Fig. 1

Experimental setup. (A) After binding to a growth factor (GF), receptor tyrosine kinases (RTKs) multimerize and signal to ERK via RAS, RAF, and MEK (the RTK/ERK pathway). Simultaneously, RTKs trigger the calcium-calcineurin pathway (RTK/calcineurin pathway). (B) The EML4-ALK oncogene suppresses a cell’s ability to respond to environmental cues such as growth factors. The ALK inhibitor (ALKi) restores responsiveness. (C) In normal cells, activity pulses are accurately transmitted by the RTK/ERK pathway from receptors to the nucleus. EML4-ALK-positive cancer cells transmit environmental information with low fidelity, which can (to some extent) be restored by ALKi. (D) Pulsatile optogenetic FGFR (optoFGFR) stimulation and fluorescent biosensors of signaling allow the quantification of information transmission through the RTK/ERK signaling axis. The ERK-KTR biosensor translocates between the nucleus and the cytoplasm upon ERK activation/inactivation. (E) OptoFGFR, in response to blue light, recapitulates endogenous RTK activation. Cell responsiveness to RTK signals may be suppressed by EML4-ALK, which associates with and sequesters GRB2-SOS. Signal transmission can be restored by ALKi, which liberates GRB2-SOS from EML4-ALK. (F) ERK-KTR migrates to the cytoplasm upon phosphorylation by ERK, and in the reverse direction after dephosphorylation by calcineurin. Inhibitors of calcineurin (cyclosporine A) or MEK (trametinib) were used to study the two pathways in isolation. (G) ERK-KTR cytoplasmic–nuclear shuttling after stimulation with a light pulse in BEAS-2B cells. Fluorescence was recorded with a one-minute resolution. Scale bar = 20 μm. (H) Light stimulus leads to ERK-KTR phosphorylation, resulting in cytoplasmic translocation (a peak is observed approximately 7 min after the light pulse), preceded by a calcineurin-mediated ‘dip’ observed 2 min after the light pulse in BEAS-2B cells. The thin gray lines represent single-cell preprocessed ERK-KTR trajectories, and the thick gray line represents their average. The green line shows a selected single-cell trajectory for which the (posterior) probability of a light pulse is given in panel K. See Methods for details of ERK-KTR trajectory preprocessing. (I) The cells were stimulated with a pseudorandom series of light pulses. Information is encoded in intervals between subsequent pulses. (J) The distribution of intervals between stimulation pulses was chosen on the basis of preliminary experiments to maximize the bitrate. (K) Single-cell ERK-KTR trajectories were used for probabilistic reconstruction of the input signal, i.e., to predict the probability of a light pulse occurrence at each time point throughout the experiment at a one-minute temporal resolution. (L) The numerically estimated entropy of the probabilistic reconstruction was used to compute the rate of information transmission through the pathway.