J, Hund TJ, Mohler PJ, Ankyrin-B regulates Cav2.1 and Cav2.2 channel expression and targeting. J Biol Chem 289, 5285– 5295 (2014). 24394417 PMC3931084 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationOligodendrocytes modulate and consolidate neural circuit refinement.
“Oligodendrocytes modulate and consolidate neural circuit refinement, and abnormal maturation of oligodendrocytes may be linked to long-lasting changes in neural wiring and brain function (38).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [38] Bercury KK, Macklin WB, Dynamics and mechanisms of CNS myelination. Dev Cell 32, 447– 458 (2015). 25710531 PMC6715306 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationAnkyrin homologs interact with ion channels in many neuronal types.
“Ankyrin homologs interact with ion channels in many neuronal types, and Ankyrin-G has been shown to stabilize GABAergic synapses (37).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [37] Tseng WC, Jenkins PM, Tanaka M, Mooney R, Bennett V, Giant ankyrin-G stabilizes somatodendritic GABAergic synapses through opposing endocytosis of GABAA receptors. Proc Natl Acad Sci U S A 112, 1214– 1219 (2015). 25552561 PMC4313813 source
Not checked in the free run.
May combine two claims.
PremisecitationAnk2 mutants show increased axonal branching and ectopic connectivity.
“Ank2 mutants showed misregulation of intracellular calcium homeostasis and calcium channel expression in excitatory neurons (25, 26), as well as increased axonal branching and ectopic connectivity (27).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [27] Yang R, ANK2 autism mutation targeting giant ankyrin-B promotes axon branching and ectopic connectivity. Proc Natl Acad Sci U S A 116, 15262– 15271 (2019). 31285321 PMC6660793 source
Not checked in the free run.
May combine two claims.
PremisecitationAnk2 loss-of-function causes hypoplasia of the corpus callosum and pyramidal tract and ultimately optic nerve degeneration, suggesting a requirement for Ank2 in maintaining premyelinated axons in excitatory neurons during early neurodevelopment.
“Ank2 loss-of-function results in hypoplasia of the corpus callosum and pyramidal tract, and ultimately optic nerve degeneration (24), suggesting that it is required in the maintenance of premyelinated axons in excitatory neurons in early neurodevelopment.”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [24] Scotland P, Zhou D, Benveniste H, Bennett V, Nervous system defects of AnkyrinB (−/−) mice suggest functional overlap between the cell adhesion molecule L1 and 440-kD AnkyrinB in premyelinated axons. J Cell Biol 143, 1305– 1315 (1998). 9832558 PMC2133070 source
Not checked in the free run.
May combine two claims.
Interpretationown dataThe data indicate that a selected group of perturbations affected recurrent gene modules with cell-type specificity and point to convergent effects across diverse ASD/ND risk genes.
“Collectively, the data indicate that a selected group of perturbations was able to affect recurrent gene modules with cell-type specificity, and point to some convergent effects across diverse ASD/ND risk genes.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductown dataAlternative approaches, including a linear mixed model-based approach and a permutation-based approach, were compared to evaluate the effects of modeling assumptions.
“To evaluate the effects of these assumptions, we also compared alternative approaches, including a linear mixed model-based approach and a permutation-based approach (fig. S9, table S9).”
Quote in the paperNot a substantial claim- The authors state this as an assumption: In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).
Interpretationnone givenThe in vivo Perturb-Seq method can be applied across diverse diseases and tissues.
“In this work, we demonstrated the application of in vivo Perturb-Seq to ASD/ND risk genes in the developing brain; this method can be applied across diverse diseases and tissues.”
Quote in the paperNot a substantial claim- Rests on an assumption: The cellular and molecular substrates of ASD/ND are not yet defined.
- Rests on an assumption: A large number of highly penetrant de novo risk genes implicated through human genetic studies provides an entry point to identify the cell types, developmental events, and mechanisms underlying ASD/ND.
- Rests on an assumption: Scalable methods are required to define the function of risk-associated genes with cell-type specificity.
Classified as probably not a substantial claim.
Study design and conductreasoningCells were down-sampled so that no perturbation had more than twice the median number of cells over all perturbations, to prevent any single perturbation or batch from dominating the linear model.
“To ensure that no single perturbation or batch dominated the linear model, we down-sampled the cells in each cell category such that no perturbation had more than two times the median number of cells over all perturbations.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationAnkyrin-G stabilizes GABAergic synapses.
