Genome-Scale CRISPR Screens Identify Human Pluripotency-Specific Genes

Area of Science:

• Stem cell biology
• Genetics
• CRISPR screening

Background:

• Human pluripotent stem cells (hPSCs) are valuable for disease modeling and preclinical research.
• Genetic screening in hPSCs is challenging, limiting their application.
• A scalable and renewable system is needed for hPSC genetic screens.

Purpose of the Study:

• To develop an inducible Cas9 and sgRNA-hPSC library for loss-of-function mutations.
• To enable genome-wide CRISPR screens in various hPSC-derived cell types.
• To identify novel regulators of hPSC self-renewal, survival, and differentiation.

Main Methods:

• Development of a scalable and renewable Cas9 and sgRNA-hPSC library.
• Induction of loss-of-function mutations at will.
• Performance of three genome-wide CRISPR screens focusing on hPSC fitness, single-cell survival, and differentiation capacity.

Main Results:

• The library successfully facilitated multiple genome-wide CRISPR screens.
• Identified known regulators of hPSC self-renewal, survival, and differentiation.
• Discovered numerous genes with previously unknown roles in these fundamental hPSC properties.

Conclusions:

• The inducible mutant hPSC library is a powerful resource for genetic screening.
• This approach enhances understanding of human development and genetics.
• It serves as a tool for identifying disease-modifying genes and pathways.