A CRISPRi/a platform in human iPSC-derived microglia uncovers regulators of disease states
Nina M Dräger1, Sydney M Sattler1, Cindy Tzu-Ling Huang2
1Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA.
Abstract:
Microglia are emerging as key drivers of neurological diseases. However, we lack a systematic understanding of the underlying mechanisms. Here, we present a screening platform to systematically elucidate functional consequences of genetic perturbations in human induced pluripotent stem cell-derived microglia. We developed an efficient 8-day protocol for the generation of microglia-like cells based on the inducible expression of six transcription factors. We established inducible CRISPR interference and activation in this system and conducted three screens targeting the 'druggable genome'. These screens uncovered genes controlling microglia survival, activation and phagocytosis, including neurodegeneration-associated genes. A screen with single-cell RNA sequencing as the readout revealed that these microglia adopt a spectrum of states mirroring those observed in human brains and identified regulators of these states. A disease-associated state characterized by osteopontin (SPP1) expression was selectively depleted by colony-stimulating factor-1 (CSF1R) inhibition. Thus, our platform can systematically uncover regulators of microglial states, enabling their functional characterization and therapeutic targeting.
Insights
Researchers developed a new platform to study human microglia, uncovering key genes involved in neurological diseases and identifying potential therapeutic targets for better brain health.
Area of Science:
- Neuroscience
- Immunology
- Stem Cell Biology
Background:
- Microglia are crucial in neurological diseases, but their mechanisms remain unclear.
- Understanding genetic influences on microglia function is essential for disease intervention.
Purpose of the Study:
- To develop a platform for systematic functional screening of human microglia.
- To identify genes regulating microglial survival, activation, and phagocytosis.
- To characterize microglial states and their regulators in neurological contexts.
Main Methods:
- Generated human induced pluripotent stem cell-derived microglia in 8 days using inducible transcription factors.
- Established inducible CRISPR interference and activation for genetic screens.
- Conducted screens targeting the druggable genome and utilized single-cell RNA sequencing.
Main Results:
- Identified genes controlling microglial survival, activation, and phagocytosis, including those linked to neurodegeneration.
- Demonstrated that screened microglia adopt diverse states resembling those in human brains.
- Discovered regulators of these microglial states and found that a disease-associated state (SPP1+) is sensitive to CSF1R inhibition.
Conclusions:
- The developed platform enables systematic functional elucidation of genetic perturbations in human microglia.
- This system facilitates the identification and functional characterization of microglial regulators.
- The findings pave the way for targeted therapeutic strategies for neurological diseases by modulating microglial states.
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