“Ankyrin homologs interact with ion channels in many neuronal types, and Ankyrin-G has been shown to stabilize GABAergic synapses (37).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [37] Tseng WC, Jenkins PM, Tanaka M, Mooney R, Bennett V, Giant ankyrin-G stabilizes somatodendritic GABAergic synapses through opposing endocytosis of GABAA receptors. Proc Natl Acad Sci U S A 112, 1214– 1219 (2015). 25552561 PMC4313813 source
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductnone givenThe staining, imaging, and quantifications were double-blinded.
“We double-blinded the staining, imaging, and quantifications.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
Interpretationnone givenAlthough the study focused on the perinatal neocortex, in vivo Perturb-Seq can be applied to study gene functions systematically across other tissues and developmental ages, revealing tissue-specific and broadly distributed functions.
“Although we focused on the perinatal neocortex in this study, in vivo Perturb-Seq can be applied to study gene functions systematically across other tissues and developmental ages, to reveal tissue-specific as well as broadly-distributed gene functions.”
Quote in the paperNot a substantial claim- Rests on an assumption: The cellular and molecular substrates of ASD/ND are not yet defined.
Classified as probably not a substantial claim. May combine two claims.
Study design and conductown dataPerturbation barcodes were identified by two complementary methods, and cells were retained only when the 10x and dial-out assignments agreed or when one method assigned a perturbation and the other did not assign any.
“We then only kept cells for which either i) the assigned 10x and dialout perturbations agree or ii) the cell was assigned to a perturbation by one method but not assigned to any perturbation in the other.”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [12] Dixit A, Perturb-Seq: Dissecting Molecular Circuits with Scalable Single-Cell RNA Profiling of Pooled Genetic Screens. Cell 167, 1853– 1866 e1817 (2016). 27984732 PMC5181115 source
Not checked in the free run.
Classified as probably not a substantial claim. May combine two claims.
Study design and conductown dataThe human differential expression analysis used DESeq2 and corrected for sex and age.
“DESeq2 v 1.20.0 (35) was then used to perform differential expression analysis between the ASD patients and the controls, correcting for sex and age.”
Quote in the paperSubstantial claim0 of 1 sources checked- cites [35] Love MI, Huber W, Anders S, Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol 15, 550 (2014). 25516281 PMC4302049 source
Not checked in the free run.
Probably not a substantial claim: may not pass S2 (checkable). May combine two claims.
InterpretationreasoningIn vivo Perturb-Seq can enable discoveries of pathways and cell types affected in heterogeneous genetic pathologies, directing downstream studies and informing refined models for genetic disorders and mechanistic studies.
“In vivo Perturb-Seq can enable discoveries of pathways and cell types affected in heterogenous genetic pathologies, directing downstream studies and informing the development of refined models for genetic disorders and mechanistic studies as we move from genetic variants to function.”
Quote in the paperNot a substantial claim- Rests on an assumption: The cellular and molecular substrates of ASD/ND are not yet defined.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationThe Wnt signaling pathway plays a crucial role in neuronal progenitor proliferation and differentiation in the forebrain.
“One of the risk genes, Chd8, encodes a protein that binds directly to β-catenin to recruit histone proteins and negatively regulates the Wnt signaling pathway, which plays a crucial role in neuronal progenitor proliferation and differentiation in the forebrain (39–42).”
Quote in the paperNot a substantial claim0 of 4 sources checked- cites [39] Platt RJ, Chd8 Mutation Leads to Autistic-like Behaviors and Impaired Striatal Circuits. Cell Rep 19, 335– 350 (2017). 28402856 PMC5455342 source
Not checked in the free run.
- cites [40] Katayama Y, CHD8 haploinsufficiency results in autistic-like phenotypes in mice. Nature 537, 675– 679 (2016). 27602517 source
Not checked in the free run.
- cites [41] Durak O, Chd8 mediates cortical neurogenesis via transcriptional regulation of cell cycle and Wnt signaling. Nat Neurosci 19, 1477– 1488 (2016). 27694995 PMC5386887 source
Not checked in the free run.
- cites [42] Sakamoto I, A novel beta-catenin-binding protein inhibits beta-catenin-dependent Tcf activation and axis formation. J Biol Chem 275, 32871– 32878 (2000). 10921920 source
Not checked in the free run.
