Genome-wide CRISPR Screens in Primary Human T Cells Reveal Key Regulators of Immune Function

Area of Science:

• Immunology
• Molecular Biology
• Genetics

Background:

• Human T cells are critical for immunity and cancer immunotherapy.
• CRISPR screens in primary human T cells are challenging but vital for target discovery.
• Identifying regulators of T cell stimulation is key for therapeutic advancements.

Purpose of the Study:

• To develop a novel CRISPR screening method for primary human T cells.
• To identify genes regulating T cell receptor signaling, proliferation, and anti-cancer activity.
• To explore mediators of immunosuppression in the tumor microenvironment.

Main Methods:

• Developed a method called single guide RNA (sgRNA) lentiviral infection with Cas9 protein electroporation (SLICE).
• Performed genome-wide loss-of-function screens in primary human T cells.
• Utilized SLICE in conjunction with single-cell RNA sequencing (RNA-seq).

Main Results:

• Identified essential T cell receptor signaling components and negative regulators of proliferation.
• Characterized gene perturbations that enhanced cancer cell killing.
• Revealed genes controlling responses to adenosine signaling, a mechanism of immunosuppression.

Conclusions:

• The SLICE platform enables efficient, large-scale CRISPR screening in primary human T cells.
• SLICE facilitates unbiased discovery of functional gene targets for immunotherapy and drug development.
• This method advances the understanding of T cell biology and therapeutic reprogramming.