Protein abundance of AKT and ERK pathway components governs cell type-specific regulation ofproliferation.

Lorenz Adlung,Sajib Chakraborty,Jens Timmer,Ursula Klingmüller,Thomas Höfer,Susen Lattermann,Melanie Boerries,Marie‐Christine Wagner,Marcel Schilling,Patrick Wuchter,Sandip Kar,Bin She,Hauke Busch,Jie Bao,Anthony D Ho

doi:10.15252/msb.20167258

Abstract

Signaling through the AKT and ERK pathways controls cell proliferation. However, the integrated regulation of this multistep process, involving signal processing, cell growth and cell cycle progression, is poorly understood. Here, we study different hematopoietic cell types, in which AKT and ERK signaling is triggered by erythropoietin (Epo). Although these cell types share the molecular network topology for pro‐proliferative Epo signaling, they exhibit distinct proliferative responses. Iterating quantitative experiments and mathematical modeling, we identify two molecular sources for cell type‐specific proliferation. First, cell type‐specific protein abundance patterns cause differential signal flow along the AKT and ERK pathways. Second, downstream regulators of both pathways have differential effects on proliferation, suggesting that protein synthesis is rate‐limiting for faster cycling cells while slower cell cycles are controlled at the G1‐S progression. The integrated mathematical model of Epo‐driven proliferation explains cell type‐specific effects of targeted AKT and ERK inhibitors and faithfully predicts, based on the protein abundance, anti‐proliferative effects of inhibitors in primary human erythroid progenitor cells. Our findings suggest that the effectiveness of targeted cancer therapy might become predictable from protein abundance.

Highlights

Eukaryotic cells use a limited number of signal transduction pathways to integrate information from extracellular stimuli and to regulate cellular decisions such as proliferation, differentiation, and apoptosis
To quantitatively assess Epo-induced proliferative responses in murine primary erythroid progenitor cells at the colony-forming unit-erythroid stage and in the immortalized murine cell line BaF3 exogenously expressing the Epo receptor (EpoR) (BaF3-EpoR), we incubated the cells in the presence of different Epo doses and measured as a readout for proliferation DNA synthesis by thymidine incorporation. mCFU-E cells showed a higher sensitivity toward Epo with an EC50 of 0.26 Æ 0.02 U/ml Epo as compared to 0.55 Æ 0.04 U/ml Epo for BaF3-EpoR cells (Fig 1A)
To study the expression and phosphorylation of the EpoR and AKT, mCFU-E cells were stimulated with 2.5 U/ml Epo, while BaF3-EpoR cells were stimulated with 5 U/ml Epo to account for the at least twofold difference in sensitivities toward Epo (Fig 1A) and the observation that responses in mCFU-E cells already saturated at 2.5 U/ml Epo, whereas BaF3-EpoR required more than 5 U/ml Epo (Appendix Fig S1)

Summary

Introduction

Eukaryotic cells use a limited number of signal transduction pathways to integrate information from extracellular stimuli and to regulate cellular decisions such as proliferation, differentiation, and apoptosis. The PI3K/AKT and Ras/MEK/ERK pathways have been implicated in the control of cell growth and proliferation in many different cell types (Saez-Rodriguez et al, 2015). Previous studies suggest that the regulation of the AKT and the ERK pathway crucially depends on the cellular context (McCubrey et al, 2011). Both pathways are activated by multiple growth factors and cytokines such as the hormone erythropoietin (Epo), which is the prime regulator of erythropoiesis. Epo is essential for survival, proliferation, and differentiation of erythroid progenitor cells and thereby facilitates continuous renewal of mature erythrocytes (Koury & Bondurant, 1990).

Methods

Results

Conclusion