Classified as probably not a substantial claim.
Study design and conductown dataLinear regression testing the relationship between perturbations and WGCNA gene scores corrected for batch and number of genes.
“Linear regression was used to test the relationship between perturbations and WGCNA gene scores, correcting for batch and number of genes.”
Quote in the paperSubstantial claimMay combine two claims.
Interpretationnone givenThe approach can uncover the impact of individual disease-associated genes, combinations of genes, and the overall set of processes they affect.
“This approach can uncover both the impact of individual disease-associated genes and of combinations of genes and the overall set of processes that they affect.”
Quote in the paperSubstantial claim- Rests on an assumption: The cellular and molecular substrates of ASD/ND are not yet defined.
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationChd8 encodes a protein that binds directly to β-catenin to recruit histone proteins and negatively regulates the Wnt signaling pathway.
“One of the risk genes, Chd8, encodes a protein that binds directly to β-catenin to recruit histone proteins and negatively regulates the Wnt signaling pathway, which plays a crucial role in neuronal progenitor proliferation and differentiation in the forebrain (39–42).”
Quote in the paperNot a substantial claim0 of 4 sources checked- cites [39] Platt RJ, Chd8 Mutation Leads to Autistic-like Behaviors and Impaired Striatal Circuits. Cell Rep 19, 335– 350 (2017). 28402856 PMC5455342 source
Not checked in the free run.
- cites [40] Katayama Y, CHD8 haploinsufficiency results in autistic-like phenotypes in mice. Nature 537, 675– 679 (2016). 27602517 source
Not checked in the free run.
- cites [41] Durak O, Chd8 mediates cortical neurogenesis via transcriptional regulation of cell cycle and Wnt signaling. Nat Neurosci 19, 1477– 1488 (2016). 27694995 PMC5386887 source
Not checked in the free run.
- cites [42] Sakamoto I, A novel beta-catenin-binding protein inhibits beta-catenin-dependent Tcf activation and axis formation. J Biol Chem 275, 32871– 32878 (2000). 10921920 source
Not checked in the free run.
The paper relies on this without support.
Few ASD/ND risk genes have been studied in animal or cellular models, leaving their developmental brain function poorly defined.
“Few of these risk genes have been studied in animal or cellular models, and thus their function during brain development remains poorly defined.”
The paper relies on this without support.
Multiple ASD/ND loss-of-function risk gene mutations rarely co-occur in patients.
“As it is rare for multiple ASD/ND loss-of-function risk gene mutations to co-occur in patients”
The authors state this as an assumption.
In the linear model, module expression in individual cells is independent after conditioning on experimental batch, and noise is normally distributed.
“Our modeling approach assumes that module expression in individual cells is independent after conditioning on the experimental batch, and that noise is normally distributed.”
The paper relies on this without support.
The cellular and molecular substrates of ASD/ND are not yet defined.
“ASD/ND affects brain function profoundly, but its cellular and molecular substrates are not yet defined.”
The paper relies on this without support.
A large number of highly penetrant de novo risk genes implicated through human genetic studies provides an entry point to identify the cell types, developmental events, and mechanisms underlying ASD/ND.
“The large number of highly penetrant de novo risk genes implicated through human genetic studies offers an entry point to identify the cell types, developmental events, and mechanisms underlying ASD/ND.”
The paper relies on this without support.
Scalable methods are required to define the function of risk-associated genes with cell-type specificity.
“However, this requires scalable methods to define the function of risk-associated genes with cell-type specificity.”
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
Existing methods that perform perturbation-effect analysis in single-cell omics data do not achieve guaranteed confounder identification, which is a necessary condition for interpretable causal-effect estimation.
“There are a number of existing methods that perform perturbation-effect analysis in single-cell omics data, but none of them achieve guaranteed confounder identification, which is a necessary condition for interpretable causal-effect estimation.”
From Causal identification of single-cell experimental perturbation effects with CINEMA-OT (2023) · cites this paper as [7]
CRISPR-Cas gene editing coupled with single-cell transcriptome readouts enables pooled genetic perturbation experiments in vivo.
“Furthermore, CRISPR–Cas gene editing coupled with single-cell transcriptome readouts15–17 enables pooled genetic perturbation experiments in vivo18.”
