The flexibility of MAPK cascade responses enables regulation of a vast array of cell fate decisions, but elucidating the mechanisms underlying this plasticity is difficult in endogenous signaling networks. We constructed insulated mammalian MAPK cascades in yeast to explore how intrinsic and extrinsic perturbations affect the flexibility of these synthetic signaling modules. Contrary to biphasic dependence on scaffold concentration, we observe monotonic decreases in signal strength as scaffold concentration increases. We find that augmenting the concentration of sequential kinases can enhance ultrasensitivity and lower the activation threshold. Further, integrating negative regulation and concentration variation can decouple ultrasensitivity and threshold from the strength of the response. Computational analyses show that cascading can generate ultrasensitivity and that natural cascades with different kinase concentrations are innately biased toward their distinct activation profiles. This work demonstrates that tunable signal processing is inherent to minimal MAPK modules and elucidates principles for rational design of synthetic signaling systems.
Bibliographical noteFunding Information:
This work was supported by the National Institutes of Health (NIH) through the NIH Director's Pioneer Award Program, grant number DP1 OD00364, and by the Ellison Medical Foundation and the Howard Hughes Medical Institute (to J.J.C.); and by the American Heart Association, grant number 0835132N, and startup funds from the University of Pennsylvania (to C.A.S.). We also thank Natalie Ahn, Lloyd Cantley, Gerald Fink, Steen Hansen, and David Levin for kindly providing cDNA.