Peptide growth factors stimulate cellular responses through activation of their trans-membrane receptors. Multiple intracellular signaling cascades are engaged following growth factor - receptor binding, leading to short- and long-term biological effects. Each receptor-activated signaling pathway does not act in isolation, but rather interacts at different levels with other pathways to shape signaling networks that are distinctive for each growth factor. To gain insights into the specifics of growth factor-regulated interactions among different signaling cascades, we developed a HeLa cell line stably expressing fluorescent live-cell imaging reporters that are readouts for two major growth factor-stimulated pathways, Ras - Raf - Mek - Erk and PI3-kinase - Akt. Incubation of cells with EGF resulted in rapid, robust, and sustained Erk signaling but shorter-term activation of Akt. In contrast, HGF induced sustained Akt signaling, but weak and short-lived Erk activity, and IGF-I stimulated strong long-term Akt responses, but negligible Erk signaling. To address potential interactions between signaling pathways, we employed specific small molecule inhibitors. In cells incubated with EGF or PDGF-AA, Raf activation and the subsequent stimulation of Erk reduced Akt signaling, while Mek inhibition, which blocked Erk activation, enhanced Akt, and turned transient effects into sustained responses. Our results reveal that individual growth factors initiate signaling cascades that vary markedly in strength and duration, and demonstrate in living cells the dramatic effects of crosstalk from Raf and Mek to PI3-kinase and Akt. Our data further indicate how specific growth factors can encode distinct cellular behaviors by promoting complex interactions among signaling pathways.
- growth factors
- live-cell imaging
- signal transduction
- Copyright © 2017, American Journal of Physiology-Cell Physiology