From Inferring and perturbing cell fate regulomes in human brain organoids (2023) · cites this paper as [18]
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
Each contradiction between two passages listed here was confirmed by an independent check.
Cell total is unchanged after additional filtering
MinorThe five listed cell-class counts sum to 35,857, exactly the number said to remain after further low-quality cells were removed. The stated filtering therefore produces no reduction from the listed counts.
“We further filtered out some remaining low-quality cells in these five major cell classes, retaining 35,857 high-quality cells”
“cortical projection neurons (8,450 cells), cortical inhibitory neurons (5,532 cells), astrocytes (9,526 cells), oligodendrocytes (4,279 cells), and microglia/macrophages (8,070 cells)”
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“Ank2 encodes an ankyrin protein and is expressed broadly in excitatory and inhibitory neurons as well as glial cells in the brain (22).”
BackgroundConserved cell types with divergent features in human versus mouse cortex
Hodge, R. D., Bakken, T. E., Miller, J. A., Smith, K. A., Barkan, E. R., Graybuck, L. T., Close, J. L., Long, B., Johansen, N., Penn, O., et al. (2019) · Nature · doi:10.1038/s41586-019-1506-7 · Cited by 1,775 · cited 3× · see citing works
“Cell type Adult human brain (Hodge et al 2019) (29)”
BackgroundA single-cell survey of the small intestinal epithelium
Haber, A. L., Biton, M., Rogel, N., Herbst, R. H., Shekhar, K., Smillie, C., Burgin, G., Delorey, T. M., Howitt, M. R., Katz, Y., et al. (2017) · Nature · doi:10.1038/nature24489 · Cited by 1,750 · see citing works
Self-citation<b>stm</b> : An <i>R</i> Package for Structural Topic Models
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Three-dimensional intact-tissue sequencing of single-cell transcriptional states
Wang, X., Allen, W. E., Wright, M. A., Sylwestrak, E. L., Samusik, N., Vesuna, S., Evans, K., Liu, C., Ramakrishnan, C., Liu, J., et al. (2018) · Science · doi:10.1126/science.aat5691 · Cited by 1,620 · see citing works
Intra- and Inter-cellular Rewiring of the Human Colon during Ulcerative Colitis
Smillie, C. S., Biton, M., Ordovás-Montañés, J., Sullivan, K. M., Burgin, G., Graham, D. B., Herbst, R. H., Rogel, N., Slyper, M., Waldman, J., et al. (2019) · Cell · doi:10.1016/j.cell.2019.06.029 · Cited by 1,386 · see citing works
Self-citationTranscriptional landscape of the prenatal human brain
Miller, J. A., Ding, S.-L., Sunkin, S. M., Smith, K. A., Ng, L., Szafer, A., Ebbert, A., Riley, Z. L., Royall, J. J., Aiona, K., et al. (2014) · Nature · doi:10.1038/nature13185 · Cited by 1,325 · see citing works
Genome-scale CRISPR-Cas9 Knockout and Transcriptional Activation Screening
Joung, J., Konermann, S., Gootenberg, J. S., Abudayyeh, O. O., Platt, R. J., Brigham, M. D., Sanjana, N. E., & Zhang, F. (2017) · Nature Protocols · doi:10.1038/nprot.2017.016 · Cited by 1,243 · see citing works
Self-citationAdamson, B., Norman, T. M., Jost, M., Cho, M. Y., Nuñez, J. K., Chen, Y., Villalta, J. E., Gilbert, L. A., Horlbeck, M. A., Hein, M. Y., et al. (2016) · Cell · doi:10.1016/j.cell.2016.11.048 · Cited by 1,215 · see citing works
Self-citationNeuronal Subtype-Specific Genes that Control Corticospinal Motor Neuron Development In Vivo
Arlotta, P., Molyneaux, B. J., Chen, J., Inoue, J., Kominami, R., & Macklis, J. D. (2005) · Neuron · doi:10.1016/j.neuron.2004.12.036 · Cited by 1,157 · see citing works
Self-citationShared molecular neuropathology across major psychiatric disorders parallels polygenic overlap
