(A) Mixed population of barcoded HeLa cells expressing 24 FRET (blue) and non-FRET (green) biosensors (Table S1) were stimulated with the indicated activators (2DG, 10 mM 2-deoxyglucose; ANI, 1 μg/mL anisomycin; EGF, 100 ng/mL EGF; ION, 1 μM ionomycin; PDB, 200 nM phorbol-12,13-dibutyrate; and UK, 10 μM UK14304). For each barcode, the activities were normalized to pre-stimulus levels across cells of the corresponding barcode and adjusted for its dynamic range (Table S2). Black boxes indicate expected responses to known activators. Red boxes indicate additional responses.
(B) Individual cell traces of EKAR and ERKKTR activities normalized to pre-stimulus levels. Thick blue lines represent the average activities.
(C) Comparison of activation kinetics of EKAR and ERKKTR obtained from mixed barcoded cells. Activities are scaled to the peak responses and represent the mean of 46 (EKAR) and 22 (EKRKTR) cells.
See also Table S3.