Scalable Method for Monitoring Protein Levels in Cells

This case study presents a scalable method for monitoring protein levels and localizations in cells, developed by researchers at the CeMM Research Center for Molecular Medicine. The method utilizes CRISPR-Cas9-based intron tagging to create cell pools expressing hundreds of GFP-fusion proteins. This allows for the monitoring of protein localization changes through time-lapse microscopy and in situ sequencing. The researchers transduced HAP1 cells with an sgRNA library targeting metabolic enzyme genes, generating a pool of GFP-tagged cells. They identified clonal cell lines and performed compound screening with BRD4-targeting PROTAC dBET6, observing both known and novel changes in protein localization. The study concludes that this approach can effectively identify cellular responses to perturbations and can be applied to other gene sets for studying protein dynamics at scale. The findings demonstrate the potential of pooled protein tagging combined with in situ sequencing for high-throughput analysis in cellular biology.