Gandal, M. J., Haney, J. R., Parikshak, N. N., Leppa, V., Ramaswami, G., Hartl, C., Schork, A. J., Appadurai, V., Buil, A., Werge, T. M., et al. (2018) · Science · doi:10.1126/science.aad6469 · Cited by 1,095 · see citing works
Self-citationIndividual brain organoids reproducibly form cell diversity of the human cerebral cortex
Velasco, S., Kedaigle, A. J., Simmons, S. K., Nash, A., Rocha, M., Quadrato, G., Paulsen, B., Nguyen, L., Adiconis, X., Regev, A., et al. (2019) · Nature · doi:10.1038/s41586-019-1289-x · Cited by 1,094 · see citing works
Self-citationSpatiotemporal gene expression trajectories reveal developmental hierarchies of the human cortex
Nowakowski, T. J., Bhaduri, A., Pollen, A. A., Alvarado, B., Mostajo-Radji, M. A., Di Lullo, E., Haeussler, M., Sandoval-Espinosa, C., Liu, S. J., Velmeshev, D., et al. (2017) · Science · doi:10.1126/science.aap8809 · Cited by 964 · see citing works
Single-cell genomics identifies cell type–specific molecular changes in autism
Velmeshev, D., Schirmer, L., Jung, D., Haeussler, M., Perez, Y., Mayer, S., Bhaduri, A., Goyal, N., Rowitch, D. H., & Kriegstein, A. R. (2019) · Science · doi:10.1126/science.aav8130 · Cited by 927 · see citing works
Advancing the understanding of autism disease mechanisms through genetics
de la Torre-Ubieta, L., Won, H., Stein, J. L., & Geschwind, D. H. (2016) · Nature Medicine · doi:10.1038/nm.4071 · Cited by 827 · see citing works
Dissecting Immune Circuits by Linking CRISPR-Pooled Screens with Single-Cell RNA-Seq
Jaitin, D. A., Weiner, A., Yofe, I., Lara-Astiaso, D., Keren-Shaul, H., David, E., Salame, T. M., Tanay, A., van Oudenaarden, A., & Amit, I. (2016) · Cell · doi:10.1016/j.cell.2016.11.039 · Cited by 777 · see citing works
β-catenin is critical for dendritic morphogenesis
Yu, X., & Malenka, R. C. (2003) · Nature Neuroscience · doi:10.1038/nn1132 · Cited by 454 · see citing works
CHD8 haploinsufficiency results in autistic-like phenotypes in mice
Katayama, Y., Nishiyama, M., Shoji, H., Ohkawa, Y., Kawamura, A., Sato, T., Suyama, M., Takumi, T., Miyakawa, T., & Nakayama, K. I. (2016) · Nature · doi:10.1038/nature19357 · Cited by 353 · see citing works
Single-cell multiomics sequencing and analyses of human colorectal cancer
Bian, S., Hou, Y., Zhou, X., Li, X.-L., Yong, J., Wang, Y., Wang, W.-D., Yan, J., Hu, B.-Q., Guo, H.-S., et al. (2018) · Science · doi:10.1126/science.aao3791 · Cited by 326 · see citing works
Dynamics and Mechanisms of CNS Myelination
Bercury, K. K., & Macklin, W. B. (2015) · Developmental Cell · doi:10.1016/j.devcel.2015.01.016 · Cited by 320 · see citing works
Rubin, A. J., Parker, K. R., Satpathy, A. T., Qi, Y., Wu, B., Ong, A. J., Mumbach, M. R., Ji, A. L., Kim, D. S., Cho, S. W., et al. (2019) · Cell · doi:10.1016/j.cell.2018.11.022 · Cited by 314 · see citing works
Role of β-Catenin in Synaptic Vesicle Localization and Presynaptic Assembly
Bamji, S. X., Shimazu, K., Kimes, N. E., Huelsken, J., Birchmeier, W., Lu, B., & Reichardt, L. F. (2003) · Neuron · doi:10.1016/s0896-6273(03)00718-9 · Cited by 310 · see citing works
The Emerging Picture of Autism Spectrum Disorder: Genetics and Pathology
Chen, J. A., Peñagarikano, O., Belgard, T. G., Swarup, V., & Geschwind, D. H. (2015) · Annual Review of Pathology: Mechanisms of Disease · doi:10.1146/annurev-pathol-012414-040405 · Cited by 254 · see citing works
Chd8 mediates cortical neurogenesis via transcriptional regulation of cell cycle and Wnt signaling
Durak, O., Gao, F., Kaeser-Woo, Y. J., Rueda, R., Martorell, A. J., Nott, A., Liu, C. Y., Watson, L. A., & Tsai, L.-H. (2016) · Nature Neuroscience · doi:10.1038/nn.4400 · Cited by 243 · see citing works
