Heterogeneous kinetics of AKT signaling in individual cells are accounted for by variable protein concentration

René Meyer,Bernhard Kramer,Sandip Kar,Daniel Kaschek,Bin She,Bettina Hahn,Mats Kvarnström,Ursula Klingmüller,Wolf-Dieter Lehmann,Mats Jirstrand,David Wrangborg,Johan Karlsson,Lorenza A D'Alessandro,Thomas Höfer,Jens Timmer

doi:10.3389/fphys.2012.00451

René Meyer, Bernhard Kramer + Show 13 more

Open Access

https://doi.org/10.3389/fphys.2012.00451

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Abstract

In most solid cancers, cells harboring oncogenic mutations represent only a sub-fraction of the entire population. Within this sub-fraction the expression level of mutated proteins can vary significantly due to cellular variability limiting the efficiency of targeted therapy. To address the causes of the heterogeneity, we performed a systematic analysis of one of the most frequently mutated pathways in cancer cells, the phosphatidylinositol 3 kinase (PI3K) signaling pathway. Among others PI3K signaling is activated by the hepatocyte growth factor (HGF) that regulates proliferation of hepatocytes during liver regeneration but also fosters tumor cell proliferation. HGF-mediated responses of PI3K signaling were monitored both at the single cell and cell population level in primary mouse hepatocytes and in the hepatoma cell line Hepa1_6. Interestingly, we observed that the HGF-mediated AKT responses at the level of individual cells is rather heterogeneous. However, the overall average behavior of the single cells strongly resembled the dynamics of AKT activation determined at the cell population level. To gain insights into the molecular cause for the observed heterogeneous behavior of individual cells, we employed dynamic mathematical modeling in a stochastic framework. Our analysis demonstrated that intrinsic noise was not sufficient to explain the observed kinetic behavior, but rather the importance of extrinsic noise has to be considered. Thus, distinct from gene expression in the examined signaling pathway fluctuations of the reaction rates has only a minor impact whereas variability in the concentration of the various signaling components even in a clonal cell population is a key determinant for the kinetic behavior.

Highlights

Cancer heterogeneity is considered a result of clonal instability, followed by clonal evolution (Campbell and Polyak, 2007; Marusyk and Polyak, 2010), as it has been shown in cultured cell lines (Odoux et al, 2008; Dalerba et al, 2011)
To determine the dynamics of hepatocyte growth factor (HGF) signaling at the cell population level, primary mouse hepatocytes were stimulated with HGF and lysed at different time points
The mCherry-AKT localization was monitored by live cell imaging in transiently transfected primary mouse hepatocytes stimulated with HGF or left untreated

Summary

Introduction

Cancer heterogeneity is considered a result of clonal instability, followed by clonal evolution (Campbell and Polyak, 2007; Marusyk and Polyak, 2010), as it has been shown in cultured cell lines (Odoux et al, 2008; Dalerba et al, 2011). The heterogeneity of individual cells in a tumor is an extremely important issue since it can cause differential responses to treatment resulting in incomplete tumor regression and contributing to overall poor efficiency of therapy in hepatocellular carcinoma (HCC) patients (Unsal et al, 1994; Shachaf et al, 2004) and other cancers (Brognard et al, 2001). Cells harbor non-genetic sources of random variability that are likely to contribute to heterogeneous responses to therapy. Cells die at very different time points after the administration of pro-apoptotic drugs, and a sizable fraction of cells usually survives treatment (Spencer et al, 2009). Such non-genetic cell-to-cell variability has been extensively studied in gene expression. Two alleles of the same gene in mammalian cells show random differences in transcription in the absence of allelic imprinting

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