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

Area of Science:

• Cell Biology
• Neuroscience
• Genomics

Background:

• CRISPR/Cas9 technology has revolutionized mammalian cell biology research.
• Previous CRISPR screens primarily used cancer cell lines, limiting insights into healthy, differentiated cells.

Purpose of the Study:

• To develop and validate a CRISPR interference (CRISPRi) platform for genetic screens in human induced pluripotent stem cell-derived neurons (iPSC-neurons).
• To identify neuron-specific essential genes and genes influencing neuronal survival and morphology.
• To uncover cell-type-specific gene functions in human neurons.

Main Methods:

• Established a CRISPR interference (CRISPRi) platform for gene knockdown in iPSC-neurons.
• Conducted three complementary genetic screens: survival-based, single-cell transcriptomic, and longitudinal imaging.
• Validated robust and durable gene knockdown in differentiated human neurons.

Main Results:

• Identified neuron-specific essential genes and genes that enhance neuronal survival.
• Uncovered cell-type-specific consequences of gene knockdown using transcriptomic analysis.
• Observed distinct effects of gene knockdown on neuronal morphology through imaging.

Conclusions:

• Demonstrated the utility of unbiased genetic screens in iPSC-derived differentiated cells.
• Provided a powerful platform for systematic investigation of normal and disease states in human neurons.
• Highlighted the importance of studying gene function in relevant cellular contexts.