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Regulation of EGFR signal transduction by analogue-to-digital conversion in endosomes.

Villaseñor R, Nonaka H, Del Conte-Zerial P, Kalaidzidis Y, Zerial M - Elife (2015)

Bottom Line: By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output.Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation.We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.

ABSTRACT
An outstanding question is how receptor tyrosine kinases (RTKs) determine different cell-fate decisions despite sharing the same signalling cascades. Here, we uncovered an unexpected mechanism of RTK trafficking in this process. By quantitative high-resolution FRET microscopy, we found that phosphorylated epidermal growth factor receptor (p-EGFR) is not randomly distributed but packaged at constant mean amounts in endosomes. Cells respond to higher EGF concentrations by increasing the number of endosomes but keeping the mean p-EGFR content per endosome almost constant. By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output. Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation. We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response.

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The total amount of p-EGFR in endosomes decays with the same kinetics asthe number of endosomes with p-EGFR.Time course of total integral p-EGFR intensity in endosomes (red) andendosomes with p-EGFR (black) per 1000 μm2 of the areacovered by cells (black) after stimulation with 10 ng/ml EGF as in Figure 1. Points show mean ± SEM.All measurements were done in three independent experiments with a total of∼150 cells per time point or condition.DOI:http://dx.doi.org/10.7554/eLife.06156.010
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fig1s6: The total amount of p-EGFR in endosomes decays with the same kinetics asthe number of endosomes with p-EGFR.Time course of total integral p-EGFR intensity in endosomes (red) andendosomes with p-EGFR (black) per 1000 μm2 of the areacovered by cells (black) after stimulation with 10 ng/ml EGF as in Figure 1. Points show mean ± SEM.All measurements were done in three independent experiments with a total of∼150 cells per time point or condition.DOI:http://dx.doi.org/10.7554/eLife.06156.010

Mentions: We next determined the distribution of EGFR and p-EGFR in individual endosomes. Thenumber of endosomes with p-EGFR decayed with similar kinetics as the total p-EGFR signal(τdecay N-p-EGFR = 45.24 ± 11.39 vsτdecay p-EGFR = 30.97 ± 1.69; compare red with blackcurve in Figure 1—figure supplement 6).The mean content of total EGFR per endosome increased over time and then rapidly decayedreaching steady state, due to the balance of continuous EGF uptake and degradation(Figure 1B, green curve). After a rapidincrease, the mean content of p-EGFR in each endosome stabilized to a fairly constantlevel after ∼20 min (Figure 1B, redcurve). Similar results were obtained when EGF was pulsed for 1 min and chased fordifferent periods of time (Figure 1B, blue andblack points).


Regulation of EGFR signal transduction by analogue-to-digital conversion in endosomes.

Villaseñor R, Nonaka H, Del Conte-Zerial P, Kalaidzidis Y, Zerial M - Elife (2015)

The total amount of p-EGFR in endosomes decays with the same kinetics asthe number of endosomes with p-EGFR.Time course of total integral p-EGFR intensity in endosomes (red) andendosomes with p-EGFR (black) per 1000 μm2 of the areacovered by cells (black) after stimulation with 10 ng/ml EGF as in Figure 1. Points show mean ± SEM.All measurements were done in three independent experiments with a total of∼150 cells per time point or condition.DOI:http://dx.doi.org/10.7554/eLife.06156.010
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4384751&req=5

fig1s6: The total amount of p-EGFR in endosomes decays with the same kinetics asthe number of endosomes with p-EGFR.Time course of total integral p-EGFR intensity in endosomes (red) andendosomes with p-EGFR (black) per 1000 μm2 of the areacovered by cells (black) after stimulation with 10 ng/ml EGF as in Figure 1. Points show mean ± SEM.All measurements were done in three independent experiments with a total of∼150 cells per time point or condition.DOI:http://dx.doi.org/10.7554/eLife.06156.010
Mentions: We next determined the distribution of EGFR and p-EGFR in individual endosomes. Thenumber of endosomes with p-EGFR decayed with similar kinetics as the total p-EGFR signal(τdecay N-p-EGFR = 45.24 ± 11.39 vsτdecay p-EGFR = 30.97 ± 1.69; compare red with blackcurve in Figure 1—figure supplement 6).The mean content of total EGFR per endosome increased over time and then rapidly decayedreaching steady state, due to the balance of continuous EGF uptake and degradation(Figure 1B, green curve). After a rapidincrease, the mean content of p-EGFR in each endosome stabilized to a fairly constantlevel after ∼20 min (Figure 1B, redcurve). Similar results were obtained when EGF was pulsed for 1 min and chased fordifferent periods of time (Figure 1B, blue andblack points).

Bottom Line: By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output.Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation.We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.

ABSTRACT
An outstanding question is how receptor tyrosine kinases (RTKs) determine different cell-fate decisions despite sharing the same signalling cascades. Here, we uncovered an unexpected mechanism of RTK trafficking in this process. By quantitative high-resolution FRET microscopy, we found that phosphorylated epidermal growth factor receptor (p-EGFR) is not randomly distributed but packaged at constant mean amounts in endosomes. Cells respond to higher EGF concentrations by increasing the number of endosomes but keeping the mean p-EGFR content per endosome almost constant. By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output. Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation. We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response.

Show MeSH
Related in: MedlinePlus