Genome-wide CRISPRi screen in human iNeurons identifies novel negative mTOR regulator genes associated with focal cortical dysplasia
What this study shows
First unbiased genome-wide functional screen for negative regulators of mTOR signalling carried out in human neurons rather than in cancer lines or non-neural cells. Six genes (LRRC4, EIF3A, TSN, HIP1, PIK3R3, URI1) raise phospho-S6 when knocked down; only PIK3R3 and HIP1 raise phosphorylation across the whole AKT/mTOR/S6 axis, and each of those two has an independently reported candidate pathogenic variant in resected FCD brain tissue, which gives the screen external validity. Boundary: pS6 is a readout of mTORC1 output, not of mTORC1 itself, and four of the six hits did not show pathway-wide hyperphosphorylation, so their placement relative to mTORC1 is unresolved. The screen is in vitro in iPSC-derived neurons; no animal or human causal test of these genes in FCD is included. Relevant to the Atlas as an expansion of the upstream regulator set, and because the GDNF-withdrawal result shows the hits uncouple mTOR output from growth-factor input.
Abstract
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Focal cortical dysplasia (FCD) is a common cause of focal epilepsy that typically results from brain mosaic mutations in the mTOR cell signaling pathway. To identify new potential FCD genes, we developed an in vitro CRISPRi screen in human neurons and used FACS enrichment based on the FCD biomarker, phosphorylated S6 ribosomal protein (pS6). Using whole-genome (110,000 gRNAs) and candidate (129 gRNAs) libraries, we discovered 6 new genes in which loss of function significantly increases pS6 levels: LRRC4, EIF3A, TSN, HIP1, PIK3R3, and URI1.
Read the full abstract on PubMed →At a glance
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| Evidence type | M Molecular — cells, biochemistry, structure Marked M because it is molecular or in-vitro work (model: Human iPSC-derived neurons (genome-wide CRISPRi screen)) rather than a whole-organism health-outcome study. That is often exactly where causal biology gets established -- the code says which system the finding was shown in, and nothing about how good the work is. |
| Study type | 5 - Mechanistic / In Vitro |
| Model system | Human iPSC-derived neurons (genome-wide CRISPRi screen) |
| Journal | Neurobiology of Disease |
| Year | 2026 |
| Peer reviewed | Yes |
| Record last updated | 2026-10-01 |
| Source | DOI 10.1016/j.nbd.2026.107602 · PMID 42731765 |
Where this study fits
How this study connects to the pathway map, open questions, people and other studies in the Atlas.
People in the Atlas
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Stem-cell neuroscientist who builds human neuron and organoid models to find the genes behind brain malformations
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Neurologist and stem-cell researcher who rebuilds human epilepsies in patient-derived neurons to find what goes wrong
More: 4 other authors
No profile yet: Luo J, Walker JC, Takla TN, Carvill GL
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Topics
Extracted findings
These fields were extracted from the paper with AI assistance and have not been checked against the source. Read them as a pointer into the paper, not as curated claims; where they disagree with the curated summary above, the curated summary stands.
| Intervention | CRISPR interference (CRISPRi) knockdown; FACS enrichment on phospho-S6 (pS6); GDNF withdrawal |
| Target | Negative regulators of neuronal mTOR signalling (LRRC4, EIF3A, TSN, HIP1, PIK3R3, URI1) |
| Model | Human iPSC-derived neurons (iNeurons) |
| Effect | A genome-wide CRISPRi screen in human iNeurons identifies six genes whose loss raises pS6; two of them (PIK3R3, HIP1) raise phosphorylation throughout the AKT/mTOR/S6 axis, and knockdown of all six makes mTOR signalling resistant to GDNF withdrawal |
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