RT Journal
A1 Reicher, Andreas
A1 Koren, Anna
A1 Kubicek, Stefan
T1 Pooled protein tagging, cellular imaging, and in situ sequencing for monitoring drug action in real time
JF Genome Research 
JO Genome Research 
YR 2020 
FD December 01 
VO 30 
IS 12 
SP 1846 
OP 1855 
DO 10.1101/gr.261503.120 
UL http://genome.cshlp.org/content/30/12/1846.abstract 
AB The levels and subcellular localizations of proteins regulate critical aspects of many cellular processes and can become targets of therapeutic intervention. However, high-throughput methods for the discovery of proteins that change localization either by shuttling between compartments, by binding larger complexes, or by localizing to distinct membraneless organelles are not available. Here we describe a scalable strategy to characterize effects on protein localizations and levels in response to different perturbations. We use CRISPR-Cas9-based intron tagging to generate cell pools expressing hundreds of GFP-fusion proteins from their endogenous promoters and monitor localization changes by time-lapse microscopy followed by clone identification using in situ sequencing. We show that this strategy can characterize cellular responses to drug treatment and thus identify nonclassical effects such as modulation of protein–protein interactions, condensate formation, and chemical degradation.