Cited by 556CRISPR and Genetic EngineeringPluripotent Stem Cells ResearchVirus-based gene therapy research

The DOI matches its Crossref record; 67 of the 68 references it cites were found in a registry or an index; none of the 20 sources its claims cite is checked for support yet.

CRISPR Interference-Based Platform for Multimodal Genetic Screens in Human iPSC-Derived Neurons

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Tian, Ruilin, Gachechiladze, Mariam A., Ludwig, Connor H., Laurie, Matthew T., Hong, Jason Y., Nathaniel, Diane, Prabhu, Anika V., Fernandopulle, Michael S., Patel, Rajan, Abshari, Mehrnoosh, Ward, Michael E., Kampmann, Martin·2019·Neuron

DOI 10.1016/j.neuron.2019.07.014PMC PMC6813890

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Abstract

CRISPR/Cas9-based functional genomics have transformed our ability to elucidate mammalian cell biology. However, most previous CRISPR-based screens were conducted in cancer cell lines rather than healthy, differentiated cells. Here, we describe a CRISPR interference (CRISPRi)-based platform for genetic screens in human neurons derived from induced pluripotent stem cells (iPSCs). We demonstrate robust and durable knockdown of endogenous genes in such neurons and present results from three complementary genetic screens. First, a survival-based screen revealed neuron-specific essential genes and genes that improved neuronal survival upon knockdown. Second, a screen with a single-cell transcriptomic readout uncovered several examples of genes whose knockdown had strikingly cell-type-specific consequences. Third, a longitudinal imaging screen detected distinct consequences of gene knockdown on neuronal morphology. Our results highlight the power of unbiased genetic screens in iPSC-derived differentiated cell types and provide a platform for systematic interrogation of normal and disease states of neurons. VIDEO ABSTRACT.

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In the paper
“We demonstrate robust and durable knockdown of endogenous genes in such neurons”
- Also stated in: § Robust CRISPR interference in human iPSC-derived neurons · boosted
  “Our platform thus enables potent CRISPRi knockdown of endogenous genes in iPSC-derived neurons.”
§ CRISPR interference-based platform for multimodal genetic screens in human iPSC-derived neurons > Abstract
Load-bearingStrength:boosted· not yet calibratedCausal:direct· not yet calibrated
Tests✓ Asserted: passes✓ Checkable: passes✓ Load-bearing: passes✓ Based: passes

Show all 62 claims

It may state two separate claims as one. 20
Probably not a substantial claim. 20

None of the sources cited for this claim have been checked yet.
In the paper
“which yields large quantities of highly homogeneous neurons, a prerequisite for robust population-based screens.”
§ CRISPR interference-based platform for multimodal genetic screens in human iPSC-derived neurons > INTRODUCTION
What the sources say
- Not checked yetcites Fernandopulle et al., 2018 source
- Not checked yetcites Wang et al., 2017 source
Load-bearingStrength:plain· not yet calibratedCausal:direct· not yet calibrated
Tests✓ Asserted: passes✓ Checkable: passes✓ Load-bearing: passes✓ Based: passes
It may state two separate claims as one.
None of the sources cited for this claim have been checked yet.
In the paper
“induced expression of different transcription factors yield other types of neurons, such as motor neurons (Hester et al., 2011; Shi et al., 2018) and inhibitory neurons (Yang et al., 2017).”
§ CRISPR interference-based platform for multimodal genetic screens in human iPSC-derived neurons > DISCUSSION
What the sources say
- Not checked yetcites Hester et al., 2011 source
- Not checked yetcites Shi et al., 2018 source
- Not checked yetcites Yang et al., 2017 source
Strength:plain· not yet calibratedCausal:direct· not yet calibrated
Tests✓ Asserted: passes✓ Checkable: passes✗ Load-bearing: fails✓ Based: passes
Probably not a substantial claim. The paper’s conclusions may not depend on it.
In the paper
“Second, it is inducible and reversible (Gilbert et al., 2014)”
§ CRISPR interference-based platform for multimodal genetic screens in human iPSC-derived neurons > DISCUSSION
What the sources say
- Not checked yetcites Gilbert et al., 2014 source
Load-bearingStrength:plain· not yet calibrated
Tests✓ Asserted: passes✓ Checkable: passes✓ Load-bearing: passes✓ Based: passes
It may state two separate claims as one.
None of the sources cited for this claim have been checked yet.
In the paper
“In addition to the Ngn2-driven generation of glutamatergic neurons (Fernandopulle et al., 2018; Wang et al., 2017; Zhang et al., 2013) used here”
§ CRISPR interference-based platform for multimodal genetic screens in human iPSC-derived neurons > DISCUSSION
What the sources say
- Not checked yetcites Fernandopulle et al., 2018 source
- Not checked yetcites Wang et al., 2017 source
- Not checked yetcites Zhang et al., 2013 source
Strength:plain· not yet calibratedCausal:direct· not yet calibrated
Tests✓ Asserted: passes✓ Checkable: passes✗ Load-bearing: fails✓ Based: passes
Probably not a substantial claim. The paper’s conclusions may not depend on it.

