Encoding Growth Factor Identity in the Temporal Dynamics of FOXO3 under the Combinatorial Control of ERK and AKT Kinases

Abstract

Extracellular growth factors signal to transcription factors via a limited number of cytoplasmic kinase cascades. It remains unclear how such cascades encode ligand identities and concentrations. In this paper, we use live-cell imaging and statistical modeling to study FOXO3, a transcription factor regulating diverse aspects of cellular physiology that is under combinatorial control. We show that FOXO3 nuclear-to-cytosolic translocation has two temporally distinct phases varying in magnitude with growth factor identity and cell type. These phases comprise synchronous translocation soon after ligand addition followed by an extended back-and-forth shuttling; this shuttling is pulsatile and does not have a characteristic frequency, unlike a simple oscillator. Early and late dynamics are differentially regulated by Akt and ERK and have low mutual information, potentially allowing the two phases to encode different information. In cancer cells in which ERK and Akt are dysregulated by oncogenic mutation, the diversity of states is lower.

Collective ERK/Akt activity waves orchestrate epithelial homeostasis by driving apoptosis-induced survival

“…S1C). As previously shown (Albeck et al 2013;Sampattavanich et al 2018), we observed that starved MCF10A cells show synchronous ERK and Akt activity pulses whose frequency increases with EGF stimulation (Fig.1A). The amplitudes of both ERK/Akt pulses were similar in presence or absence of EGF ( Fig.S1D).…”

Communication codes in developmental signaling pathways

“…As EGF and NGF elicit different cellular phenotypes, it would be interesting to investigate whether different RTKs engage similar or distinct downstream feedback loops, or whether the different dynamics result from different structural features of various ligand-receptor complexes. Combinatorial control of a cell response by AKT and ERK has also been observed in individual human mammary gland epithelial cells stimulated with various growth factors (Sampattavanich et al, 2018). The RTK signaling system thus provides an important and intriguing example of how signaling pathways not only transmit inputs but actively process them to control population structure, and how quantitative, multi-dimensional, single cell measurements can provide insights into the relationship between signal processing and cell fate decision-making.…”

Cracking the context-specific PI3K signaling code

“…For instance, a quantitative study of NGF signaling in PC12 cells revealed that the cellular decision to differentiate or proliferate is determined by a two-dimensional phospho-ERK/phospho-AKT response map that integrates the activation strength of both pathways (61). In addition, the early and late dynamics of FOXO3 nuclear-cytoplasmic shuttling is differentially regulated by AKT and ERK downstream of different growth factors, potentially serving as a mechanism to encode the identity of upstream ligands (62).…”

“…These include (i) the use of CRISPR-mediated tagging of endogenous effector proteins, such as AKT or FOXO, to follow their dynamic translocation live and without stoichiometric changes and (ii) the integration of quantitative, multiplexed immunofluorescence in time course studies that seek to assess a wider repertoire of signaling responses at the single-cell level (65)(66)(67). Proof-of-concept studies from Sorger and his team illustrate the detailed cell signaling insight that can be obtained with some of these approaches and the translational potential of the acquired knowledge (62,68). Adoption of these methodologies will likely be instrumental in closing the PI3K signaling knowledge gaps that will be discussed next.…”