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From the Journals

Brain Cell Networks Mapped with CRISPR

A new Perturb-seq platform utilizes adeno-associated viruses vectors and transposons to examine cell-specific transcriptional networks within the developing brain <em>in vivo</em>.

Conexiant

A new method to study how genes turn on and off in specific cell types of a developing brain, all within a living organism, was developed.

Researchers have developed a novel platform utilizing adeno-associated viruses (AAVs) for rapid, in vivo CRISPR screening, enabling single-cell transcriptomic phenotyping across the brain and peripheral nervous systems from embryonic to adult stages. The study explored the cell-type-specific effects of transcriptional networks in cortical development.

The research, published in Cell, utilized the versatile tropism and labeling capacity of AAVs, facilitating large-scale in vivo CRISPR screening. The team tested 86 vectors across AAV serotypes, combined with a transposon system, increasing labeling efficiency and accelerating in vivo gene delivery from weeks to days. In a proof-of-principle in utero screen, they observed the pleiotropic effects of Foxg1, a transcription factor, on distinct networks involved in the cell fate specification of Layer 6 corticothalamic neurons.

The platform can label more than 6% of cerebral cells, compared to the current state-of-the-art efficacy of less than 0.1% by lentivirus. This capability enables the analysis of over 30,000 cells in a single experiment, supporting massively parallel in vivo Perturb-seq. The platform is compatible with various phenotypic measurements, including single-cell and spatial multi-omics, allowing for comprehensive analysis of gene function across cell types in vivo.

Key Findings:

  • Rapid and efficient gene delivery using AAVs, achieved high expression within two days.
  • Increased expression and perturbation in the brain and peripheral nervous systems using a transposon system.
  • Identification of cell-type-specific effects of Foxg1, leading to hybrid cell states.
  • Ability to label more than 6% of cerebral cells, enabling the analysis of over 30,000 cells in one experiment.

This study presented a novel AAV-based, parallel in vivo Perturb-seq platform with capabilities for high-resolution, high-throughput gene function analysis across various cell types in vivo. 

Funding support was provided by the NIH and the authors reported no conflict of interest.