36 of 36

The paper relies on this without support.
The AAVS1 locus, where the inducible Ngn2 transgene was integrated, lies within the PPP1R12C gene.
“Interestingly, the AAVS1 locus, into which the inducible Ngn2 transgene was integrated, resides within the PPP1R12C gene.”
The paper relies on this without support.
Primary human neurons are difficult to obtain from human donors and cannot be expanded because they are post-mitotic.
“primary neurons are difficult to obtain from human donors, and non-expandable due to their post-mitotic nature.”
The paper relies on this without support.
Most previous CRISPR-based screens were conducted in cancer cell lines or stem cells rather than healthy differentiated human cells.
“Most previous CRISPR-based screens were implemented in cancer cell lines or stem cells rather than healthy differentiated human cells”
The paper relies on this without support.
CRISPRi reduces gene function by partial knockdown rather than complete knockout, allowing study of essential genes.
“Furthermore, CRISPRi perturbs gene function by partial knockdown, rather than knockout, thereby enabling the investigation of the biology of essential genes.”
The paper relies on this without support.
sgRNA abundance in pooled screens reflects cell survival or proliferation.
“Based on the depletion or enrichment of sgRNAs targeting specific genes at different time points compared to day 0, we identified hit genes for which knockdown was toxic or beneficial to either iPSCs or neurons at different time points (Fig. 2B, Fig. S2B).”
The paper relies on this without support.
iPSCs can be made from readily available cells, genetically engineered, expanded, and differentiated into various cell types.
“They can be made from readily available cells, such as skin fibroblasts or peripheral mononuclear blood cells; they can be genetically engineered and subsequently expanded to generate large numbers of isogenic cells; and they can then be differentiated into a variety of cell types”
The paper relies on this without support.
The presence of astrocytes in co-culture is representative of the brain environment.
“In the brain, many neuronal functions are supported by glial cells, particularly astrocytes.”
The paper relies on this without support.
CRISPR interference reduces gene expression partially rather than completely eliminating it.
“Third, it perturbs gene function via partial knockdown, as opposed to knockout”

8 of 8

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Doxycycline concentration differs by a factor of 1,000
Minor
The neuronal-plating procedures specify 2 μg/μL doxycycline in Cortical Neuron Culture Medium, whereas the formulation of that same medium specifies 2 μg/mL.

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Authors: Tian, R., Gachechiladze, M. A., Ludwig, C. H., Laurie, M. T., Hong, J. Y., Nathaniel, D., Prabhu, A. V., Fernandopulle, M. S., Patel, R., Abshari, M., Ward, M. E., & Kampmann, M.
Affiliations: University of California, San Francisco (US)
National Institutes of Health (US)
Corresponding author: Ward, Michael E.
Kampmann, Martin
ORCID: Tian, Ruilin
Gachechiladze, Mariam A.
Ludwig, Connor H.
Laurie, Matthew T.
Prabhu, Anika V.
Fernandopulle, Michael S.
Patel, Rajan
Abshari, Mehrnoosh
Ward, Michael E.
Kampmann, Martin
Venue: Neuron
Publisher: Elsevier BV
Published: 2019-10
Volume: 104
Issue: 2
Pages: 239-255.e12
ISSN: 0896-6273 (ISSN-L)
DOI: 10.1016/j.neuron.2019.07.014
PMC: PMC6813890
OpenAlex: W2967789674
Cited by: 543
References: 68
Citations by year: 2026: 85 · 2025: 116 · 2024: 103 · 2023: 60 · 2022: 66 · 2021: 62 · 2020: 45 · 2019: 6
FWCI: 15.72
Type: journal article
Language: en
Open access: bronze
doi.org
Licence: www.elsevier.com/tdm/userlicense/1.0/ (tdm)
www.elsevier.com/legal/tdmrep-license (tdm)
www.elsevier.com/open-access/userlicense/1.0/ (vor, 388 d)
Locations: Neuron (publishedVersion)
PubMed (publishedVersion)
PubMed Central (acceptedVersion)
eScholarship (California Digital Library) (submittedVersion)
Funders: National Institute of General Medical Sciences
Paul G. Allen Family Foundation
National Institute of Neurological Disorders and Stroke
National Institute on Aging
Topics: Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Pluripotent Stem Cells Research
Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
Keywords: CRISPR interference, iPSC-derived neurons, functional genetic screens, gene knockdown, neuronal survival, essential genes, single-cell transcriptomics, neuronal morphology
MeSH terms: RNA-Seq, Cell Survival, Humans, Neurons, Microscopy, Confocal, Gene Knockdown Techniques, Induced Pluripotent Stem Cells, Single-Cell Analysis, CRISPR-Cas Systems

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CRISPR Interference-Based Platform for Multimodal Genetic Screens in Human iPSC-Derived Neurons

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