Chd8 Mutation Leads to Autistic-like Behaviors and Impaired Striatal Circuits
Platt, R. J., Zhou, Y., Slaymaker, I. M., Shetty, A. S., Weisbach, N. R., Kim, J.-A., Sharma, J., Desai, M., Sood, S., Kempton, H. R., et al. (2017) · Cell Reports · doi:10.1016/j.celrep.2017.03.052 · Cited by 240 · see citing works
Self-citationSkin-resident innate lymphoid cells converge on a pathogenic effector state
Bielecki, P., Riesenfeld, S. J., Hütter, J.-C., Triglia, E. T., Kowalczyk, M. S., Ricardo-González, R. R., Lian, M., Vesely, M. C. A., Kroehling, L., Xu, H. Y., et al. (2021) · Nature · doi:10.1038/s41586-021-03188-w · Cited by 218 · see citing works
Self-citationScotland, P., Zhou, D., Benveniste, H., & Bennett, V. (1998) · The Journal of Cell Biology · doi:10.1083/jcb.143.5.1305 · Cited by 185 · see citing works
Unifying Views of Autism Spectrum Disorders: A Consideration of Autoregulatory Feedback Loops
Mullins, C., Fishell, G., & Tsien, R. W. (2016) · Neuron · doi:10.1016/j.neuron.2016.02.017 · Cited by 167 · see citing works
Overcoming confounding plate effects in differential expression analyses of single-cell RNA-seq data
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“Our results showed that Chd8 modulates gene programs for oligodendrocyte differentiation and maturation, consistent with previously reported ChIP-Seq results showing that CHD8 interacts directly with OPC maturation genes in the neonatal stage (Marie et al., 2018; Zhao et al., 2018).”
ResultSelf-citationAnkyrin-B Is Required for Intracellular Sorting of Structurally Diverse Ca2+ Homeostasis Proteins
Tuvia, S., Buhusi, M., Davis, L., Reedy, M., & Bennett, V. (1999) · The Journal of Cell Biology · doi:10.1083/jcb.147.5.995 · Cited by 127 · see citing works
Gao, X., Arlotta, P., Macklis, J. D., & Chen, J. (2007) · Journal of Neuroscience · doi:10.1523/jneurosci.3206-07.2007 · Cited by 108 · see citing works
Self-citationA Novel β-Catenin-binding Protein Inhibits β-Catenin-dependent Tcf Activation and Axis Formation
Sakamoto, I., Kishida, S., Fukui, A., Kishida, M., Yamamoto, H., Hino, S.-I., Michiue, T., Takada, S., Asashima, M., & Kikuchi, A. (2000) · Journal of Biological Chemistry · doi:10.1074/jbc.m004089200 · Cited by 106 · see citing works
Marie, C., Clavairoly, A., Frah, M., Hmidan, H., Yan, J., Zhao, C., Van Steenwinckel, J., Daveau, R., Zalc, B., Hassan, B., et al. (2018) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1802620115 · Cited by 103 · see citing works
Yang, R., Walder-Christensen, K. K., Kim, N., Wu, D., Lorenzo, D. N., Badea, A., Jiang, Y.-H., Yin, H. H., Wetsel, W. C., & Bennett, V. (2019) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1904348116 · Cited by 99 · see citing works
Model-based understanding of single-cell CRISPR screening
Duan, B., Zhou, C., Zhu, C., Yu, Y., Li, G., Zhang, S., Zhang, C., Ye, X., Ma, H., Qu, S., et al. (2019) · Nature Communications · doi:10.1038/s41467-019-10216-x · Cited by 98 · see citing works
Tseng, W. C., Jenkins, P. M., Tanaka, M., Mooney, R., & Bennett, V. (2015) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1417989112 · Cited by 90 · see citing works
Early Embryonic Death in Mice Lacking the β-Catenin-Binding Protein Duplin
Nishiyama, M., Nakayama, K., Tsunematsu, R., Tsukiyama, T., Kikuchi, A., & Nakayama, K. I. (2004) · Molecular and Cellular Biology · doi:10.1128/mcb.24.19.8386-8394.2004 · Cited by 86 · see citing works
Dscam Expression Levels Determine Presynaptic Arbor Sizes in Drosophila Sensory Neurons
