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Peer-reviewedCC BY 4.0eLife

Arrayed single-gene perturbations identify drivers of human anterior neural tube closure

  • Roya E Huang1,2,3
  • Giridhar M Anand1,2,3
  • Heitor C Megale1,2,3
  • Jason Chen1,3
  • Chudi Abraham-Igwe3
  • Sharad Ramanathan1,2,3
  1. 1Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, United States
  2. 2School of Engineering and Applied Sciences, Harvard University, Cambridge, United States
  3. 3Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States
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Abstract

Genetic studies of human embryonic morphogenesis are constrained by ethical and practical challenges, restricting insights into developmental mechanisms and disorders. Human pluripotent stem cell (hPSC)-derived organoids provide a powerful alternative for the study of embryonic morphogenesis. However, screening for genetic drivers of morphogenesis in vitro has been infeasible due to organoid variability and the high costs of performing scaled tissue-wide single-gene perturbations. By overcoming both these limitations, we developed a platform that integrates reproducible organoid morphogenesis with uniform single-gene perturbations, enabling high-throughput arrayed CRISPR interference screening in hPSC-derived organoids. To demonstrate the power of this platform, we screened 77 transcription factors in an organoid model of anterior neurulation to identify ZIC2, SOX11, and ZNF521 as essential regulators of neural tube closure. We discovered that ZIC2 and SOX11 are required for closure, while ZNF521 prevents ectopic closure points. Single-cell transcriptomic analysis of perturbed organoids revealed co-regulated gene targets of ZIC2 and SOX11 and an opposing role for ZNF521, suggesting that these transcription factors jointly govern a gene regulatory program driving neural tube closure in the anterior forebrain region. Our single-gene perturbation platform enables high-throughput genetic screening of in vitro models of human embryonic morphogenesis.

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Cite

BibTeXRIS
BibTeX
@article{huang2026arrayed,
  title = {{Arrayed single-gene perturbations identify drivers of human anterior neural tube closure}},
  author = {Roya E Huang and Giridhar M Anand and Heitor C Megale and Jason Chen and Chudi Abraham-Igwe and Sharad Ramanathan},
  journal = {eLife},
  year = {2026},
  month = jul,
  doi = {10.7554/elife.108224},
  url = {https://elifesciences.org/articles/108224},
  note = {CC BY 4.0},
}
RIS
TY  - JOUR
TI  - Arrayed single-gene perturbations identify drivers of human anterior neural tube closure
AU  - Roya E Huang
AU  - Giridhar M Anand
AU  - Heitor C Megale
AU  - Jason Chen
AU  - Chudi Abraham-Igwe
AU  - Sharad Ramanathan
PY  - 2026
DA  - 2026/07/07
JO  - eLife
DO  - 10.7554/elife.108224
UR  - https://elifesciences.org/articles/108224
AB  - Genetic studies of human embryonic morphogenesis are constrained by ethical and practical challenges, restricting insights into developmental mechanisms and disorders. Human pluripotent stem cell (hPSC)-derived organoids provide a powerful alternative for the study of embryonic morphogenesis. However, screening for genetic drivers of morphogenesis in vitro has been infeasible due to organoid variability and the high costs of performing scaled tissue-wide single-gene perturbations. By overcoming both these limitations, we developed a platform that integrates reproducible organoid morphogenesis with uniform single-gene perturbations, enabling high-throughput arrayed CRISPR interference screening in hPSC-derived organoids. To demonstrate the power of this platform, we screened 77 transcription factors in an organoid model of anterior neurulation to identify ZIC2, SOX11, and ZNF521 as essential regulators of neural tube closure. We discovered that ZIC2 and SOX11 are required for closure, while ZNF521 prevents ectopic closure points. Single-cell transcriptomic analysis of perturbed organoids revealed co-regulated gene targets of ZIC2 and SOX11 and an opposing role for ZNF521, suggesting that these transcription factors jointly govern a gene regulatory program driving neural tube closure in the anterior forebrain region. Our single-gene perturbation platform enables high-throughput genetic screening of in vitro models of human embryonic morphogenesis.
SN  - 2050-084X
ER  -