Kim, J. H., Wang, X., Coolon, R., & Ye, B. (2013) · Neuron · doi:10.1016/j.neuron.2013.05.020 · Cited by 86 · see citing works
Mancinelli, S., & Lodato, S. (2018) · Current Opinion in Neurobiology · doi:10.1016/j.conb.2018.08.001 · Cited by 36 · see citing works
Self-citationAnkyrin-B Regulates Cav2.1 and Cav2.2 Channel Expression and Targeting
Kline, C. F., Scott, J., Curran, J., Hund, T. J., & Mohler, P. J. (2014) · Journal of Biological Chemistry · doi:10.1074/jbc.m113.523639 · Cited by 33 · see citing works
Bielecki, P., Riesenfeld, S. J., Kowalczyk, M. S., et al. (2018) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.1101/461228 · Cited by 21 · see citing works
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Inferring and perturbing cell fate regulomes in human brain organoids
Fleck, J. S., Jansen, S. M. J., Wollny, D., Zenk, F., Seimiya, M., Jain, A., Okamoto, R., Santel, M., He, Z., Camp, J. G., et al. (2023) · Nature · doi:10.1038/s41586-022-05279-8 · Cited by 315 · see citing works
Single-cell and spatial transcriptomics: deciphering brain complexity in health and disease
Piwecka, M., Rajewsky, N., & Rybak-Wolf, A. (2023) · Nature Reviews Neurology · doi:10.1038/s41582-023-00809-y · Cited by 320 · see citing works
A new era in functional genomics screens
Przybyla, L., & Gilbert, L. A. (2022) · Nature Reviews Genetics · doi:10.1038/s41576-021-00409-w · Cited by 281 · see citing works
Impact of the Human Cell Atlas on medicine
Rood, J. E., Maartens, A., Hupalowska, A., Teichmann, S. A., & Regev, A. (2022) · Nature Medicine · doi:10.1038/s41591-022-02104-7 · Cited by 222 · see citing works
Self-citationRood, J. E., Hupalowska, A., & Regev, A. (2024) · Cell · doi:10.1016/j.cell.2024.07.035 · Cited by 139 · see citing works
Self-citationScalable genetic screening for regulatory circuits using compressed Perturb-seq
Yao, D., Binan, L., Bezney, J., Simonton, B., Freedman, J., Frangieh, C. J., Dey, K., Geiger-Schuller, K., Eraslan, B., Gusev, A., et al. (2024) · Nature Biotechnology · doi:10.1038/s41587-023-01964-9 · Cited by 101 · see citing works
Self-citationCausal identification of single-cell experimental perturbation effects with CINEMA-OT
Dong, M., Wang, B., Wei, J., de O. Fonseca, A. H., Perry, C. J., Frey, A., Ouerghi, F., Foxman, E. F., Ishizuka, J. J., Dhodapkar, R. M., et al. (2023) · Nature Methods · doi:10.1038/s41592-023-02040-5 · Cited by 94 · see citing works
Nicheformer: a foundation model for single-cell and spatial omics
Tejada-Lapuerta, A., Schaar, A. C., Gutgesell, R., Palla, G., Halle, L., Minaeva, M., Vornholz, L., Dony, L., Drummer, F., Richter, T., et al. (2025) · Nature Methods · doi:10.1038/s41592-025-02814-z · Cited by 100 · see citing works
Time-resolved single-cell RNA-seq using metabolic RNA labelling
Erhard, F., Saliba, A.-E., Lusser, A., Toussaint, C., Hennig, T., Prusty, B. K., Kirschenbaum, D., Abadie, K., Miska, E. A., Friedel, C. C., et al. (2022) · Nature Reviews Methods Primers · doi:10.1038/s43586-022-00157-z · Cited by 73 · see citing works
In vivo locus-specific editing of the neuroepigenome
Yim, Y. Y., Teague, C. D., & Nestler, E. J. (2020) · Nature Reviews Neuroscience · doi:10.1038/s41583-020-0334-y · Cited by 76 · see citing works
Multiplexed, image-based pooled screens in primary cells and tissues with PerturbView
Kudo, T., Meireles, A. M., Moncada, R., Chen, Y., Wu, P., Gould, J., Hu, X., Kornfeld, O., Jesudason, R., Foo, C., et al. (2025) · Nature Biotechnology · doi:10.1038/s41587-024-02391-0 · Cited by 74 · see citing works
Self-citationTeter, O. M., McQuade, A., Hagan, V., Liang, W., Dräger, N. M., Sattler, S. M., Holmes, B. B., Castillo, V. C., Papakis, V., Leng, K., et al. (2025) · Molecular Psychiatry · doi:10.1038/s41380-025-02997-z · Cited by 38 · see citing works
Zhou, Y., Luo, K., Liang, L., Chen, M., & He, X. (2023) · Nature Methods · doi:10.1038/s41592-023-02017-4 · Cited by 34 · see citing works
PerturbNet predicts single-cell responses to unseen chemical and genetic perturbations
Yu, H., Qian, W., Song, Y., & Welch, J. D. (2025) · Molecular Systems Biology · doi:10.1038/s44320-025-00131-3 · Cited by 57 · see citing works
Barry, T., Mason, K., Roeder, K., & Katsevich, E. (2024) · Genome Biology · doi:10.1186/s13059-024-03254-2 · Cited by 28 · see citing works
PerturbNet predicts single-cell responses to unseen chemical and genetic perturbations
Yu, H., & Welch, J. D. (2022) · doi:10.1101/2022.07.20.500854 · Cited by 57 · see citing works
“Perturb-seq measures cellular responses at single-cell 43 resolution, revealing how cell states are impacted by genetic perturbations, and has been utilized 44 for many biomedical applications [5, 6, 7, 8, 9].”
Background
- Authors
- Jin, X., Simmons, S. K., Guo, A., Shetty, A. S., Ko, M., Nguyen, L., Jokhi, V., Robinson, E., Oyler, P., Curry, N., Deangeli, G., Lodato, S., Levin, J. Z., Regev, A., Zhang, F., & Arlotta, P.
- Affiliations
- Broad Institute (US)
- Harvard University (US)
- Broad Institute of MIT and Harvard, Cambridge, MA, USA.
- Department of Stem Cell and Regenerative Biology, Harvard University, MA, USA.
- Humanitas University (IT)
- Corresponding author
- Jin, Xin
- Regev, Aviv
- Zhang, Feng
- Arlotta, Paola
- ORCID
- Venue
- Science
- Published
- 2020-11-27
- Volume
- 370
- Issue
- 6520
- ISSN
- 0036-8075 (ISSN-L)
- 1095-9203
- PMC
- PMC7985844
- OpenAlex
- W3110610967
- Semantic Scholar
- ccebffce65b9879271b25238c491376d9c2ad3cb
- Cited by
- 317
- References
- 60
- Citations by year
- 2026: 58 · 2025: 60 · 2024: 51 · 2023: 50 · 2022: 50 · 2021: 35 · 2020: 13
- FWCI
- 19.89
- Type
- journal article
- Language
- en
- Open access
- green
- doi.org
- Licence
- cc-by-nc-nd
- Locations
- Science (publishedVersion)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc-sa)
- Funders
- National Institutes of Health (1R01-HG009761, 1R01-MH110049, 1DP1-HL141201, U01MH115727, R01MH096066, P50MH094271)
- Howard Hughes Medical Institute
- National Human Genome Research Institute
- Brain and Behavior Research Foundation
- Stanley Center for Psychiatric Research, Broad Institute
- Klarman Cell Observatory, Broad Institute
- Hock E. Tan and K. Lisa Yang Center for Autism Research at MIT
- New York Stem Cell Foundation
- G. Harold and Leila Y. Mathers Foundation
- Poitras Center for Affective Disorders Research at MIT
- Harvard William F. Milton Grant
- Topics
- Life Sciences › Neuroscience › Cognitive Neuroscience › Autism Spectrum Disorder Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genetics and Neurodevelopmental Disorders
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Keywords
- autism spectrum disorder, neurodevelopmental delay, autism risk genes, mouse brain development, single-cell RNA sequencing, neuronal cell types, glial cell types
- MeSH terms
- Animals, Autistic Disorder, Brain, DNA-Binding Proteins, Humans, Neuroglia, Neurons, Repressor Proteins, Risk, Transcription Factors, Frameshift Mutation, Ankyrins, Gene Expression Profiling, Mice, Genetic Loci, CRISPR-Cas Systems
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-05.
Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-05.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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